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Background With the arrival of chikungunya (CHIKV) and zika (ZIKV) viruses in Mexico, there was a decrease in diagnosed dengue virus (DENV) cases. During the first years of cocirculation (2015–2017), the algorithms established by epidemiological surveillance systems and the installed capacity limited us to one diagnostic test per sample, so there was an underestimation of cases until September 2017, when a multiplex algorithm was implemented. Therefore, the objective of this study was determine the impact of the introduction of CHIKV and ZIKV on the incidence of diagnosed DENV in endemic areas of Mexico, when performing the rediagnosis, using the multiplex algorithm, in samples from the first three years of co-circulation of these arboviruses. Methodology and principal findings For this, 1038 samples received by the Central Laboratory of Epidemiology between 2015 and 2017 were selected for this work. Viruses were identified by multiplex RT-qPCR, and the χ2 test was used to compare categorical variables. With the new multiplex algorithm, we identified 2.4 times the rate of arbovirosis as originally reported, evidencing an underestimation of the incidence of the three viruses. Even so, significantly less dengue was observed than in previous years. The high incidence rates of chikungunya and Zika coincided with periods of dengue decline. The endemic channel showed that the cases caused by DENV rose again after the circulation of CHIKV and ZIKV decreased. In addition, 23 cases of coinfection were identified, with combinations between all viruses. Conclusions and significance The results obtained in this study show for the first time the real impact on the detected incidence of dengue after the introduction of CHIKV and ZIKV in Mexico, the degree of underestimation of these arboviruses in the country, as well as the co-infections between these viruses, whose importance clinical and epidemiological are still unknown., Author summary Mexico is considered an endemic country for DENV as the number of cases each year is high. With the arrival of the CHIKV and ZIKV, this number decreased considerably. This led us to suspect that this decrease was not coincidental and could be attributed to two factors: the lack of expertise on the part of health teams to identify the clinical etiological agent and/or other complex ecological factors. Therefore, we decided to reanalyze 1038 suspected cases of dengue, chikungunya and Zika samples using a test that could identify any of the three viruses in a single reaction. We discovered that there was indeed an underestimation of cases, mostly of chikungunya and Zika but, contrary to what we imagined, a lower degree of dengue. We were also able to detect cases of coinfection, which had never been reported in Mexico. Thus, the present study lays the foundations for a better preparation for the arrival of other arboviruses, showing the need for the development and implementation of multiplex diagnostic techniques, in addition to providing more precise epidemiological information on this historical moment of co-circulation of these three arboviruses in our country.

Introduction: Chikungunya virus (CHIKV) is a mosquito-borne virus known to cause acute febrile illness associated with debilitating polyarthritis. In 2019, several institutions in Myanmar reported a CHIKV outbreak. There are no official reports of CHIKV cases between 2011 and 2018. Therefore, this study sought to determine the seroprevalence of CHIKV infection before the 2019 outbreak. Methods: A total of 1,544 serum samples were collected from healthy volunteers and patients with febrile illnesses in Yangon, Mandalay, and the Myeik district in 2013, 2015, and 2018. Participants ranged from one month to 65 years of age. Antibody screening was performed with in-house anti-CHIKV IgG and IgM ELISA. A neutralization assay was used as a confirmatory test. Results: The seroprevalence of anti-CHIKV IgM and anti-CHIKV IgG was 8.9% and 28.6%, respectively, with an overall seropositivity rate of 34.5%. A focus reduction neutralization assay confirmed 32.5% seroprevalence of CHIKV in the study population. Age, health status, and region were significantly associated with neutralizing antibodies (NAbs) and CHIKV seropositivity (p < 0.05), while gender was not (p = 0.9). Seroprevalence in 2013, 2015, and 2018 was 32.1%, 28.8%, and 37.3%, respectively. Of the clinical symptoms observed in participants with fevers, arthralgia was mainly noted in CHIKV-seropositive patients. Conclusion: The findings in this study reveal the circulation of CHIKV in Myanmar’s Mandalay, Yangon, and Myeik regions before the 2019 CHIKV outbreak. As no treatment or vaccine for CHIKV exists, the virus must be monitored through systematic surveillance in Myanmar., PLoS Neglected Tropical Diseases, 15(12), art. no. e0009961; 2021

Tomatidine, a natural steroidal alkaloid from unripe green tomatoes has been shown to exhibit many health benefits. We recently provided in vitro evidence that tomatidine reduces the infectivity of Dengue virus (DENV) and Chikungunya virus (CHIKV), two medically important arthropod-borne human infections for which no treatment options are available. We observed a potent antiviral effect with EC50 values of 0.82 μM for DENV-2 and 1.3 μM for CHIKV-LR. In this study, we investigated how tomatidine controls CHIKV infectivity. Using mass spectrometry, we identified that tomatidine induces the expression of p62, CD98, metallothionein and thioredoxin-related transmembrane protein 2 in Huh7 cells. The hits p62 and CD98 were validated, yet subsequent analysis revealed that they are not responsible for the observed antiviral effect. In parallel, we sought to identify at which step of the virus replication cycle tomatidine controls virus infectivity. A strong antiviral effect was seen when in vitro transcribed CHIKV RNA was transfected into Huh7 cells treated with tomatidine, thereby excluding a role for tomatidine during CHIKV cell entry. Subsequent determination of the number of intracellular viral RNA copies and viral protein expression levels during natural infection revealed that tomatidine reduces the RNA copy number and viral protein expression levels in infected cells. Once cells are infected, tomatidine is not able to interfere with active RNA replication yet it can reduce viral protein expression. Collectively, the results delineate that tomatidine controls viral protein expression to exert its antiviral activity. Lastly, sequential passaging of CHIKV in presence of tomatidine did not lead to viral resistance. Collectively, these results further emphasize the potential of tomatidine as an antiviral treatment towards CHIKV infection., Author summary Chikungunya fever is a debilitating disease caused by the mosquito-borne Chikungunya virus. Over the past two decades the geographical spread of the virus and its mosquito vector has drastically increased thereby causing millions of infections. To date there is no antiviral drug and no vaccine available to treat/prevent Chikungunya virus infection. We recently showed that the natural steroidal alkaloid tomatidine has potent antiviral activity towards Chikungunya virus at submicromolar concentrations. In this study we dissected how tomatidine reduces the production of Chikungunya virus particles. We show that tomatidine lowers viral protein expression and we hypothesize that the effect of tomatidine on viral protein translation hampers the production of progeny viral RNA copies / number of infected cells thereby leading to a reduced production of secreted virus particles. Also, we show that Chikungunya virus does not readily become resistant to tomatidine. Collectively, we deciphered the mechanism by which tomatidine exerts antiviral activity to Chikungunya virus and our results strengthen the potential of tomatidine as an antiviral treatment strategy towards Chikungunya virus.

Chikungunya virus (CHIKV) epidemics around the world have created public health concern with the unavailability of effective drugs and vaccines. This emphasizes the need for molecular understanding of host-virus interactions for developing effective targeted antivirals. Microarray analysis was carried out using CHIKV strain (Prototype and Indian) infected Vero cells and two host isozymes, MAPK activated protein kinase 2 (MK2) and MAPK activated protein kinase 3 (MK3) were selected for further analysis. The substrate spectrum of both enzymes is indistinguishable and covers proteins involved in cytokines production, endocytosis, reorganization of the cytoskeleton, cell migration, cell cycle control, chromatin remodeling and transcriptional regulation. Gene silencing and drug treatment were performed in vitro and in vivo to unravel the role of MK2/MK3 in CHIKV infection. Gene silencing of MK2 and MK3 abrogated around 58% CHIKV progeny release from the host cell and a MK2 activation inhibitor (CMPD1) treatment demonstrated 68% inhibition of viral infection suggesting a major role of MAPKAPKs during late CHIKV infection in vitro. Further, it was observed that the inhibition in viral infection is primarily due to the abrogation of lamellipodium formation through modulation of factors involved in the actin cytoskeleton remodeling pathway. Moreover, CHIKV-infected C57BL/6 mice demonstrated reduction in the viral copy number, lessened disease score and better survivability after CMPD1 treatment. In addition, reduction in expression of key pro-inflammatory mediators such as CXCL13, RAGE, FGF, MMP9 and increase in HGF (a CHIKV infection recovery marker) was observed indicating the effectiveness of the drug against CHIKV. Taken together it can be proposed that MK2 and MK3 are crucial host factors for CHIKV infection and can be considered as important target for developing effective anti-CHIKV strategies., Author summary Chikungunya virus has been a dreaded disease from the first time it occurred in 1952 Tanzania. Since then it has been affecting the different parts of the world at different time periods in large scale. It is typically transmitted to humans by bites of Aedes aegypti and Aedes albopictus mosquitoes. Although, studies have been undertaken to combat its prevalence still there are no effective strategies like vaccines or antivirals against it. Therefore it is essential to understand the virus and host interaction to overcome this hurdle. In this study two host factors MK2 and MK3 have been taken into consideration to see how they affect the multiplication of the virus. The in vitro and in vivo experiments conducted demonstrated that inhibition of MK2 and MK3 not only restricted viral release but also decreased the disease score and allowed better survivability. Therefore, MK2 and MK3 could be considered as the key targets in the anti CHIKV approach.

Background Australia is theoretically at risk of epidemic chikungunya virus (CHIKV) activity as the principal vectors are present on the mainland Aedes aegypti) and some islands of the Torres Strait (Ae. aegypti and Ae. albopictus). Both vectors are highly invasive and adapted to urban environments with a capacity to expand their distributions into south-east Queensland and other states in Australia. We sought to estimate the epidemic potential of CHIKV, which is not currently endemic in Australia, by considering exclusively transmission by the established vector in Australia, Ae. aegypti, due to the historical relevance and anthropophilic nature of the vector. Methodology/Principal findings We estimated the historical (1995–2019) epidemic potential of CHIKV in eleven Australian locations, including the Torres Strait, using a basic reproduction number equation. We found that the main urban centres of Northern Australia could sustain an epidemic of CHIKV. We then estimated future trends in epidemic potential for the main centres for the years 2020 to 2029. We also conducted uncertainty and sensitivity analyses on the variables comprising the basic reproduction number and found high sensitivity to mosquito population size, human population size, impact of vector control and human infectious period. Conclusions/Significance By estimating the epidemic potential for CHIKV transmission on mainland Australia and the Torres Strait, we identified key areas of focus for controlling vector populations and reducing human exposure. As the epidemic potential of the virus is estimated to rise towards 2029, a greater focus on control and prevention measures should be implemented in at-risk locations., Author summary Chikungunya virus (CHIKV) is transmitted primarily by Aedes aegypti and Aedes albopictus mosquitoes and causes a potentially debilitating febrile and arthralgic disease. The virus is a threat to public health in regions where the primary vectors are established, as evidenced by past epidemics in the Indian Ocean Islands, South America and the Caribbean. In Australia, there are established populations of Ae. aegypti both on the mainland and in the Torres Strait, and of Ae. albopictus in the Torres Strait. This provides a theoretical potential for CHIKV transmission, as seen historically with dengue virus (DENV). It is therefore important to understand the epidemic potential of CHIKV in Australia. We estimated the basic reproduction number (R0) of CHIKV during the years 1995–2019 for 11 Urban Centres and Localities (UCLs) in Australia, and found that Brisbane, Cairns, Darwin, Rockhampton, Thursday Island, and Townsville were all susceptible to CHIKV epidemics. We then forecasted epidemic potential from 2020–2029 and found an increase in R0 across the six main UCLs. By highlighting factors that significantly influence the epidemic potential of CHIKV in Australia, our study supports evidence-based decision making for vector control and public health programs.

The zinc finger antiviral protein (ZAP) is a broad inhibitor of virus replication. Its best-characterized function is to bind CpG dinucleotides present in viral RNAs and, through the recruitment of TRIM25, KHNYN and other cofactors, target them for degradation or prevent their translation. The long and short isoforms of ZAP (ZAP-L and ZAP-S) have different intracellular localization and it is unclear how this regulates their antiviral activity against viruses with different sites of replication. Using ZAP-sensitive and ZAP-insensitive human immunodeficiency virus type I (HIV-1), which transcribe the viral RNA in the nucleus and assemble virions at the plasma membrane, we show that the catalytically inactive poly-ADP-ribose polymerase (PARP) domain in ZAP-L is essential for CpG-specific viral restriction. Mutation of a crucial cysteine in the C-terminal CaaX box that mediates S-farnesylation and, to a lesser extent, the residues in place of the catalytic site triad within the PARP domain, disrupted the activity of ZAP-L. Addition of the CaaX box to ZAP-S partly restored antiviral activity, explaining why ZAP-S lacks antiviral activity for CpG-enriched HIV-1 despite conservation of the RNA-binding domain. Confocal microscopy confirmed the CaaX motif mediated localization of ZAP-L to vesicular structures and enhanced physical association with intracellular membranes. Importantly, the PARP domain and CaaX box together jointly modulate the interaction between ZAP-L and its cofactors TRIM25 and KHNYN, implying that its proper subcellular localisation is required to establish an antiviral complex. The essential contribution of the PARP domain and CaaX box to ZAP-L antiviral activity was further confirmed by inhibition of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) replication, which replicates in double-membrane vesicles derived from the endoplasmic reticulum. Thus, compartmentalization of ZAP-L on intracellular membranes provides an essential effector function in ZAP-L-mediated antiviral activity against divergent viruses with different subcellular replication sites., Author summary Cell-intrinsic antiviral factors, such as the zinc finger antiviral protein (ZAP), provide the first line of defence against viral pathogens. ZAP acts by selectively binding CpG dinucleotides in viral RNAs, leading to their degradation or translation inhibition. Here, we show that the ability to target these foreign elements is not only dependent on ZAP’s N-terminal RNA-binding domain, but additional determinants in the central and C-terminal regions also regulate this process. The PARP domain and its associated CaaX box, are crucial for ZAP’s CpG-specific activity and required for optimal binding to the ZAP cofactors TRIM25 and KHNYN. Furthermore, a CaaX box, known to mediate post-translational modification by a hydrophobic S-farnesyl group, caused re-localization of ZAP from the cytoplasm and increased its association with intracellular membranes. The distribution of the long isoform of ZAP to intracellular membranes was essential for inhibition of both a ZAP-sensitized HIV-1 and SARS-CoV-2. Our work unveils how the determinants outside the RNA-binding domain assist ZAP’s antiviral activity and highlights the role of S-farnesylation and membrane association in this process.

Background Biological control programs involving Wolbachia-infected Aedes aegypti are currently deployed in different epidemiological settings. New Caledonia (NC) is an ideal location for the implementation and evaluation of such a strategy as the only proven vector for dengue virus (DENV) is Ae. aegypti and dengue outbreaks frequency and severity are increasing. We report the generation of a NC Wolbachia-infected Ae. aegypti strain and the results of experiments to assess the vector competence and fitness of this strain for future implementation as a disease control strategy in Noumea, NC. Methods/principal findings The NC Wolbachia strain (NC-wMel) was obtained by backcrossing Australian AUS-wMel females with New Caledonian Wild-Type (NC-WT) males. Blocking of DENV, chikungunya (CHIKV), and Zika (ZIKV) viruses were evaluated via mosquito oral feeding experiments and intrathoracic DENV challenge. Significant reduction in infection rates were observed for NC-wMel Ae. aegypti compared to WT Ae. aegypti. No transmission was observed for NC-wMel Ae. aegypti. Maternal transmission, cytoplasmic incompatibility, fertility, fecundity, wing length, and insecticide resistance were also assessed in laboratory experiments. Ae. aegypti NC-wMel showed complete cytoplasmic incompatibility and a strong maternal transmission. Ae. aegypti NC-wMel fitness seemed to be reduced compared to NC-WT Ae. aegypti and AUS-wMel Ae. aegypti regarding fertility and fecundity. However further experiments are required to assess it accurately. Conclusions/significance Our results demonstrated that the NC-wMel Ae. aegypti strain is a strong inhibitor of DENV, CHIKV, and ZIKV infection and prevents transmission of infectious viral particles in mosquito saliva. Furthermore, our NC-wMel Ae. aegypti strain induces reproductive cytoplasmic incompatibility with minimal apparent fitness costs and high maternal transmission, supporting field-releases in Noumea, NC., Author summary Dengue represents a risk for almost half of the world’s population, especially throughout the tropics. In New Caledonia, dengue outbreaks have become more frequent in the past decade along with the recent circulation of chikungunya and Zika viruses. The opportunity to use the biocontrol method involving the release of Wolbachia-infected Ae. aegypti mosquitoes has been investigated as an alternative solution to the traditional control methods, like elimination of larval habitats and pyrethroid insecticide application to kill adults, which are becoming insufficient. A local strain of Ae. aegypti carrying Wolbachia (NC-wMel) has been generated and tested to evaluate its pathogen blocking capacity for the four dengue virus serotypes as well as chikungunya and Zika viruses. The fitness of NC-wMel strain has also been assessed to estimate its ability to compete with the wild-type strain in the field. Noumea city, where a third of the population of New Caledonia resides, has been chosen as the first site to implement the method in New Caledonia. As Ae. aegypti is the only proven vector in New Caledonia, we expect a significant impact on dengue outbreaks occurring in Noumea as soon as a high frequency of NC-wMel is established in the population.

Dengue has become endemic in Pakistan with annual recurrence. A sudden increase in the dengue cases was reported from Rawalpindi in 2016, while an outbreak occurred for the first time in Peshawar in 2017. Therefore, a multi-center study was carried out to determine the circulating dengue virus (DENV) serotypes and Chikungunya virus (CHIKV) co-infection in Lahore, Rawalpindi, and Peshawar cities in 2016–18. A hospital-based cross-sectional study was carried out in Lahore and Rawalpindi in 2016–18, while a community-based study was carried out in Peshawar in 2017. The study participants were tested for dengue NS1 antigen using an immunochromatographic device while anti-dengue IgM/IgG antibodies were detected by indirect ELISA. All NS1 positive samples were used for DENV serotyping using multiplex real-time PCR assay. Additionally, dengue samples were tested for CHIKV co-infection using IgM/IgG ELISA. A total of 6291 samples were collected among which 8.11% were NS1 positive while 2.5% were PCR positive. DENV-2 was the most common serotype (75.5%) detected, followed by DENV-1 in 16.1%, DENV-3 in 3.9% and DENV-4 in 0.7% while DENV-1 and DENV-4 concurrent infections were detected in 3.9% samples. DENV-1 was the predominant serotype (62.5%) detected from Lahore and Rawalpindi, while DENV-2 was the only serotype detected from Peshawar. Comorbidities resulted in a significant increase (p-value, Author summary Mosquitoes are responsible for the transmission of many different types of infectious pathogens and parasites to humans. Some of these pathogens include viruses like dengue and chikungunya viruses which cause diseases in humans. Dengue virus has four different types (1, 2, 3, and 4) present in nature. In this study, we detected all four dengue virus types in hospitalized patients in Lahore and Rawalpindi. While only dengue virus type-2 was detected from Peshawar city. The secondary infections with different dengue virus types than the previous one might result in a more severe form of the disease. Therefore, the population of Peshawar is at risk of developing a severe form of dengue fever in the future in case of infection from another dengue virus type. The data of hospitalized patients showed that other diseases in addition to dengue fever result in an increased hospital stay. Comorbidities like diabetes also increase the risk of developing a more severe form of dengue fever. Combined infections of both dengue and chikungunya viruses were detected from Lahore, Rawalpindi, and Peshawar. The transmission of dengue and chikungunya viruses and the co-circulation of multiple dengue virus types not only impose public health burden but also result in economic losses.

Zika virus (ZIKV) and chikungunya virus (CHIKV) were recently introduced into the Americas resulting in significant disease burdens. Understanding their spatial and temporal dynamics at the subnational level is key to informing surveillance and preparedness for future epidemics. We analyzed anonymized line list data on approximately 105,000 Zika virus disease and 412,000 chikungunya fever suspected and laboratory-confirmed cases during the 2014–2017 epidemics. We first determined the week of invasion in each city. Out of 1,122, 288 cities met criteria for epidemic invasion by ZIKV and 338 cities by CHIKV. We analyzed risk factors for invasion using linear and logistic regression models. We also estimated that the geographic origin of both epidemics was located in Barranquilla, north Colombia. We assessed the spatial and temporal invasion dynamics of both viruses to analyze transmission between cities using a suite of (i) gravity models, (ii) Stouffer’s rank models, and (iii) radiation models with two types of distance metrics, geographic distance and travel time between cities. Invasion risk was best captured by a gravity model when accounting for geographic distance and intermediate levels of density dependence; Stouffer’s rank model with geographic distance performed similarly well. Although a few long-distance invasion events occurred at the beginning of the epidemics, an estimated distance power of 1.7 (95% CrI: 1.5–2.0) from the gravity models suggests that spatial spread was primarily driven by short-distance transmission. Similarities between the epidemics were highlighted by jointly fitted models, which were preferred over individual models when the transmission intensity was allowed to vary across arboviruses. However, ZIKV spread considerably faster than CHIKV., Author summary Understanding the spread of infectious diseases across space and time is critical for preparedness, designing interventions, and elucidating mechanisms underlying transmission. We analyzed human case data from over 500,000 reported cases to investigate the spread of the recent Zika virus (ZIKV) and chikungunya virus (CHIKV) epidemics in Colombia. Both viruses were introduced into northern Colombia. We found that gravity models and Stouffer’s rank models best described transmission and that transmission mainly occurred over short distances. Our results highlight similarities and key differences between the ZIKV and CHIKV epidemics in Colombia, which can be used to anticipate future epidemic waves and prioritize cities for active surveillance and targeted interventions.

Mosquito-borne viruses cause diseases of great public health concern. Arboviral disease case distributions have complex relationships with socioeconomic and environmental factors. We combined information about socio-economic (population, and poverty rate) and environmental (precipitation, and land use) characteristics with reported human cases of arboviral disease in the counties of Alabama, USA, from 2007–2017. We used county level data on West Nile virus (WNV), dengue virus (DENV), chikungunya virus (CHIKV), Zika virus (ZIKV), California serogroup virus, Eastern equine encephalitis virus, and Saint Louis encephalitis virus to provide a detailed description of their spatio-temporal pattern. We found a significant spatial convergence between incidence of WNV and poverty rate clustered in the southern part of Alabama. DENV, CHIKV and ZIKV cases showed a different spatial pattern, being mostly located in the northern part, in areas of high socioeconomic status. The results of our study establish that poverty-driven inequities in arboviral risk exist in the southern USA, and should be taken into account when planning prevention and intervention strategies., Author summary Mosquito-borne arboviruses like West Nile virus (WNV), dengue virus (DENV), chikungunya virus (CHIKV), Zika virus (ZIKV), California serogroup virus (CSV), Eastern equine encephalitis virus (EEE), and Saint Louis encephalitis virus (SLE) are on the rise globally. Socioeconomic and environmental conditions have played a role in directing in this expansion by creating conditions ideal for mosquito vectors and transmission. In this study, we used 10 years (2007–2017) of county level human arboviral case data from the US state of Alabama to better understand the roles socioeconomics (poverty rate) and environmental (land use, precipitation, land cover) conditions may play in driving patterns of arboviral disease in the southern US. We found a significant association between poverty rate and incidence of WNV, an arbovirus primarily transmitted by Culex spp. mosquitoes, which are known for thriving in contaminated water sources and sewage overflow. Conversely, cases of DENV, CHIKV, and ZIKV, arboviruses primarily transmitted by Aedes spp. mosquitoes, were reported in areas of high socioeconomic status. These findings suggest differential distribution of arboviruses relevant to human health in Alabama, and that poverty in the southern US is a significant factor that should be considered when planning WNV prevention and intervention strategies.

Most alphaviruses are mosquito-borne and can cause severe disease in humans and domesticated animals. In North America, eastern equine encephalitis virus (EEEV) is an important human pathogen with case fatality rates of 30–90%. Currently, there are no therapeutics or vaccines to treat and/or prevent human infection. One critical impediment in countermeasure development is the lack of insight into clinically relevant parameters in a susceptible animal model. This study examined the disease course of EEEV in a cynomolgus macaque model utilizing advanced telemetry technology to continuously and simultaneously measure temperature, respiration, activity, heart rate, blood pressure, electrocardiogram (ECG), and electroencephalography (EEG) following an aerosol challenge at 7.0 log10 PFU. Following challenge, all parameters were rapidly and substantially altered with peak alterations from baseline ranged as follows: temperature (+3.0–4.2°C), respiration rate (+56–128%), activity (-15-76% daytime and +5–22% nighttime), heart rate (+67–190%), systolic (+44–67%) and diastolic blood pressure (+45–80%). Cardiac abnormalities comprised of alterations in QRS and PR duration, QTc Bazett, T wave morphology, amplitude of the QRS complex, and sinoatrial arrest. An unexpected finding of the study was the first documented evidence of a critical cardiac event as an immediate cause of euthanasia in one NHP. All brain waves were rapidly (~12–24 hpi) and profoundly altered with increases of up to 6,800% and severe diffuse slowing of all waves with decreases of ~99%. Lastly, all NHPs exhibited disruption of the circadian rhythm, sleep, and food/fluid intake. Accordingly, all NHPs met the euthanasia criteria by ~106–140 hpi. This is the first of its kind study utilizing state of the art telemetry to investigate multiple clinical parameters relevant to human EEEV infection in a susceptible cynomolgus macaque model. The study provides critical insights into EEEV pathogenesis and the parameters identified will improve animal model development to facilitate rapid evaluation of vaccines and therapeutics., Author summary In North America, EEEV causes the most severe mosquito-borne disease in humans highlighted by fatal encephalitis and permeant debilitating neurological sequelae in survivors. The first confirmed human cases were reported more than 80 years ago and since then multiple sporadic outbreaks have occurred including one of the largest in 2019. Unfortunately, most human infections are diagnosed at the on-set of severe neurological symptoms and consequently a detailed disease course in humans is lacking. This gap in knowledge is a significant obstacle in the development of appropriate animal models to evaluate countermeasures. Here, we performed a cutting-edge study by utilizing a new telemetry technology to understand the course of EEEV infection in a susceptible macaque model by measuring multiple physiological parameters relevant to human disease. Our study demonstrates that the infection rapidly produces considerable alterations in many critical parameters including the electrical activity of the heart and the brain leading to severe disease. The study also highlights the extraordinary potential of new telemetry technology to develop the next generation of animal models to comprehensively investigate pathogenesis as well as evaluate countermeasures to treat and/or prevent EEEV disease.

We report the identification of two orthobunyaviruses, Melao virus (MELV) and Oropouche virus (OROV), in plasma specimens from Haitian children with acute febrile illness who presented during outbreaks caused by alpha- and flaviviruses in 2014. Heretofore not described as a human pathogen, MELV was isolated in cell culture from the plasma of five case patients. OROV RNA was detected in the plasma of an additional child, using an unbiased sequencing approach, with phylogenetic inference suggesting a close relationship with strains from Brazil. Abdominal pain was reported by four case patients with MELV infections, with lymphadenopathy noted in two cases. Our findings document the occurrence of these orthobunyaviruses within the Caribbean region and highlight the critical importance of surveillance with viral genome sequence analyses to identify outbreaks caused by these and other emerging viruses., Author summary Melao and Oropuche virus infections were detected in Haitian children who developed acute febrile illnesses in year 2014. As these viruses were not previously known to circulate in Haiti, our findings highlight the critical importance of surveillance to identify outbreaks caused by these and other emerging viruses.

Background Chikungunya fever (CHIKF) is a serious public health problem with a high rate of infection and chronic disabling manifestations that has affected more than 2 million people worldwide since 2005. In spite of this, epidemiological data on vulnerable groups such as Indigenous people are scarce, making it difficult to implement public policies in order to prevent this disease and assist these populations. Objective To describe the serological and epidemiological profile of chikungunya virus (CHIKV) in two Indigenous populations in Northeast Brazil, as well as in an urbanized control community, and to explore associations between CHIKV and anthropometric variables in these populations. Methodology/Principal findings This is a cross-sectional ancillary study of the Project of Atherosclerosis among Indigenous Populations (PAI) that included people 30 to 70 years old, recruited from two Indigenous tribes (the less urbanized Fulni-ô and the more urbanized Truká people) and an urbanized non-Indigenous control group from the same area. Subjects underwent clinical evaluation and were tested for anti-CHIKV IgG by enzyme-linked immunosorbent assay. Serological profile was described according to ethnicity, sex, and age. The study population included 433 individuals distributed as follows: 109 (25·2%) Truká, 272 (62·8%) Fulni-ô, and 52 (12%) from the non-Indigenous urbanized control group. Overall prevalence of CHIKV IgG in the study sample was 49.9% (216; 95% CI: 45·1–54·7). When the sample was stratified, positive CHIKV IgG was distributed as follows: no individuals in the Truká group, 78·3% (213/272; 95% CI: 72·9–83·1) in the Fulni-ô group, and 5.8% (3/52; 95% CI: 1.21–16) in the control group. Conclusions/Significance Positive tests for CHIKV showed a very high prevalence in a traditional Indigenous population, in contrast to the absence of anti-CHIKV serology in the Truká people, who are more urbanized with respect to physical landscape, socio-cultural, and historical aspects, as well as a low prevalence in the non-Indigenous control group, although all groups are located in the same area., Author summary Chikungunya fever is a serious public health problem, with a high rate of infection and disease. Chikungunya virus (CHIKV) is a cosmopolitan virus, which has inflicted severe damage in 50 countries in the Americas and is responsible for chronic disabling manifestations. In spite of this, epidemiological data on vulnerable groups such as Indigenous people are scarce. We report on a cross-sectional study describing the seroprevalence of CHIKV in Indigenous groups in the São Francisco Valley, Brazil, in association with anthropometric data. The study population included 433 individuals distributed in the following ethnic groups: 109 (25.2%) Truká, 272 (62·8%) Fulni-ô, and 52 (12%) from the non-Indigenous urbanized control group When the sample was stratified, positive CHIKV IgG was distributed as follows: no individuals in the Truká group, 213/272 (78.3%; 95% CI: 72·9–83·1) individuals in the Fulni-ô group, and 3/52 (5.8%; 95% CI: 1·21–16) individuals in the control group. This study shows, for the first time, that CHIKV circulated in an Indigenous population (Fulni-ô) in the São Francisco Valley, in 2016 and 2017. The finding strikingly differs from the absence of anti-CHIKV serology found in the Truká people and from the low prevalence in the urban region of Juazeiro, Bahia.

The rapid evolution of RNA viruses has been long considered to result from a combination of high copying error frequencies during RNA replication, short generation times and the consequent extensive fixation of neutral or adaptive changes over short periods. While both the identities and sites of mutations are typically modelled as being random, recent investigations of sequence diversity of SARS coronavirus 2 (SARS-CoV-2) have identified a preponderance of C->U transitions, proposed to be driven by an APOBEC-like RNA editing process. The current study investigated whether this phenomenon could be observed in datasets of other RNA viruses. Using a 5% divergence filter to infer directionality, 18 from 36 datasets of aligned coding region sequences from a diverse range of mammalian RNA viruses (including Picornaviridae, Flaviviridae, Matonaviridae, Caliciviridae and Coronaviridae) showed a >2-fold base composition normalised excess of C->U transitions compared to U->C (range 2.1x–7.5x), with a consistently observed favoured 5’ U upstream context. The presence of genome scale RNA secondary structure (GORS) was the only other genomic or structural parameter significantly associated with C->U/U->C transition asymmetries by multivariable analysis (ANOVA), potentially reflecting RNA structure dependence of sites targeted for C->U mutations. Using the association index metric, C->U changes were specifically over-represented at phylogenetically uninformative sites, potentially paralleling extensive homoplasy of this transition reported in SARS-CoV-2. Although mechanisms remain to be functionally characterised, excess C->U substitutions accounted for 11–14% of standing sequence variability of structured viruses and may therefore represent a potent driver of their sequence diversification and longer-term evolution., Author summary The rapid evolution of RNA viruses is thought to arise from high mutation frequencies during replication and the rapid accumulation of genetic changes over time in response to its changing environments. This study describes an additional potent factor that contributes to the evolution of RNA viruses infecting mammals, the occurrence of specific C->U mutations in a subset that possess intensely structured genomes. This mutational process substantially damages the virus’s ability to replicate and is potentially akin to “genome editing” of HIV-1 and other retrovirus genome sequences by APOBEC, one of the principal components of vertebrate antiretroviral defence mechanisms. This study however provides evidence for a wider mutational activity against several human and veterinary RNA viruses, including HCV and foot and mouth disease virus. The C->U mutational process accounted for 15–20% of standing sequence variability of these RNA viruses, representing a potent driver of their sequence diversification and longer-term evolution.

Background Chikungunya fever is considered an abrupt onset arbovirus transmitted by mosquitoes, mainly Aedes aegypti and Aedes albopictus. The disease has a significant impact on the quality of life of affected persons, and many of its numerous symptoms have not yet been properly clarified, such as the manifestations that can occur in the oral cavity. The aim of this study was to identify the main oral manifestations related to chikungunya fever, as well as describe the demographic characteristics of patients, by conducting a systematic review of the literature. Methods and findings Searches were performed in MEDLINE (PubMed), Embase (Elsevier), LILACS (VHL), Cochrane Library, Scopus, and CAPES electronic databases for theses and dissertations published up to January 16, 2021 without language and date restrictions. Additional manual searches of gray literature, reference list, and Google Scholar were carried out. We included 27 studies highlighting mainly oral manifestations that cause masticatory discomfort such as ulcers and oral thrush, gingival bleeding, pain and burning of the oral mucous membranes, temporomandibular joint (TMJ) arthralgia, opportunistic infections, and changes in taste. Conclusions There seems to be a predominance of oral manifestations that cause discomfort when chewing, such as ulcerations in the acute phase of the disease, with complete remission within 3 to 10 days after the onset, apparently mostly affecting women and older persons. These oral manifestations can be compatible with basic viral infections related to inflammatory response and transitory immunosuppression., Author summary Since its resurgence in 2004, control of the transmission of chikungunya fever has become a challenge to public health worldwide. As it is capable of diminishing the quality of life of infected persons, it has aroused interest in the scientific community in developing more in-depth studies. Many of its numerous symptoms have not yet been properly clarified, such as the manifestations that can occur in the oral cavity. A study conducted in India in 2010, with the aim of verifying the prevalence of oral manifestations in chikungunya patients, found a positive outcome in 96.37% of the 110 suspected cases and 100% of the 37 confirmed cases. Given the above findings, our aim in this study was to carry out a systematic review on the subject of oral manifestations in patients with chikungunya. Based on the 27 studies selected, we were able to point out the main oral manifestations that cause masticatory discomfort such as ulcers and oral thrush, gingival bleeding, pain and burning of the oral mucous membranes, temporomandibular joint (TMJ) arthralgia, opportunistic infections, and changes in taste. Studies have reported that both frequency and worsening of oral mucosal lesions occur with increasing age and higher predominance in females. However, many studies have pointed out that most oral manifestations such as ulcerations were more frequently observed in the acute phase of the disease, with complete remission occurring 3 to 10 days after the onset of the symptoms.

Background The Old world Alphavirus, Middelburg virus (MIDV), is not well known and although a few cases associated with animal illness have previously been described from Southern Africa, there has been no investigation into the association of the virus with human illness. The current study aimed to investigate possible association of MIDV infection with febrile or neurological manifestations in hospitalized or symptomatic patients fromGauteng, South Africa. Methods This study is a descriptive retrospective and prospective laboratory based study. Archived cerebrospinal fluid (CSF) samples submitted to the National Health Laboratory Service (NHLS), Tshwane Academic division for viral investigation from public sector hospitals in Gauteng as well as EDTA (ethylenediaminetetraacetic acid) whole blood samples from ad hoc cases of veterinary students, presenting with neurological and febrile illness, were selected and screened for the presence of alphaviruses using real-time reverse transcription(rtRT) PCR.Virus isolations from rtRT-PCR positive samples were conducted in Vero cell culture and used to obtain full genome sequences. Basic descriptive statistical analysis was conducted using EpiInfo. Results MIDV was detected by rtRT-PCR in 3/187 retrospective CSF specimens obtained from the NHLS from hospitalised patients in the Tshwane region of Gauteng and 1/2 EDTA samples submitted in the same year (2017) from ad hoc query arbovirus cases from veterinary students from the Faculty of Veterinary Science University of Pretoria.Full genome sequences were obtained for virus isolates from two cases; one from an EDTA whole blood sample (ad hoc case) and another from a CSF sample (NHLS sample).Two of the four Middelburg virus positive cases,for which clinical information was available, had other comorbidities or infections at the time of infection. Conclusion Detection of MIDV in CSF of patients with neurological manifestations suggests that the virus should be investigated as a human pathogen with the potential of causing or contributing to neurological signs in children and adults., Author summary Old World alphaviruses are mainly associated with polyarthritis in humans. Global emergence of chikungunya virus raises concerns regarding disease potential of neglected African arboviruses.New World alphaviruses are associated with severe neurological disease in humans and horses in the Americas with a high mortality rate. Middelburg virus (MIDV) is an Old World alphavirus previously associated with neurological disease and fatalities in horses and wildlife in southern Africa but not yet linked to disease in humans. This study investigated MIDV in humans presenting with acute neurological symptoms in South Africa. MIDV infection was confirmed in 4/189 (2%) such cases using a generic Alphavirus real-time RT-PCR and phylogenetic analysis. MIDV full genomes were obtained from a cerebrospinal fluid sample of a 2-year-old child and a whole blood sample of a veterinary student, both presenting with neurological signs. This study suggests MIDV may be associated with unsolved neurological disease in humans in Africa.

Background Aedes (Stegomyia) albopictus (Skuse) impacts human outdoor activity because of its aggressive biting behavior, and as a major vector of mosquito-borne diseases, it is also of public health importance. Although most mosquito species exhibit crepuscular activity by primarily host seeking at dawn and dusk, Ae. albopictus has been traditionally characterized as a diurnal or day-biting mosquito. With the global expansion and increased involvement of Ae. albopictus in mosquito-borne diseases, it is imperative to elucidate the diel activity of this species, particularly in newly invaded areas. Methodology and principal findings Human sweep netting and carbon dioxide-baited rotator traps were used to evaluate the diel activity of Ae. albopictus in two study sites. Both trapping methods were used in New Jersey’s Mercer County, USA (temperate/urban), while only human sweep netting was used in Florida’s Volusia County, USA (subtropical/suburban). Human sweep netting was performed to determine adult mosquito activity at Sunrise, Solar Noon, Sunset, and Lunar Midnight. Because New Jersey is in a temperate area, diel activity was investigated during the early season (3–19 July), peak season (25 July-19 September), and late season (22 September- 22 October). Aedes albopictus showed the highest activity during peak and late seasons at Solar Noon (P < 0.05). At Sunrise and Sunset during the peak season, Ae. albopictus activity was similar. Lunar Midnight activity was significantly lower than Sunrise and Solar Noon (P < 0.05) but was similar to that of Sunset. In the late season, the highest activity was observed during Solar Noon while the least activity was observed during Sunrise and Lunar Midnight (P, Author summary The Asian tiger mosquito, Aedes albopictus, is an invasive mosquito which is now established in at least 40 states in the USA. Lack of efficient surveillance and control methods against Ae. albopictus, in addition to human-aided accidental transportations, have played a great role in its rapid expansion. Although surveillance measures are becoming more systematic and effective, control of this species still poses a great challenge. Aedes albopictus is difficult to control in the larval stage because it primarily develops in artificial containers that are widespread in peridomestic habitats. These habitats are not only ubiquitous in these environments, they are also cryptic, inaccessible, and extremely difficult to control. Therefore, control of Ae. albopictus in these environments often relies on adult control measures which utilize insecticides dispersed through ultra-low volume equipment as a cold aerosol space spray. These adulticide applications are often conducted at night against endemic mosquito species which are primarily active between dawn and dusk. However, since Ae. albopictus has been traditionally classified as a day-biting mosquito, mosquito control specialists have had doubts about the efficacy of a nocturnal application against a diurnally active mosquito. These uncertainties about intervention efforts become even more important during public health outbreaks of mosquito-borne pathogens such as chikungunya, dengue, or Zika viruses when protection of public health is of paramount importance in peridomestic habitats. Our investigations provide evidence that Ae. albopictus exhibits activity throughout the day and night and that nighttime adulticide applications may indeed be effective against this species, and should not be disregarded.

Background Serological diagnosis of Zika virus (ZIKV) infection is challenging because of the antibody cross-reactivity among flaviviruses. At the same time, the role of Nucleic Acid Testing (NAT) is limited by the low proportion of symptomatic infections and the low average viral load. Here, we compared the diagnostic performance of commercially available IgM, IgAM, and IgG ELISAs in sequential samples during the ZIKV and chikungunya (CHIKV) epidemics and co-circulation of dengue virus (DENV) in Brazil and Venezuela. Methodology/Principal findings Acute (day of illness 1–5) and follow-up (day of illness ≥ 6) blood samples were collected from nine hundred and seven symptomatic patients enrolled in a prospective multicenter study between June 2012 and August 2016. Acute samples were tested by RT-PCR for ZIKV, DENV, and CHIKV. Acute and follow-up samples were tested for IgM, IgAM, and IgG antibodies to ZIKV using commercially available ELISAs. Among follow-up samples with a RT-PCR confirmed ZIKV infection, anti-ZIKV IgAM sensitivity was 93.5% (43/46), while IgM and IgG exhibited sensitivities of 30.3% (10/33) and 72% (18/25), respectively. An additional 24% (26/109) of ZIKV infections were detected via IgAM seroconversion in ZIKV/DENV/CHIKV RT-PCR negative patients. The specificity of anti-ZIKV IgM was estimated at 93% and that of IgAM at 85%. Conclusions/Significance Our findings exemplify the challenges of the assessment of test performance for ZIKV serological tests in the real-world setting, during co-circulation of DENV, ZIKV, and CHIKV. However, we can also demonstrate that the IgAM immunoassay exhibits superior sensitivity to detect ZIKV RT-PCR confirmed infections compared to IgG and IgM immunoassays. The IgAM assay also proves to be promising for detection of anti-ZIKV seroconversions in sequential samples, both in ZIKV PCR-positive as well as PCR-negative patients, making this a candidate assay for serological monitoring of pregnant women in future ZIKV outbreaks., Author summary Zika virus (ZIKV) is transmitted through the bite of infected Aedes mosquitos but can also be transmitted sexually or vertically from mother-to-child. The same mosquitoes transmit dengue virus (DENV) and chikungunya virus (CHIKV), which cause similar clinical syndromes. The ZIKV epidemics in the Pacific and the Americas that occurred between 2015 and 2017 were linked to congenital abnormalities, most prominently microcephaly, in newborns. Because most infections are asymptomatic, diagnosis via indirect serological assays is an important strategy. On the other hand, many serological assays are affected by cross-reactivity resulting from prior infections by closely related viruses, such as DENV. This study evaluated three commercially available and widely used immunoassays that detect IgG, IgM or IgA and M (IgAM) antibodies to ZIKV. Our results suggest that the IgAM test performs best by detecting around 90% of RT-PCR confirmed infections. We also detected additional infections that were not detected by RT-PCR. The strength of this study is that it was carried out in two different countries of the American region where several arboviruses are endemic and that sequential blood samples from individual patients were available to evaluate the performance of the tests over time.

Chikungunya virus (CHIKV) is considered a public health problem due to its rapid spread and high morbidity. In 2016–2017 an outbreak of CHIKV was occurred in Pakistan but the data regarding the genomic diversity of CHIKV was not reported. Hence, the current study aimed to determine the genetic diversity of CHIKVs in Pakistan. A cross sectional study was carried out using sera of infected CHIKV patients (n = 1549) during the outbreak in Pakistan (2016–2018). Nucleotide sequencing of non-structural genes of CHIKV from eight isolates were performed followed by phylogenetic analysis using Bayesian method. Phylogenetic analysis suggested that the Pakistani CHIKV strains belonged to Indian Ocean Lineage (IOL) of genotype ECSA and C1.3a clade. Furthermore, the Pakistani isolates showed several key mutations (nsP2-H130Y, nsP2-E145D, nsP4-S55N and nsP4- R85G) corresponding to mutations reported in 2016 Indian strains of CHIKV. The molecular analysis revealed high evolutionary potential of CHIKV strains as well as better understanding of enhanced virulence and pathogenesis of this outbreak. The study highlights the need to continue surveillance in order to understand viral diversity over time and to devise preventive measures to limit diseases transmission in the region.

International audience; Mosquito control is implemented when arboviruses are detected in patients or in field-collected mosquitoes. However, mass screening of mosquitoes is usually laborious and expensive, requiring specialized expertise and equipment. Detection of virus in mosquito saliva using honey-impregnated filter papers seems to be a promising method as it is non-destructive and allows monitoring the viral excretion dynamics over time from the same mosquito. Here we test the use of filter papers to detect chikungunya virus in mosquito saliva in laboratory conditions, before proposing this method in large-scale mosquito surveillance programs. We found that 0.9 cm 2 cards impregnated with a 50% honey solution could replace the forced salivation technique as they offered a viral RNA detection until 7 days after oral infection of Aedes aegypti and Aedes albopictus mosquitoes with CHIKV.

Background Mosquitoes host and transmit numerous arthropod-borne viruses (arboviruses) that cause disease in both humans and animals. Effective surveillance of virome profiles in mosquitoes is vital to the prevention and control of mosquito-borne diseases in northwestern China, where epidemics occur frequently. Methods Mosquitoes were collected in the Shaanxi-Gansu-Ningxia region (Shaanxi Province, Gansu Province, and Ningxia Hui Autonomous Region) of China from June to August 2019. Morphological methods were used for taxonomic identification of mosquito species. High-throughput sequencing and metagenomic analysis were used to characterize mosquito viromes. Results A total of 22,959 mosquitoes were collected, including Culex pipiens (45.7%), Culex tritaeniorhynchus (40.6%), Anopheles sinensis (8.4%), Aedes (5.2%), and Armigeres subalbatus (0.1%). In total, 3,014,183 (0.95% of clean reads) viral sequences were identified and assigned to 116 viral species (including pathogens such as Japanese encephalitis virus and Getah virus) in 31 viral families, including Flaviviridae, Togaviridae, Phasmaviridae, Phenuiviridae, and some unclassified viruses. Mosquitoes collected in July (86 species in 26 families) showed greater viral diversity than those from June and August. Culex pipiens (69 species in 25 families) and Culex tritaeniorhynchus (73 species in 24 families) carried more viral species than Anopheles sinensis (50 species in 19 families) or Aedes (38 species in 20 families) mosquitoes. Conclusion Viral diversity and abundance were affected by mosquito species and collection time. The present study elucidates the virome compositions of various mosquito species in northwestern China, improving the understanding of virus transmission dynamics for comparison with those of disease outbreaks., Author summary Most traditional viral research methods are limited to screening for a few types of viruses with low throughput. Here, we relied on metagenomic sequencing, which is a broadly non-specific screen for any virus, for monitoring of mosquito-borne viruses in the Shaanxi-Gansu-Ningxia region of China in 2019. The baseline of viral abundance and diversity in local Culex pipiens, Culex tritaeniorhynchus, Anopheles sinensis, and Aedes mosquitoes were investigated, with 116 viral species identified in 31 viral families, including Flaviviridae, Togaviridae, Phasmaviridae, Phenuiviridae, and some unclassified viruses. This study provides a baseline survey of the viromes of mosquitoes in the study area, highlighting the necessity of strengthened monitoring of mosquito-borne pathogens.

Background As the three major arthropod-borne viruses, dengue virus (DENV), chikungunya virus (CHIKV), and zika virus (ZIKV) are posing a growing threat to global public health and socioeconomic development. Our study aimed to systematically review the global seroprevalences of these arboviruses from existing publications. Methods Articles published between Jan 01, 2000 and Dec 31, 2019 in the databases of Embase, Pubmed and Web of Science were searched and collected. Countries or areas with known local presence of Aedes vector mosquitoes were included. Random effects model was utilized to estimate the pooled seroprevalences and the proportion of inapparent infection. Results Out of 1375, a total of 133 articles involving 176,001 subjects were included for our analysis. The pooled seroprevalences of DENV, CHIKV and ZIKV were 38%, 25% and 18%, respectively; and their corresponding proportions of inapparent infections were 80%, 40% and 50%. The South-East Asia Region had the highest seroprevalences of DENV and CHIKV, while the Region of the Americas had the highest seroprevalence of ZIKV. The seroprevalences of DENV and CHIKV were similar when comparing developed and developing countries, urban and rural areas, or among different populations. In addition, we observed a decreased global seroprevalences in the new decade (2010–2019) comparing to the decade before (2000–2009) for CHIKV. For ZIKV, the positive rates tested with the nucleic acid detection method were lower than those tested with the antibody detection method. Lastly, numerous cases of dual seropositivity for CHIKV and DENV were reported. Conclusions Our results revealed a varied prevalence of arbovirus infections in different geographical regions and countries, and the inapparent infection accounted an unneglected portion of infections that requires more attention. This study will shed lights on our understanding of the true burden of arbovirus infections and promote appropriate vaccination in the future., Author summary DENV, CHIKV, and ZIKV infections are the three major neglected tropical diseases continually posing threats to the public health and the socioeconomic development in recent decades. Given a high fraction of inapparent infections of these viruses, it is necessary to obtain informative knowledge of the seroepidemiology of DENV, CHIKV, and ZIKV worldwide, so as to estimate the true burden of these diseases and evaluate effects on vaccine efficacy and uptake. Given the fact that the reported infection rates may vary greatly due to the measurement, criteria of inclusion, and etc, a systematic review with meta-analysis is essential to collect and summarize the seroepidemiologic evidence of these arboviruses. In this study, the geographical distributions of DENV, CHIKV, and ZIKV and the proportions of their inapparent infections were analyzed, which revealed an overall high disease burden to the world, especially in specific regions. The epidemiological features of these arboviruses were also characterized via comparisons made in different countries/areas, populations, and time periods. In addition, evidence on the co-infection of DENV and CHIKV was assessed.

Mayaro virus (MAYV) is an alphavirus endemic to South and Central America associated with sporadic outbreaks in humans. MAYV infection causes severe joint and muscle pain that can persist for weeks to months. Currently, there are no approved vaccines or therapeutics to prevent MAYV infection or treat the debilitating musculoskeletal inflammatory disease. In the current study, a prophylactic MAYV vaccine expressing the complete viral structural polyprotein was developed based on a non-replicating human adenovirus V (AdV) platform. Vaccination with AdV-MAYV elicited potent neutralizing antibodies that protected WT mice against MAYV challenge by preventing viremia, reducing viral dissemination to tissues and mitigating viral disease. The vaccine also prevented viral-mediated demise in IFN⍺R1-/- mice. Passive transfer of immune serum from vaccinated animals similarly prevented infection and disease in WT mice as well as virus-induced demise of IFN⍺R1-/- mice, indicating that antiviral antibodies are protective. Immunization with AdV-MAYV also generated cross-neutralizing antibodies against two related arthritogenic alphaviruses–chikungunya and Una viruses. These cross-neutralizing antibodies were protective against lethal infection in IFN⍺R1-/- mice following challenge with these heterotypic alphaviruses. These results indicate AdV-MAYV elicits protective immune responses with substantial cross-reactivity and protective efficacy against other arthritogenic alphaviruses. Our findings also highlight the potential for development of a multi-virus targeting vaccine against alphaviruses with endemic and epidemic potential in the Americas., Author summary Mayaro virus is an understudied alphavirus that is currently circulating in tropical environments in South and Central America without an approved vaccine. Recent outbreaks have suggested a broadening range and higher likelihood of urban outbreaks, increasing the public health risk. Mayaro virus is closely related to other arthritogenic alphaviruses with overlapping circulation such as chikungunya and Una viruses, both of which also lack clinically approved vaccines. Identification of a safe, easily manufactured, and effective strategy to vaccinate at risk populations is important to control outbreak potential. Here we report on a vaccination approach using a non-replicating adenovirus viral vector that encodes Mayaro virus structural proteins that assemble into non-infectious virus-like particles upon expression following vaccination. These particles stimulate strong immune responses against Mayaro virus. Upon testing against other alphaviruses, it was determined that the vaccine elicits cross-reactive neutralizing antibodies against chikungunya and Una viruses, significantly diminishes disease severity, and protects immunocompromised, highly susceptible mice from death following viral challenge. Our study provides new approaches to protect against these co-circulating viruses using a single vaccine. This approach is highly amenable to other virus targets for vaccine development and its ability to provide protection against chikungunya virus has global ramifications.

Background In 2014, a first outbreak of chikungunya hit the Caribbean area where chikungunya virus (CHIKV) had never circulated before. Methodology/Principal findings We conducted a cross-sectional study to measure the seroprevalence of CHIKV immediately after the end of the 2014 outbreak in HIV-infected people followed up in two clinical cohorts at the University hospitals of Guadeloupe and Martinique. Study patients were identified during the first months of 2015 and randomly selected to match the age and sex distribution of the general population in the two islands. They were invited to complete a survey that explored the symptoms consistent with chikungunya they could have developed during 2014 and to have a blood sample drawn for CHIKV serology. The study population consisted of 377 patients (198 in Martinique and 179 in Guadeloupe, 178 men and 199 women), 182 of whom reported they had developed symptoms consistent with chikungunya. CHIKV serology was positive in 230 patients, which accounted for an overall seroprevalence rate of 61% [95%CI 56–66], with only 153 patients who reported symptoms consistent with chikungunya. Most frequent symptoms included arthralgia (94.1%), fever (73.2%), myalgia (53.6%), headache (45.8%), and skin rash (26.1%). Conclusions/Significance This study showed that the seroprevalence of CHIKV infection was 61% after the 2014 outbreak, with one third of asymptomatic infections. Trial registration ClinicalTrials.gov NCT 02553369., Author summary After the termination of the first documented outbreak of chikungunya in the Caribbean in 2014, the seroprevalence rate of CHIKV infection in French West Indies was estimated at 61% [95% CI 56.0–65.8] in a population-matched cohort of HIV-infected patients. This high seroprevalence may explain the abrupt and persistent cessation CHIKV circulation in these territories. One third of participants had not developed clinical symptoms of chikungunya.

Throughout the last decade, chikungunya virus (CHIKV) and Zika virus (ZIKV) infections have spread globally, causing a spectrum of disease that ranges from self-limited febrile illness to permanent severe disability, congenital anomalies, and early death. Nevertheless, estimates of their aggregate health impact are absent from the literature and are currently omitted from the Global Burden of Disease (GBD) reports. We systematically reviewed published literature and surveillance records to evaluate the global burden caused by CHIKV and ZIKV between 2010 and 2019, to calculate estimates of their disability-adjusted life year (DALY) impact. Extracted data on acute, chronic, and perinatal outcomes were used to create annualized DALY estimates, following techniques outlined in the GBD framework. This study is registered with PROSPERO (CRD42020192502). Of 7,877 studies identified, 916 were screened in detail, and 21 were selected for inclusion. Available data indicate that CHIKV and ZIKV caused the average yearly loss of over 106,000 and 44,000 DALYs, respectively, between 2010 and 2019. Both viruses caused substantially more burden in the Americas than in any other World Health Organization (WHO) region. This unequal distribution is likely due to a combination of limited active surveillance reporting in other regions and the lack of immunity that left the previously unexposed populations of the Americas susceptible to severe outbreaks during the last decade. Long-term rheumatic sequelae provided the largest DALY component for CHIKV, whereas congenital Zika syndrome (CZS) contributed most significantly for ZIKV. Acute symptoms and early mortality accounted for relatively less of the overall burden. Suboptimal reporting and inconsistent diagnostics limit precision when determining arbovirus incidence and frequency of complications. Despite these limitations, it is clear from our assessment that CHIKV and ZIKV represent a significant cause of morbidity that is not included in current disease burden reports. These results suggest that transmission-blocking strategies, including vector control and vaccine development, remain crucial priorities in reducing global disease burden through prevention of potentially devastating arboviral outbreaks., Author summary Chikungunya and Zika are 2 mosquito-borne viral diseases that can cause both acute symptoms and long-term, debilitating complications in infected individuals. Chikungunya is best known as a cause of persistent arthritis in otherwise recovered patients and Zika as a cause of cognitive, motor, and sensory anomalies in newborn children. Both diseases emerged in the Americas within the last decade and have since spread rapidly throughout the region. Despite their widespread transmission there and throughout much of the world, chikungunya and Zika remain neglected diseases. One of the most significant obstacles to address their spread is a lack of data involving their burden. We searched the published literature and surveillance reports to collect information about the incidence, mortality, and morbidity associated with each of these diseases to estimate their regional and global burden during the last decade. Our estimates confirm that chikungunya and Zika caused substantial burden throughout this time frame and place them among the most problematic mosquito-borne viral diseases worldwide. We found that the largest proportion of global burden linked to each disease between 2010 and 2019 occurred in the Americas, although this observation is likely due to limited reporting in other regions.

Dengue, Zika and chikungunya are diseases of global health significance caused by arboviruses and transmitted by the mosquito Aedes aegypti, which is of worldwide circulation. The arrival of the Zika and chikungunya viruses to South America increased the complexity of transmission and morbidity caused by these viruses co-circulating in the same vector mosquito species. Here we present an integrated analysis of the reported arbovirus cases between 2007 and 2017 and local climate and socio-economic profiles of three distinct Colombian municipalities (Bello, Cúcuta and Moniquirá). These locations were confirmed as three different ecosystems given their contrasted geographic, climatic and socio-economic profiles. Correlational analyses were conducted with both generalised linear models and generalised additive models for the geographical data. Average temperature, minimum temperature and wind speed were strongly correlated with disease incidence. The transmission of Zika during the 2016 epidemic appeared to decrease circulation of dengue in Cúcuta, an area of sustained high incidence of dengue. Socio-economic factors such as barriers to health and childhood services, inadequate sanitation and poor water supply suggested an unfavourable impact on the transmission of dengue, Zika and chikungunya in all three ecosystems. Socio-demographic influencers were also discussed including the influx of people to Cúcuta, fleeing political and economic instability from neighbouring Venezuela. Aedes aegypti is expanding its range and increasing the global threat of these diseases. It is therefore vital that we learn from the epidemiology of these arboviruses and translate it into an actionable local knowledge base. This is even more acute given the recent historical high of dengue cases in the Americas in 2019, preceding the COVID-19 pandemic, which is itself hampering mosquito control efforts., Author summary Viruses transmitted by Ae. aegypti mosquitoes (dengue, Zika, chikungunya) are amongst the most significant public health concerns of recent years due to the increase in global cases and the rapid spread of the vectors. The primary method of controlling the spread of these arboviruses is through mosquito control. Understanding factors associated with risk of these viruses is key for informing control programmes and predicting when outbreaks may occur. Climate is an important driver in mosquito development and virus reproduction and hence the association of climate with disease risk. Socio-economic factors contribute to perpetuate disease risk. Areas of high poverty have abundance of suitable habitat for Ae. aegypti (e.g. due to poor housing and sanitation). This study investigated the factors affecting arbovirus incidence in three distinct regions of Colombia: Bello, Cúcuta and Moniquirá. The results show significant relationships between disease incidence and temperature, precipitation and wind speed. A decline in dengue following outbreaks of Zika (2016) is also reported. Measures of poverty, including critical overcrowding and no access to improved water source were also found to be higher in areas of higher disease incidence. The results of this study highlight the importance of using a multifactorial approach when designing vector control programs in order to effectively distribute local health care resources.

During the 2015–2017 Zika epidemic, dengue and chikungunya–two other viral diseases with the same vector as Zika–were also in circulation. Clinical presentation of these diseases can vary from person to person in terms of symptoms and severity, making it difficult to differentially diagnose them. Under these circumstances, it is possible that numerous cases of Zika could have been misdiagnosed as dengue or chikungunya, or vice versa. Given the importance of surveillance data for informing epidemiological analyses, our aim was to quantify the potential extent of misdiagnosis during this epidemic. Using basic principles of probability and empirical estimates of diagnostic sensitivity and specificity, we generated revised estimates of reported cases of Zika that accounted for the accuracy of diagnoses made on the basis of clinical presentation with or without laboratory confirmation. Applying this method to weekly reported case data from 43 countries throughout Latin America and the Caribbean, we estimated that 944,700 (95% CrI: 884,900–996,400) Zika cases occurred when assuming all confirmed cases were diagnosed using molecular methods versus 608,400 (95% CrI: 442,000–821,800) Zika cases that occurred when assuming all confirmed cases were diagnosed using serological methods. Our results imply that misdiagnosis was more common in countries with proportionally higher reported cases of dengue and chikungunya, such as Brazil. Given that Zika, dengue, and chikungunya appear likely to co-circulate in the Americas and elsewhere for years to come, our methodology has the potential to enhance the interpretation of passive surveillance data for these diseases going forward. Likewise, our methodology could also be used to help resolve transmission dynamics of other co-circulating diseases with similarities in symptomatology and potential for misdiagnosis., Author summary In 2016, the Zika epidemic in the Americas was declared a Public Health Emergency of International Concern by the World Health Organization, due to the finding that Zika virus infection was associated with microcephaly. In total, nearly 700,000 cases of Zika were reported to the Pan American Health Organization between 2015 and 2017. During this time, there were also nearly 4 million cases of dengue and chikungunya, diseases that share a common mosquito vector, and similar symptomatology, with Zika. In a region with co-circulation of these diseases, high potential for clinical misdiagnosis among these diseases has been found. This raises the question of how misdiagnosis could have affected reports of the overall size of the Zika epidemic. We addressed this question by leveraging passive surveillance data and empirical estimates of misdiagnosis to quantify the spatial and temporal extent of misdiagnosis between Zika, dengue, and chikungunya. Our results suggest that, across the Americas, the Zika epidemic was likely larger than indicated by passive surveillance data taken at face value, but that conclusions about the overall size of the epidemic depend on assumptions made about sensitivities and specificities of laboratory diagnostic tests. We estimated that many misdiagnosed Zika cases occurred in 2015, prior to the start of reporting of Zika in most countries.

Made available in DSpace on 2021-06-25T10:55:22Z (GMT). No. of bitstreams: 0 Previous issue date: 2021-03-01 National Institute of Biological Resources The potential outcome of flavivirus and alphavirus co-infections is worrisome due to the development of severe diseases. Hundreds of millions of people worldwide live under the risk of infections caused by viruses like chikungunya virus (CHIKV, genus Alphavirus), dengue virus (DENV, genus Flavivirus), and zika virus (ZIKV, genus Flavivirus). So far, neither any drug exists against the infection by a single virus, nor against co-infection. The results described in our study demonstrate the inhibitory potential of two flavonoids derived from citrus plants: Hesperetin (HST) against NS2B/NS3pro of ZIKV and nsP2pro of CHIKV and, Hesperidin (HSD) against nsP2pro of CHIKV. The flavonoids are noncompetitive inhibitors and the determined IC50 values are in low µM range for HST against ZIKV NS2B/NS3pro (12.6 ± 1.3 µM) and against CHIKV nsP2pro (2.5 ± 0.4 µM). The IC50 for HSD against CHIKV nsP2pro was 7.1 ± 1.1 µM. The calculated ligand efficiencies for HST were > 0.3, which reflect its potential to be used as a lead compound. Docking and molecular dynamics simulations display the effect of HST and HSD on the protease 3D models of CHIKV and ZIKV. Conformational changes after ligand binding and their effect on the substrate-binding pocket of the proteases were investigated. Additionally, MTT assays demonstrated a very low cytotoxicity of both the molecules. Based on our results, we assume that HST comprise a chemical structure that serves as a starting point molecule to develop a potent inhibitor to combat CHIKV and ZIKV co-infections by inhibiting the virus proteases. Multiuser Center for Biomolecular Innovation Departament of Physics Instituto de Biociências Letras e Ciências Exatas (Ibilce) Universidade Estadual Paulista (UNESP) Institute of Biological Information Processing (IBI-7: Structural Biochemistry) Forschungszentrum Jülich Federal University of Tocantins Instituto de Biociências Letras e Ciências Exatas (Ibilce) Universidade Estadual Paulista (UNESP) FACERES Medical School Faculdade de Medicina de São José do Rio Preto-FAMERP Institute of Physics Federal University of Mato Grosso do Sul Institut für Physikalische Biologie Heinrich-Heine-Universität Düsseldorf JuStruct: Jülich Centre for Structural Biology Forchungszentrum Jülich Multiuser Center for Biomolecular Innovation Departament of Physics Instituto de Biociências Letras e Ciências Exatas (Ibilce) Universidade Estadual Paulista (UNESP) Instituto de Biociências Letras e Ciências Exatas (Ibilce) Universidade Estadual Paulista (UNESP)

Background Statistical models are regularly used in the forecasting and surveillance of infectious diseases to guide public health. Variable selection assists in determining factors associated with disease transmission, however, often overlooked in this process is the evaluation and suitability of the statistical model used in forecasting disease transmission and outbreaks. Here we aim to evaluate several modelling methods to optimise predictive modelling of Ross River virus (RRV) disease notifications and outbreaks in epidemiological important regions of Victoria and Western Australia. Methodology/Principal findings We developed several statistical methods using meteorological and RRV surveillance data from July 2000 until June 2018 in Victoria and from July 1991 until June 2018 in Western Australia. Models were developed for 11 Local Government Areas (LGAs) in Victoria and seven LGAs in Western Australia. We found generalised additive models and generalised boosted regression models, and generalised additive models and negative binomial models to be the best fit models when predicting RRV outbreaks and notifications, respectively. No association was found with a model’s ability to predict RRV notifications in LGAs with greater RRV activity, or for outbreak predictions to have a higher accuracy in LGAs with greater RRV notifications. Moreover, we assessed the use of factor analysis to generate independent variables used in predictive modelling. In the majority of LGAs, this method did not result in better model predictive performance. Conclusions/Significance We demonstrate that models which are developed and used for predicting disease notifications may not be suitable for predicting disease outbreaks, or vice versa. Furthermore, poor predictive performance in modelling disease transmissions may be the result of inappropriate model selection methods. Our findings provide approaches and methods to facilitate the selection of the best fit statistical model for predicting mosquito-borne disease notifications and outbreaks used for disease surveillance., Author summary Mosquito-borne diseases cause significant illness worldwide. Mosquito breeding, which leads to disease transmission, is driven by favorable climatic and meteorological events (e.g., rainfall and warm temperatures). Understanding the association meteorological conditions have with mosquito breeding aids in directing mosquito control activities when there is a likelihood of disease transmission. Predictive models are used in public health decision making and resource allocation to guide mosquito control programs. However, there are multiple modelling methods, all of which provide differing degrees of accuracy in their predictions and suitability to the disease transmission dynamics. This study aims to assess commonly used statistical models for predicting mosquito-borne disease notifications and outbreaks. We demonstrate that statistical model selection plays an important role in accurately forecasting mosquito-borne disease and poor predictive performance may be due to inappropriate model selection. Furthermore, a model suited to predicting disease notifications may not always be the best model to accurately predict the occurrence of disease outbreaks. The methods used here can aid in public health to establish suitable predictive mosquito-borne disease surveillance systems to help guide disease prevention and resource allocation, and mosquito control activities.

Patients following infection by chikungunya virus (CHIKV) can suffer for months to years from arthralgia and arthritis. Interestingly, methotrexate (MTX) a major immune-regulatory drug has proved to be of clinical benefit. We have previously shown that CHIKV can persist in the joint of one patient 18 months post-infection and plausibly driving chronic joint inflammation but through ill-characterized mechanisms. We have pursued our investigations and report novel histological and in vitro data arguing for a plausible role of a COX-2-mediated inflammatory response post-CHIKV. In the joint, we found a robust COX-2 staining on endothelial cells, synovial fibroblasts and more prominently on multinucleated giant cells identified as CD11c+ osteoclasts known to be involved in bone destruction. The joint tissue was also strongly stained for CD3, CD8, CD45, CD14, CD68, CD31, CD34, MMP2, and VEGF (but not for NO synthase and two B cell markers). Dendritic cells were rarely detected. Primary human synovial fibroblasts were infected with CHIKV or stimulated either by the synthetic molecule polyriboinosinic:polyribocytidylic acid (PIC) to mimic chronic viral infection or cytokines. First, we found that PIC and CHIKV enhanced mRNA expression of COX-2. We further found that PIC but not CHIKV increased the mRNA levels of cPLA2α and of mPGES-1, two other central enzymes in PGE2 production. IFNβ upregulated cPLA2α and COX-2 transcription levels but failed to modulated mPGES-1 mRNA expression. Moreover, PIC, CHIKV and IFNβ decreased mRNA expression of the PGE2 degrading enzyme 15-PGDH. Interestingly, MTX failed to control the expression of all these enzymes. In sharp contrast, dexamethasone was able to control the capacity of pro-inflammatory cytokines, IL-1β as well as TNFα, to stimulate mRNA levels of cPLA2α, COX-2 and mPGES-1. These original data argue for a concerted action of CHIKV (including viral RNA) and cytokines plausibly released from recruited leukocytes to drive a major COX-2-mediated PGE2 proinflammatory responses to induce viral arthritis., Author summary It is important to have a better understanding of the immuno-pathogenesis of Chikungunya virus (CHIKV) and particularly focusing on the chronic phase associated to arthralgia and arthritis. Benefiting from our prospective cohort studies, we herein provide novel in vivo data identifying for the first time the implication of COX-2 and several other enzymes involved in prostaglandin biosynthesis and the persistence of the virus on the joint. Prostaglandin has major activities in inflammation and joint destruction. In vitro, we have used a model of human synovial fibroblasts to decipher the regulatory mechanisms of prostaglandin biosynthesis pathway. We have made important observations showing that the virus itself as well as major inflammatory cytokines can dramatically control the expression of all enzymes involved in the metabolism of prostaglandin. Interestingly, pharmacological investigations further revealed that dexamethasone, but not methotrexate (currently used to treat patients with chikungunya) may be of clinical values.

Defective viral genomes (DVGs) are truncated and/or rearranged viral genomes produced during virus replication. Described in many RNA virus families, some of them have interfering activity on their parental virus and/or strong immunostimulatory potential, and are being considered in antiviral approaches. Chikungunya virus (CHIKV) is an alphavirus transmitted by Aedes spp. that infected millions of humans in the last 15 years. Here, we describe the DVGs arising during CHIKV infection in vitro in mammalian and mosquito cells, and in vivo in experimentally infected Aedes aegypti mosquitoes. We combined experimental and computational approaches to select DVG candidates most likely to have inhibitory activity and showed that, indeed, they strongly interfere with CHIKV replication both in mammalian and mosquito cells. We further demonstrated that some DVGs present broad-spectrum activity, inhibiting several CHIKV strains and other alphaviruses. Finally, we showed that pre-treating Aedes aegypti with DVGs prevented viral dissemination in vivo., Author summary Defective viral genomes (DVGs) are produced during virus replication. On their own they cannot replicate, but some of them can compete with wild-type virus for viral and/or cellular resources. For chikungunya virus, interference by DVGs has not been described. Here, we use a new approach based on experimental evolution and computational analysis to characterize all DVGs generated in a virus population and identify those with the highest antiviral potential. We confirm their antiviral activity in both mammalian and mosquito host environments and show that some can broadly interfere with other strains or related alphaviruses. Finally, we show that DVGs can inhibit virus dissemination in mosquitoes.

Background As an invasive mosquito species in the United States, Aedes albopictus is a potential vector of arboviruses including dengue, chikungunya, and Zika, and may also be involved in occasional transmission of other arboviruses such as West Nile, Saint Louis encephalitis, eastern equine encephalitis, and La Crosse viruses. Aedes albopictus feeds on a wide variety of vertebrate hosts, wild and domestic, as well as humans. Methodology/Principal findings In order to investigate blood feeding patterns of Ae. albopictus, engorged specimens were collected from a variety of habitat types using the Centers for Disease Control and Prevention light traps, Biogents Sentinel 2 traps, and modified Reiter gravid traps in southeast Virginia. Sources of blood meals were determined by the analysis of mitochondrial cytochrome b gene sequences amplified in PCR assays. Our aims were to quantify degrees of Ae. albopictus interactions with vertebrate hosts as sources of blood meals, investigate arboviral infection status, assess the influence of key socioecological conditions on spatial variability in blood feeding, and investigate temporal differences in blood feeding by season. Analysis of 961 engorged specimens of Ae. albopictus sampled between 2017–2019 indicated that 96%, 4%, and less than 1% obtained blood meals from mammalian, reptilian, and avian hosts, respectively. Domestic cats were the most frequently identified (50.5%) hosts followed by Virginia opossums (17.1%), white-tailed deer (12.2%), and humans (7.3%), together representing 87.1% of all identified blood hosts. We found spatial patterns in blood feeding linked to socioecological conditions and seasonal shifts in Ae. albopictus blood feeding with implications for understanding human biting and disease risk. In Suffolk Virginia in areas of lower human development, the likelihood of human blood feeding increased as median household income increased and human blood feeding was more likely early in the season (May-June) compared to later (July-October). Screening of the head and thorax of engorged Ae. albopictus mosquitoes by cell culture and RT-PCR resulted in a single isolate of Potosi virus. Conclusion and significance Understanding mosquito-host interactions in nature is vital for evaluating vectorial capacity of mosquitoes. These interactions with competent reservoir hosts support transmission, maintenance, and amplification of zoonotic agents of human diseases. Results of our study in conjunction with abundance in urban/suburban settings, virus isolation from field-collected mosquitoes, and vector competence of Ae. albopictus, highlight the potential involvement of this species in the transmission of a number of arboviruses such as dengue, chikungunya, and Zika to humans. Limited interaction with avian hosts suggests that Ae. albopictus is unlikely to serve as a bridge vector of arboviruses such as West Nile and eastern equine encephalitis in the study region, but that possibility cannot be entirely ruled out., Author summary Native to Southeast Asia, breeding populations of Aedes albopictus were first discovered in Harris County, Texas, in 1985, and as of 2017, seasonal populations of this species have been reported in more than 40 states and the District of Columbia. Aedes albopictus breed readily in natural or man-made environment where stagnant water can accumulate such as gutters, flowerpots, discarded tires, and tree holes. This mosquito species has been implicated in outbreaks of chikungunya, dengue, and Zika viruses and is a competent vector of many arboviruses including West Nile, eastern equine encephalitis, yellow fever, Rift Valley fever, and Japanese encephalitis. Aedes albopictus is regarded as an opportunistic mosquito feeding on a variety of domestic and wild mammals, birds, reptiles and amphibians; however, a preference for human blood meals has been noted in blood meal analysis of field-collected mosquitoes and in laboratory investigations. We studied vector-host interactions of Ae. albopictus in Virginia, United States using molecular methods and identified ten mammalian, three reptilian, and two avian species as blood hosts of this mosquito species. Our study clarifies the host associations of Ae. albopictus and highlights concerns about the potential role of this mosquito species in transmission of emerging and reemerging arboviruses.

Background Chikungunya is now of public health concern globally due to its re-emergence in endemic areas and introduction into new areas of the world. Worldwide, the vectors for transmission of the chikungunya virus are Aedes mosquitoes and these are prevalent in Ghana. Despite its global significance, the true burden of chikungunya virus infection in Ghana is largely unknown and the threat of outbreak remains high owing to international travel. This study sought to determine chikungunya virus infection among febrile patients suspected of having malaria infections at some selected health facilities in the Ashanti, Bono East, and Bono Regions of Ghana. Methodology This cross-sectional study recruited six hundred (600) febrile patients suspected of having malaria who submitted their clinical samples to the laboratories of the selected health facilities for the diagnosis of their infections. Five to ten millilitres (5-10ml) of venous blood were collected from each study participant. Sera were separated and tested for anti-chikungunya (IgM and IgG) antibodies using InBios ELISA kit following the manufacturer’s instruction. Samples positive for chikungunya IgM and IgG were selected and tested for chikungunya virus RNA using Reverse Transcription-quantitative Polymerase Chain Reaction. Malaria Rapid Diagnostic Test kits were used to screen the participants for malaria. Structured questionnaires were administered to obtain demographic and clinical information of the study participants. Result Of the 600 samples tested, the overall seroprevalence of chikungunya was 6%. The seroprevalence of chikungunya IgM and IgG antibodies were 1.8% and 4.2% respectively. None of the chikungunya IgM and IgG positive samples tested positive for chikungunya RNA by RT-qPCR. Of the 600 samples, tested 32.3% (194/600) were positive for malaria parasites. Malaria and chikungunya co-infection was detected in 1.8% (11/600) of the participants. Conclusion Findings from the current study indicate low-level exposure to the chikungunya virus suggesting the virus is circulating and potentially causing morbidity in Ghana., Author summary Chikungunya fever is a mosquito-borne disease of Chikungunya virus (CHIKV) origin that shares similar symptoms with malaria fever. The symptoms include fever, joint pains, muscle pains, skin rashes and headache. There is a paucity of information about CHIKV epidemiology in Ghana although the mosquito vector (Aedes) responsible for the transmission is present in the country. Ghana’s neighbours (Ivory Coast and Burkina Faso) have also reported isolated cases of CHIKV infections. The authors sought to find evidence of the circulation of CHIKV among suspected malaria patients in Ghana and to shed light on the importance of the investigation of non-malaria causes of fever such as chikungunya fever. And the possible inclusion of chikungunya fever in the differential diagnoses of unexplained febrile cases in Ghana. We found out that 6% of the 600 people studied had antibodies to the CHIKV i.e. they were previously infected with CHIKV. Malaria prevalence was 32.3% among the study participants. The rate of malaria-chikungunya co-infections was 1.8% among the study participants. Our findings show a low-level exposure to the CHIKV suggesting the virus is circulating and potentially causing morbidity in Ghana that may be misdiagnosed as malaria.

Background Epidemic arbovirus transmission occurs among humans by mosquito bites and the sylvatic transmission cycles involving non-human primates (NHPs) still exists. However, limited data are available on the extent in NHPs infections and their role. In this study, we have developed and validated a high-throughput serological screening tool to study the circulation of multiple arboviruses that represent a significant threat to human health, in NHPs in Central Africa. Methodology/Principal findings Recombinant proteins NS1, envelope domain-3 (DIII) for the dengue (DENV), yellow fever (YFV), usutu (USUV), west nile (WNV) and zika (ZIKV) and envelope 2 for the chikungunya (CHIKV) and o'nyong-nyong (ONNV) were coupled to Luminex beads to detect IgG directed against these viruses. Evaluation of test performance was made using 161 human sera of known arboviral status (66 negative and 95 positive). The sensitivity and specificity of each antigen were determined by statistical methods and ROC curves (except for ONNV and USUV). All NS1 antigens (except NS1-YFV), CHIKV-E2 and WNV-DIII had sensitivities and specificities > 95%. For the other DIII antigens, the sensitivity was low, limiting the interest of their use for seroprevalence studies. Few simultaneous reactions were observed between the CHIKV+ samples and the NS1 antigens to the non-CHIKV arboviruses. On the other hand, the DENV+ samples crossed-reacted with NS1 of all the DENV serotypes (1 to 4), as well as with ZIKV, USUV and to a lesser extent with YFV. A total of 3,518 samples of 29 species of NHPs from Cameroon and the Democratic Republic of Congo (DRC) were tested against NS1 (except YFV), E2 (CHIKV/ONNV) and DIII (WNV) antigens. In monkeys (n = 2,100), the global prevalence varied between 2 and 5% for the ten antigens tested. When we stratified by monkey’s biotope, the arboreal species showed the highest reactivity. In monkeys from Cameroon, the highest IgG prevalence were observed against ONNV-E2 and DENV2-NS1 with 3.95% and 3.40% respectively and in DRC, ONNV-E2 (6.63%) and WNV-NS1 (4.42%). Overall prevalence was low in apes (n = 1,418): ranging from 0% for USUV-NS1 to 2.6% for CHIKV-E2. However, a very large disparity was observed among collection site and ape species, e.g. 18% (9/40) and 8.2% (4/49) of gorillas were reactive with CHIKV-E2 or WNV-NS1, respectively in two different sites in Cameroon. Conclusions/Significance We have developed a serological assay based on Luminex technology, with high specificity and sensitivity for simultaneous detection of antibodies to 10 antigens from 6 different arboviruses. This is the first study that evaluated on a large scale the presence of antibodies to arboviruses in NHPs to evaluate their role in sylvatic cycles. The overall low prevalence (, Author summary In the last decades, chikungunya, zika, yellow fever, usutu and dengue viruses have (re)-emerged in different parts of the world and many of these outbreaks occur in resource-limited countries with limited or under-equipped health facilities and where endemic malaria with very similar clinical symptoms confounds surveillance. Most arboviruses that circulate today likely originated in Africa where sporadic human outbreaks occur. In this work, we developed a serological tool that allows simultaneous detection of IgG antibodies to multiple arbovirus in a biological sample. With this highly sensitive and specific multiplex assay, we screened more than 3,500 samples collected from 29 species of monkeys and apes in Africa. We found a global IgG antibody prevalence of less than 5%. However, this seroprevalence varied by collection site, NPHs species and virus type. Given these findings, we concluded that African non-human primates are most likely not the reservoirs, but rather are intermediate hosts.

Poxvirus systems have been extensively used as vaccine vectors. Herein a RNA-Seq analysis of intramuscular injection sites provided detailed insights into host innate immune responses, as well as expression of vector and recombinant immunogen genes, after vaccination with a new multiplication defective, vaccinia-based vector, Sementis Copenhagen Vector. Chikungunya and Zika virus immunogen mRNA and protein expression was associated with necrosing skeletal muscle cells surrounded by mixed cellular infiltrates. The multiple adjuvant signatures at 12 hours post-vaccination were dominated by TLR3, 4 and 9, STING, MAVS, PKR and the inflammasome. Th1 cytokine signatures were dominated by IFNγ, TNF and IL1β, and chemokine signatures by CCL5 and CXCL12. Multiple signatures associated with dendritic cell stimulation were evident. By day seven, vaccine transcripts were absent, and cell death, neutrophil, macrophage and inflammation annotations had abated. No compelling arthritis signatures were identified. Such injection site vaccinology approaches should inform refinements in poxvirus-based vector design., Author summary Poxvirus vector systems have been widely developed for vaccine applications. Despite considerable progress, so far only one recombinant poxvirus vectored vaccine has to date been licensed for human use, with ongoing efforts seeking to enhance immunogenicity whilst minimizing reactogenicity. The latter two characteristics are often determined by early post-vaccination events at the injection site. We therefore undertook an injection site vaccinology approach to analyzing gene expression at the vaccination site after intramuscular inoculation with a recombinant, multiplication defective, vaccinia-based vaccine. This provided detailed insights into inter alia expression of vector-encoded immunoregulatory genes, as well as host innate and adaptive immune responses. We propose that such injection site vaccinology can inform rational vaccine vector design, and we discuss how the information and approach elucidated herein might be used to improve immunogenicity and limit reactogenicity of poxvirus-based vaccine vector systems.

Arboviruses are arthropod-dependent viruses to complete their zoonotic cycle. Among the transmitting arthropods, culicids stand out, which participate in the cycle of several arboviruses that can affect humans. The present study aimed to identify species of culicidae and to point out the risk of circulation, emergency, or reemergence of pathogenic arboviruses to humans in the region of the Jequitibá headquarters of the Parque Estadual dos Três Picos (PETP), in Cachoeiras de Macacu, state of Rio de Janeiro, Brazil. Sampling was carried out at five Sample Points (SP) demarcated on trails from the headquarters, with CDC light traps, HP model with dry ice attached to the side, for 48 hours of activity each month. Additionally, active catches were made with a castro catcher in the period of one hour per day in the field, from six to eleven o’clock in the morning, in each PM. After the captures, thematic map was assembled using the ArcGIS 10 software and performing a multidimensional scaling (MDS). A total of 1151 specimens were captured and the presence of culicids already incriminated as vectors of arboviruses circulating in the region was observed: Aedes fluviatilis Lutz, 1904 (71 specimens); Aedes scapularis Rondani, 1848 (55 specimens); Haemagogus leococelaenus Dyar and Shannon, 1924 (29 specimens). In addition to the subgenus Culex (culex) spp. (163 specimens). In this sense, we highlight the importance of strengthening the actions of continuous entomological surveillance of the emergence and re-emergence of new arboviruses in ecotourism visitation parks.

Following an outbreak of chikungunya virus (CHIKV) infections in Thailand in 2019, numerous cases of CHIKV infection have been diagnosed in Bangkok, the capital of the country. In our previous investigation of the vectors for disease transmission, we found natural infection of CHIKV in both male and female Aedes aegypti mosquitoes collected from the outbreak areas in Bangkok. Some reports mentioned the detection of CHIKV in Culex mosquitoes. In Thailand, the Culex quinquefasciatus Say mosquito is a common species found in urban and rural settings that coexists with Ae. aegypti. However, the role of Cx. quinquefasciatus mosquitoes in the spread of the Indian Ocean Lineage (IOL) of CHIKV in Thailand has never been investigated. In this study, Cx. quinquefasciatus were collected (16 males and 27 females) from an outbreak area in Bangkok. Eight of the 27 in field-caught female Cx. quinquefasciatus were positive for IOL CHIKV RNA, and 99–100% identity and full 100% coverage of sequences similar to CHIKV isolated from female Ae. aegypti in Bangkok, Thailand, whereas viral RNA was not detected in male samples using nested-RT-PCR. To determine whether CHIKV is able to replicate in Cx. quinquefasciatus, the laboratory strain of Cx. quinquefasciatus was allowed to feed on blood containing IOL CHIKV isolated from patient serum. The nested-RT-PCR, virus isolation, and immunofluorescence assay (IFA) were performed for CHIKV detection and replication. The results showed that CHIKV RNA was detected in Cx. quinquefasciatus until day 4 post infection. CHIKV did not produce any remarkable signs of infection, dissemination, or transmission in Cx. quinquefasciatus, and cytopathic effect (CPE) was not observed in C6/36 cells when infected with supernatant obtained from Cx. quinquefasciatus at days 7, 10, 14, and 21 post infection when compared to Ae. aegypti. The data from this study infer that CHIKV may be detected in Cx. quinquefasciatus but that the mosquito is not able to transmit CHIKV in Thailand.

Aedes aegypti and Aedes albopictus vector dengue, chikungunya and Zika viruses. With both species expanding their global distributions at alarming rates, developing effective surveillance equipment is a continuing priority for public health researchers. Sound traps have been shown, in limited testing, to be highly species-specific when emitting a frequency corresponding to a female mosquito wingbeat. Determining male mosquito capture rates in sound traps based on lure frequencies in endemic settings is the next step for informed deployment of these surveillance tools. We field-evaluated Male Aedes Sound Traps (MASTs) set to either 450 Hz, 500 Hz, 550 Hz or 600 Hz for sampling Aedes aegypti and/or Aedes albopictus and compared catch rates to BG-Sentinel traps within Pacific (Madang, Papua New Guinea) and Latin American (Molas, Mexico and Orange Walk Town, Belize) locations. MASTs set to 450–550 Hz consistently caught male Ae. aegypti at rates comparable to BG-Sentinel traps in all locations. A peak in male Ae. albopictus captures in MASTs set at 550 Hz was observed, with the lowest mean abundance recorded in MASTs set to 450 Hz. While significantly higher abundances of male Culex were sampled in MASTs emitting lower relative frequencies in Molas, overall male Culex were captured in significantly lower abundances in the MASTs, relative to BG-Sentinel traps within all locations. Finally, significant differences in rates at which male Aedes and Culex were positively detected in trap-types per weekly collections were broadly consistent with trends in abundance data per trap-type. MASTs at 550 Hz effectively captured both male Ae. aegypti and Ae. albopictus while greatly reducing bycatch, especially male Culex, in locations where dengue transmission has occurred. This high species-specificity of the MAST not only reduces staff-time required to sort samples, but can also be exploited to develop an accurate smart-trap system—both outcomes potentially reducing public health program expenses., Author summary The continued global expansion of Aedes aegypti and Aedes albopictus, which transmit viruses causing dengue and Zika, necessitates the further development of effective mosquito traps for monitoring and predicting transmission risk. Mosquito sound traps attract male mosquitoes by producing a frequency which mimics the wingbeat sound of female mosquitoes. As few insects will respond to such sounds, these traps can be highly species-specific, saving significant person-time to sort samples, potentially paving the way for more accurate smart-trap systems and reduced surveillance costs. To date, limited research has occurred within field-settings investigating the capture rates of male mosquitoes in sound traps set to different frequencies. To this point, we performed trials in dengue endemic environments to quantify capture rates of mosquitoes in Male Aedes Sound Traps set to different frequencies in Papua New Guinea, Mexico and Belize. We found that both male Ae. aegypti and Ae. albopictus responded positively to frequencies between 450 and 600 Hz. Additionally, male Culex mosquitoes were also caught, but at lower abundances in traps set to higher sound frequencies. These traps, when set to 550 Hz, effectively capture male Ae. aegypti or Ae. albopictus with limited bycatch.

The macrodomain of nsP3 (nsP3MD) is highly conserved among the alphaviruses and ADP-ribosylhydrolase activity of Chikungunya Virus (CHIKV) nsP3MD is critical for CHIKV viral replication and virulence. No small molecule drugs targeting CHIKV nsP3 have been identified to date. Here we report small fragments that bind to nsP3MD which were discovered by virtually screening a fragment library and X-ray crystallography. These identified fragments share a similar scaffold, 2-pyrimidone-4-carboxylic acid, and are specifically bound to the ADP-ribose binding site of nsP3MD. Among the fragments, 2-oxo-5,6-benzopyrimidine-4-carboxylic acid showed anti-CHIKV activity with an IC50 of 23 μM. Our fragment-based drug discovery approach provides valuable information to further develop a specific and potent nsP3 inhibitor of CHIKV viral replication based on the 2-pyrimidone-4-carboxylic acid scaffold. In silico studies suggest this pyrimidone scaffold could also bind to the macrodomains of other alphaviruses and coronaviruses and thus, have potential pan-antiviral activity.

Chikungunya virus (CHIKV) is an arthropod-borne virus that causes large outbreaks world-wide leaving millions of people with severe and debilitating arthritis. Interestingly, clinical presentation of CHIKV arthritides have many overlapping features with rheumatoid arthritis including cellular and cytokine pathways that lead to disease development and progression. Currently, there are no specific treatments or vaccines available to treat CHIKV infections therefore advocating the need for the development of novel therapeutic strategies to treat CHIKV rheumatic disease. Herein, we provide an in-depth analysis of an efficacious new treatment for CHIKV arthritis with a semi-synthetic sulphated polysaccharide, Pentosan Polysulfate Sodium (PPS). Mice treated with PPS showed significant functional improvement as measured by grip strength and a reduction in hind limb foot swelling. Histological analysis of the affected joint showed local inflammation was reduced as seen by a decreased number of infiltrating immune cells. Additionally, joint cartilage was protected as demonstrated by increased proteoglycan staining. Using a multiplex-immunoassay system, we also showed that at peak disease, PPS treatment led to a systemic reduction of the chemokines CXCL1, CCL2 (MCP-1), CCL7 (MCP-3) and CCL12 (MCP-5) which may be associated with the reduction in cellular infiltrates. Further characterisation of the local effect of PPS in its action to reduce joint and muscle inflammation was performed using NanoString™ technology. Results showed that PPS altered the local expression of key functional genes characterised for their involvement in growth factor signalling and lymphocyte activation. Overall, this study shows that PPS is a promising treatment for alphaviral arthritis by reducing inflammation and protecting joint integrity.

Aedes-borne viral diseases mainly Yellow Fever (YF), Dengue (DEN), Zika (ZIK) and Chikungunya (CHK) have contributed to many deaths’ in the world especially in Africa. There have been major outbreaks of these diseases in West Africa. Although, YF outbreaks have occurred in Ghana over the years, no outbreak of DEN, ZIK and CHK has been recorded. However, the risk of outbreak is high due to its proximity to West African countries where outbreaks have been recently been recorded. This study surveyed the mosquito fauna to assess the risk of transmission of Yellow fever (YFV), Dengue (DENV), Chikungunya (CHKV) and Zika (ZIKV) viruses in Larabanga and Mole Game Reserve areas in Northern Ghana. The immature and adult stages ofAedesmosquitoes were collected from Larabanga and Mole Game Reserve area. There was a significant (P>0.001) number of mosquitoes collected during the rainy season than the dry season. A total of 1,930Aedesmosquitoes were collected during the rainy season and morphologically identified. Of these, 1,915 (99.22%) wereAedes aegyptiand 15 (0.22%) wereAedes vittatus. During the dry season, 27Ae.aegyptimosquitoes were collected. A total of 415Ae.aegyptimosquitoes were molecularly identified to subspecies level of whichAe.(Ae) aegypti aegyptiwas the predominant subspecies. BothAe.aegypti aegypti and Ae aegypti formosusexist in sympatry in the area. AllAedespools (75) were negative for DENV, ZIKV and CHKV when examined by RT- PCR. Three Larval indices namely House Index, HI (percentage of houses positive forAedeslarvae or pupae), Container Index, CI (the percentage of containers positive forAedeslarvae or pupae) and Breteau Index, BI (number of positive containers per 100 houses inspected) were assessed as a measure for risk of transmission in the study area. The HI, CI and BI for both sites were as follows; Mole Game Reserve (HI, 42.1%, CI, 23.5% and BI, 100 for rainy season and 0 for all indices for dry season) and Larabanga (39%, 15.5% and 61 for rainy season and 2.3%, 1.3% and 2.3 for dry season). The spatial distribution ofAedesbreeding sites in both areas indicated thatAedeslarvae were breeding in areas with close proximity to humans. Lorry tires were the main source ofAedeslarvae in all the study areas. Information about the species composition and the potential role ofAedesmosquitoes in future outbreaks of the diseases that they transmit is needed to design efficient surveillance and vector control tools.

Between 2018 and 2019, the incidence of chikungunya was approximately 15,000 cases across 60 provinces in Thailand. Here, the clinical presentations in chikungunya, emergent pattern, and genomic diversity of the chikungunya virus (CHIKV) causing this massive outbreak were demonstrated. A total of 1,806 sera samples from suspected cases of chikungunya were collected from 13 provinces in Thailand, and samples were tested for the presence of CHIKV RNA, IgG, and IgM using real-time PCR, enzyme-linked immunoassay (ELISA), commercial immunoassay (rapid test). The phylogenetic tree of CHIKV whole-genome and CHIKV E1 were constructed using the maximum-likelihood method. CHIKV infection was confirmed in 547 (42.2%) male and 748 (57.8%) female patients by positive real-time PCR results and/or CHIKV IgM antibody titers. Unsurprisingly, CHIKV RNA was detected in >80% of confirmed cases between 1 and 5 days after symptom onset, whereas anti-CHIKV IgM was detectable in >90% of cases after day 6. Older age was clearly one of the risk factors for the development of arthralgia in infected patients. Although phylogenetic analysis revealed that the present CHIKV Thailand strain of 2018–2020 belongs to the East, Central, and Southern African (ECSA) genotype similar to the CHIKV strains that caused outbreaks during 2008–2009 and 2013, all present CHIKV Thailand strains were clustered within the recent CHIKV strain that caused an outbreak in South Asia. Interestingly, all present CHIKV Thailand strains possess two mutations, E1-K211E, and E2-V264A, in the background of E1-226A. These mutations are reported to be associated with virus-adapted Aedes aegypti. Taken together, it was likely that the present CHIKV outbreak in Thailand occurred as a result of the importation of the CHIKV strain from South Asia. Understanding with viral genetic diversity is essential for epidemiological study and may contribute to better disease management and preventive measures.

Arbovirus infection in Aedes aegypti has historically been quantified from a sample of the adult population by pooling collected mosquitoes to increase detectability. However, there is a significant knowledge gap about the magnitude of natural arbovirus infection within areas of active transmission, as well as the sensitivity of detection of such an approach. We used indoor Ae. aegypti sequential sampling with Prokopack aspirators to collect all mosquitoes inside 200 houses with suspected active ABV transmission from the city of Mérida, Mexico, and tested all collected specimens by RT-PCR to quantify: a) the absolute arbovirus infection rate in individually tested Ae. aegypti females; b) the sensitivity of using Prokopack aspirators in detecting ABV-infected mosquitoes; and c) the sensitivity of entomological inoculation rate (EIR) and vectorial capacity (VC), two measures ABV transmission potential, to different estimates of indoor Ae. aegypti abundance. The total number of Ae. aegypti (total catch, the sum of all Ae. aegypti across all collection intervals) as well as the number on the first 10-min of collection (sample, equivalent to a routine adult aspiration session) were calculated. We individually tested by RT-PCR 2,161 Aedes aegypti females and found that 7.7% of them were positive to any ABV. Most infections were CHIKV (77.7%), followed by DENV (11.4%) and ZIKV (9.0%). The distribution of infected Aedes aegypti was overdispersed; 33% houses contributed 81% of the infected mosquitoes. A significant association between ABV infection and Ae. aegypti total catch indoors was found (binomial GLMM, Odds Ratio > 1). A 10-min indoor Prokopack collection led to a low sensitivity of detecting ABV infection (16.3% for detecting infected mosquitoes and 23.4% for detecting infected houses). When averaged across all infested houses, mean EIR ranged between 0.04 and 0.06 infective bites per person per day, and mean VC was 0.6 infectious vectors generated from a population feeding on a single infected host per house/day. Both measures were significantly and positively associated with Ae. aegypti total catch indoors. Our findings provide evidence that the accurate estimation and quantification of arbovirus infection rate and transmission risk is a function of the sampling effort, the local abundance of Aedes aegypti and the intensity of arbovirus circulation., Author summary Aedes-borne diseases comprise a serious public health burden in many parts of the world, usually affecting low income areas. The ability to detect virus circulation within a population may be key in responding to the threat of outbreaks, providing a cost-effective approach for triggering vector control. Unfortunately, gaps in the knowledge of natural Aedes-borne virus (ABV) infection in Aedes aegypti have led to uncertainties in the consideration of arbovirus surveillance in mosquitoes. Here, we show that the natural infection rate in a mosquito population may not be a function of where Aedes aegypti are, but rather where key human-mosquito contacts occur. Sampling 200 houses with suspected ABV active transmission led us to quantify high virus infection rates in all Aedes aegypti present in the house and use such information to estimate the sensitivity of indoor aspiration with Prokopack devices and two measures of ABV transmission potential. Our findings provide evidence that the accurate quantification of arbovirus infection rate and transmission risk is a function of the sampling effort, the local abundance of Aedes aegypti and the intensity of arbovirus circulation. Results from this study are relevant to understand the value of virus testing of vector populations, and for the design of entomological endpoints relevant for epidemiological trials quantifying the impact of vector control on ABVs.

The p53 transcription factor plays a key role both in cancer and in the cell-intrinsic response to infections. The ORFEOME project hypothesized that novel p53-virus interactions reside in hitherto uncharacterized, unknown, or hypothetical open reading frames (orfs) of human viruses. Hence, 172 orfs of unknown function from the emerging viruses SARS-Coronavirus, MERS-Coronavirus, influenza, Ebola, Zika (ZIKV), Chikungunya and Kaposi Sarcoma-associated herpesvirus (KSHV) were de novo synthesized, validated and tested in a functional screen of p53 signaling. This screen revealed novel mechanisms of p53 virus interactions and two viral proteins KSHV orf10 and ZIKV NS2A binding to p53. Originally identified as the target of small DNA tumor viruses, these experiments reinforce the notion that all viruses, including RNA viruses, interfere with p53 functions. These results validate this resource for analogous systems biology approaches to identify functional properties of uncharacterized viral proteins, long non-coding RNAs and micro RNAs., Author summary New viruses are constantly emerging. The ORFEOME project was based on the hypothesis that every virus, regardless of its molecular makeup and biology should encode functions that intersect the p53 signaling network, since p53 guards the cell from genomic insults, of which depositing a foreign, viral nucleic acid is one. The result of the ORFEOME screen of proteins without any known function, of predicted open reading frames and of suspected non-coding RNAs is the identification of two viral proteins that interact with p53. The first one, orf10, is encoded by Kaposi Sarcoma-associated herpesvirus and the second one, NS2A, is encoded by the Zika virus.

Since the emergence of yellow fever in the Americas and the devastating 1918 influenza pandemic, biologists and clinicians have been drawn to human infecting viruses to understand their mechanisms of infection better and develop effective therapeutics against them. However, the complex molecular and cellular processes that these viruses use to infect and multiply in human cells have been a source of great concern for the scientific community since the discovery of the first human infecting virus. Viral disease outbreaks, such as the recent COVID-19 pandemic caused by a novel coronavirus, have claimed millions of lives and caused significant economic damage worldwide. In this study, we investigated the mechanisms of host-virus interaction and the molecular machinery involved in the pathogenesis of some common human viruses. We also performed a phylogenetic analysis of viral proteins involved in host-virus interaction to understand the changes in the sequence organization of these proteins during evolution for various strains of viruses to gain insights into the viral origin’s evolutionary perspectives.

Aedes aegypti and Culex pipiens complex mosquitoes are prolific vectors of arboviruses that are a global threat to human and animal health. Increased globalization and ease of travel have facilitated the worldwide dissemination of these mosquitoes and the viruses they transmit. To assess disease risk, we determined the frequency of arboviruses in western Kenyan counties bordering an area of high arboviral activity. In addition to pathogenic viruses, insect-specific flaviviruses (ISFs), some of which are thought to impair the transmission of specific pathogenic arboviruses, were also evaluated. We trapped mosquitoes in the short and long rainy seasons in 2018 and 2019 at livestock markets and hospitals. Mosquitoes were screened for dengue, chikungunya and other human pathogenic arboviruses, ISFs, and their blood-meal sources as determined by high-resolution melting analysis of (RT-)PCR products. Of 6,848 mosquitoes collected, 89% were trapped during the long rainy season, with A. aegypti (59%) and Cx. pipiens sensu lato (40%) being the most abundant. Most blood-fed mosquitoes were Cx. pipiens s.l. with blood-meals from humans, chicken, and sparrow (Passer sp.). We did not detect dengue or chikungunya viruses. However, one Culex poicilipes female was positive for Sindbis virus, 30 pools of Ae. aegypti had cell fusing agent virus (CFAV; infection rate (IR) = 1.27%, 95% CI = 0.87%-1.78%); 11 pools of Ae. aegypti had Aedes flavivirus (AeFV; IR = 0.43%, 95% CI = 0.23%-0.74%); and seven pools of Cx. pipiens s.l. (IR = 0.23%, 95% CI = 0.1%-0.45%) and one pool of Culex annulioris had Culex flavivirus. Sindbis virus, which causes febrile illness in humans, can complicate the diagnosis and prognosis of patients with fever. The presence of Sindbis virus in a single mosquito from a population of mosquitoes with ISFs calls for further investigation into the role ISFs may play in blocking transmission of other arboviruses in this region.

Mayaro virus (MAYV) is an arbovirus that is endemic to tropical forests in Central and South America, particularly within the Amazon basin. In recent years, concern has increased regarding MAYV’s ability to invade urban areas and cause epidemics across the region. We conducted a systematic literature review to characterise the evolutionary history of MAYV, its transmission potential, and exposure patterns to the virus. We analysed data from the literature on MAYV infection to produce estimates of key epidemiological parameters, including the generation time and the basic reproduction number, R0. We also estimated the force-of-infection (FOI) in epidemic and endemic settings. Seventy-six publications met our inclusion criteria. Evidence of MAYV infection in humans, animals, or vectors was reported in 14 Latin American countries. Nine countries reported evidence of acute infection in humans confirmed by viral isolation or reverse transcription-PCR (RT-PCR). We identified at least five MAYV outbreaks. Seroprevalence from population based cross-sectional studies ranged from 21% to 72%. The estimated mean generation time of MAYV was 15.2 days (95% CrI: 11.7–19.8) with a standard deviation of 6.3 days (95% CrI: 4.2–9.5). The per-capita risk of MAYV infection (FOI) ranged between 0.01 and 0.05 per year. The mean R0 estimates ranged between 2.1 and 2.9 in the Amazon basin areas and between 1.1 and 1.3 in the regions outside of the Amazon basin. Although MAYV has been identified in urban vectors, there is not yet evidence of sustained urban transmission. MAYV’s enzootic cycle could become established in forested areas within cities similar to yellow fever virus., Author summary Each year, diseases that are transmitted by mosquitoes cause substantial deaths and disability across the world. We performed a systematic literature review of Mayaro virus (MAYV) and estimated key epidemiological parameters that can be used to improve future outbreak response. We estimated the generation time and basic reproduction number for a historical outbreak. Our results suggest that the force-of-infection of MAYV in endemic settings is low. We did not find evidence of substantial urban transmission of MAYV. Nevertheless, similarities between MAYV and yellow fever virus epidemiology suggest that MAYV could emerge in urban areas. Local transmission of MAYV has never been reported outside of Central and South America. Our results highlight the need to continue monitoring emerging arboviruses in the Americas.

Chikungunya virus (CHIKV) is a mosquito-borne pathogen that is responsible for numerous large and geographical epidemics, causing millions of cases. However, there is no vaccine or therapeutics against CHIKV infection available. Interferon-alpha (IFN-α) has been shown to produce potent antiviral responses during viral infection. Herein we demonstrated the use of an adenovirus-vectored expressed mouse IFN-α (mDEF201) as a prophylactic and therapeutic treatment against CHIKV in vivo. 6-day-old BALB/c mice were pre- or post-treated intranasally with single dose of mDEF201 at 5 x 106 PFU per mouse and challenged with lethal dose of CHIKV. Complete survival protection was observed in mice upon a single dose of mDEF201 administration 1 days prior to virus challenge. Viral load in the serum and multiple organs were significantly reduced upon mDEF201 administration in a dose dependent manner as compare with adenovirus 5 vector placebo set. Histological analysis of the mice tissue revealed that mDEF201 could significantly reduce the tissue morphological abnormities, mainly infiltration of immune cells and muscle fibre necrosis caused by CHIKV infection. In addition, administration of mDEF201 at 6 hours post CHIKV challenge also showed promising inhibitory effect against viral replication and dissemination. In conclusion, single-dose of intranasal administration with mDEF201 as a prophylactic or therapeutic agent within 6 hours post CHIKV infection is highly protective against a lethal challenge of CHIKV in the murine model., Author summary Chikungunya has emerged as one of the most important arbovirus diseases of global health significance. Over the last decade, millions of CHIKV infections have been reported in over 80 countries across several continents. However, there is no approved vaccines or specific therapeutics available, development of antiviral strategies against CHIKV infection has become a pressing issue. In this study, the authors examined using 6-day-old BALB/c mice as an animal model to study CHIKV infection. After shown the 6-day-old mice were permissive and susceptible to CHIKV infection, the authors also evaluated the effectiveness of adenovirus-vectored mouse IFN-α (mDEF201) against systemic and lethal challenge of CHIKV infection in 6-day-old BALB/c mice. Single-dose of mDEF201 can protect mice against a lethal challenge of CHIKV. The authors believe that mDEF201 has immense clinical potential as a single dose could protect individuals at high risks during an outbreak. In addition, mDEF201 could also potentially be used to treat CHIKV infections especially during the acute phase.

Background Like many countries from the Americas, Cuba is threatened by Aedes aegypti-associated arboviruses such as dengue (DENV), Zika (ZIKV), and chikungunya (CHIKV) viruses. Curiously, when CHIKV was actively circulating in the region in 2013–2014, no autochthonous transmission of this virus was detected in Havana, Cuba, despite the importation of chikungunya cases into this city. To investigate if the transmission ability of local mosquito populations could explain this epidemiological scenario, we evaluated for the first time the vector competence of two Ae. aegypti populations (Pasteur and Párraga) collected from Havana for dengue virus type 1 (DENV-1), CHIKV, and ZIKV. Methodology/Principal findings Mosquito populations were fed separately using blood containing ZIKV, DENV-1, or CHIKV. Infection, dissemination, and transmission rates, were estimated at 3 (exclusively for CHIKV), 7, and 14 days post exposure (dpe) for each Ae. aegypti population-virus combination. Both mosquito populations were susceptible to DENV-1 and ZIKV, with viral infection and dissemination rates ranging from 24–97% and 6–67% respectively. In addition, CHIKV disseminated in both populations and was subsequently transmitted. Transmission rates were low (, Author summary Dengue (DENV), Zika (ZIKV), and chikungunya (CHIKV) viruses are mainly transmitted by Aedes aegypti mosquitoes and together caused near 10 million human cases in the last decade in the Americas. The diseases caused by these viruses share symptoms ranging from mild clinical manifestations to severe complications, including fatal outcomes. Cuba has not been exempted from these viruses and is frequently subjected to dengue and more recently by diseases caused by ZIKV infection. Surprisingly, despite multiple CHIKV outbreaks recorded in neighboring countries, local circulation has not been reported in Havana. Although Ae. aegypti has been implicated as the major DENV and ZIKV vector because of its wide distribution on the island, neither field nor experimental approaches have been used to demonstrate the ability of Cuban Ae. aegypti populations to transmit DENVs, ZIKV, and CHIKV. In this laboratory study, we demonstrated that two Ae. aegypti populations from Havana could become infected with and transmit these viruses, albeit at low rates. Our paper suggests that the absence of CHIKV circulation in Havana was not related to refractoriness of Ae. aegypti populations to this virus; therefore, vector control remains being essential in preventing future arboviruses outbreaks.

Introduction Dengue, Zika and Chikungunya are RNA Arboviruses present in some areas of Mexico, mainly in the endemic state of Chiapas that is characterized by presence of the vector that transmit them and an ecology that favors high transmission. According to the national epidemiological surveillance system, Dengue has intensified since 2018 and outbreaks continue in various states while for Zika and Chikungunya a decrease in cases has been reported in recent years. The main objective of this study was to determine the incidence of Dengue, Zika and Chikungunya infections during pregnancy in the state of Chiapas. Principal findings The presence of previous and current infections and coinfections diagnosed by molecular (RT-PCR) and immunological (ELISA for IgG determination) techniques indicates a wide circulation of viruses in asymptomatic people, specifically in pregnant women showing that silent infections in dry season contributes to the preservation of viruses. Conclusions From 136 studied samples, 27.7% tested positive for DENV, 8% for ZIKV and 24.1% for CHIKV by RTPCR and the values of IgG in sera show that 83.9% were positive for IgG antibodies against DENV, 65% against ZIKV and 59.1% against CHIKV. Results demonstrated presence of ZIKV and CHIKV, not detected by the epidemiological surveillance system, so the importance of establishing proactive epidemiological systems more strict, especially because these infections in pregnant women can cause severe health problems for newborn children., Author summary A study was conducted in a group of pregnant women from a region of southern Mexico where Dengue, Zika and Chikungunya are endemic to determine the presence of three viral infections, clinical manifestations and congenital damage in newborns. Epidemiological surveillance of diseases is mainly based on the incidence of these diseases; however, asymptomatic carriers also play an important role in disease transmission and are generally not included in the systems. In the case of the three arbovirosis studied, asymptomatic infections can occur until 60–70% of cases and play an important role in the disease transmission and level of protection of population. The three arboviruses, transmitted by the bite of the Aedes aegypti mosquito, in the same endemic scenario have the possibility of producing a high rate of asymptomatic infections and coinfections, which may go unnoticed. The study provides valuable information to improve knowledge of transmission in a period of low transmission (dry season) and also to understand that silent transmission has medical importance, especially in pregnancy, due the congenital damage that can cause to newborn. Therefore, it is important to design strategies to incorporate information into the national and international epidemiological surveillance system.