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Background: In this study, we aimed to investigate the specific-antibody response to the COVID-19 vaccination and the immunophenotyping of T cells in older adults who were engaged or not in an exercise training program before the pandemic. Methods: Ninety-three aged individuals (aged between 60 and 85 years) were separated into 3 groups: practitioners of physical exercise vaccinated with CoronaVac (PE-Co, n = 46), or vaccinated with ChadOx-1 (PE-Ch, n = 23), and non-practitioners vaccinated with ChadOx-1 (NPE-Ch, n = 24). Blood samples were collected before (pre) and 30 days after vaccination with the second vaccine dose. Results. Higher IgG levels and immunogenicity were found in the PE-Ch and NPE-Ch groups, whereas increased IgA levels were found only in the PE-Ch group post-vaccination. The PE-Co group showed a positive correlation between the IgA and IgG values, and lower IgG levels post-vaccination were associated with age. Significant alterations in the percentage of naive (CD28+CD57-), double-positive (CD28+CD57+), and senescent (CD28-CD57+) CD4+ T and CD8+ T cells were found post-vaccination, particularly in the PE-Ch group. Conclusions: The volunteers vaccinated with the ChadOx-1 presented not only a better antibody response but also a significant modulation in the percentage of T cell profiles, mainly in the previously exercised group.

Genetic variants of SARS-CoV-2 have been emerging and circulating in many places across the world. Rapid detection of these variants is essential since their dissemination can impact transmission rates, diagnostic procedures, disease severity, response to vaccines or patient management. Sanger sequencing has been used as the preferred approach for variant detection among circulating human immunodeficiency and measles virus genotypes. Using primers to amplify a fragment of the SARS-CoV-2 genome encoding part of the Spike protein, we showed that Sanger sequencing allowed us to rapidly detect the introduction and spread of three distinct SARS-CoV-2 variants in two major Brazilian cities. In both cities, after the predominance of variants closely related to the virus first identified in China, the emergence of the P.2 variant was quickly followed by the identification of the P1 variant, which became dominant in less than one month after it was first detected.

OBJECTIVE: COVID-19 is a systemic disease affecting multiple organs, beyond the respiratory system. Although sensorineural symptoms, including anosmia and ageusia, have been frequently associated with milder forms of SARS-CoV-2 infection, the pathogenic mechanism and its relation with oromucosal immunomodulation remains poorly understood. Therefore, the main objective of this pilot study was to provide some pieces of evidence concerning the cytokines pattern in the mucosa of the upper airways in COVID-19 patients with and without anosmia and/or ageusia. METHODS: Samples of nasopharyngeal/oropharyngeal swabs were obtained from 12 patients diagnosed with COVID-19. The patients were separated into two groups: with anosmia and/or ageusia (A/A,n=4), and a control group with patients without anosmia and/or ageusia (NA/A,n=8). RESULTS: Mucosal cytokines concentration was determined by ELISA. Levels of IFN-γ, IL-6, and IL-10 in the mucosa of the upper airways were similar between the groups. In addition, no significant differences were found in the ratio of IFN-γ/IL-6, IFN-γ/IL-10, and IL-10/IL-6 between the two groups. However, using Pearson's correlation coefficient we found a significant positive correlations between IL-6 and IFN-γ, IL-10 and IFN-γ, and also IL-10 and IL-6 in the control group (NA/A), whereas a significant negative correlation between IL-10 and IFN-γ was found in A/A group. CONCLUSION: Our results suggest that patients with mild forms of COVID-19, presenting anosmia and/or ageusia, have an altered mucosal immune response in the upper airways. DESCRIPTORS: SARS-CoV-2, Sensorineural symptoms, IL-6, IL-10, IFN-gamma.

Summary Objectives Although primarily transmitted by Aedes aegypti mosquitos, Zika virus (ZIKV) can also be transmitted by blood transfusion, due to the fact that some of the infected donors can establish asymptomatic viremia. ZIKV seroprevalence in Brazilian blood donors is unknown. The main reason for this gap in the knowledge originates from the difficulty in evaluating ZIKV humoral immunity due to antigenic cross-reactivity between the different Brazilian flaviviruses and, in particular, dengue virus (DENV). The objective of this study was to evaluate the anti-ZIKV IgG prevalence in blood donors from the Northeast region of the Sao Paulo State, Brazil, which experienced a ZIKV outbreak in 2016. Methods We evaluated the ZIKV seroprevalence using the NS1 anti-ZIKV IgG test (Euroimmun), followed by confirmation of the positive and borderline results using the Plaque Reduction Neutralization Test (PRNT). ZIKV seroprevalence was estimated by testing plasma samples collected in 2015 (before the ZIKV outbreak), 2016 (during the outbreak) and 2017 (after the outbreak). In order to investigate possible antigenic cross – reactivity between ZIKV and DENV we also included samples that were taken well before the ZIKV outbreak, in years 2010 and 2013. Results The results obtained by the Euroimmun anti-ZIKV IgG test demonstrated ZIKV seroreactivity in 2015, 2016, and 2017 with prevalences of 5.3%, 12.8% and 13.2%, respectively. The inclusion of blood donor samples from 2010 to 2013, demonstrated anti-ZIKV IgG reactivity only for 2013 (1.7%). The PRNT testing of the ZIKV positive and borderline ELISA reacting samples generated positive results only for the years of 2016 and 2017 (prevalences of 5.6% and 9.1%) which coincided with the introduction of ZIKV in our region. Conclusions Our results estimate for the first time the ZIKV seroprevalence among Brazilian blood donors from a region with apparently extensive ZIKV circulation and which, at the same time, is highly endemic for DENV. We detected relatively low ZIKV seroprevalence in blood donors from the studied region probably due to the lower intensity of the outbreak compared to other Brazilian locations. Our study adds to the global understanding of ZIKV circulation and the herd immunity of the exposed population.

Objectives: Defense against respiratory viruses depends on an immune response present in the mucosa, as saliva IgA secretes antibodies. During the pandemic, such as influenza or SARS-CoV-2, most infected patients are asymptomatic but retain specific antibodies post-infection. The authors evaluated IgG and IgA antibodies against SARS-CoV-2 and influenza in the saliva of asymptomatic volunteers, validated with controls or vaccinated individuals. Methods: The authors detected specific antibodies by validated conventional ELISA using natural SARS-CoV-2 antigens from infected Vero cells or capture-ELISA for influenza using natural antigens of the influenza vaccine. Results: Saliva from influenza-vaccinated individuals had more IgA than paired serum, contrary to the findings for specific IgG. In COVID-19-vaccinated samples, specific IgA in saliva increased after vaccination, but IgG levels were high after the first dose. In saliva from the asymptomatic population (226), anti-Influenza IgG was found in 57.5% (130) of samples, higher than IgA, found in 35% (79) of samples. IgA results were similar for SARS-CoV-2, with IgA present in 30% (68) of samples, while IgG was less present, in 44.2% (100) of samples. The proportion of influenza IgG responders was higher than that for SARS-CoV-2 IgG, but both populations presented similar proportions of IgA responders, possibly due to variable memory B cell survival. For both viruses, the authors found an important proportion (> 10%) of IgA+IgG- samples, suggesting the occurrence of humoral immunity directed to the mucosa. Conclusion: Specific antibodies for respiratory viruses in saliva are found in either infection or vaccination and are a convenient and sensitive diagnostic tool for host immune response.

SARS-CoV-2 papain-like protease (PLpro) covers multiple functions. Beside the cysteine-protease activity, PLpro has the additional and vital function of removing ubiquitin and ISG15 (Interferon-stimulated gene 15) from host-cell proteins to aid coronaviruses in evading the host’s innate immune responses. We established a high-throughput X-ray screening to identify inhibitors by elucidating the native PLpro structure refined to 1.42 Å and performing co-crystallization utilizing a diverse library of selected natural compounds. We identified three phenolic compounds as potential inhibitors. Crystal structures of PLpro inhibitor complexes, obtained to resolutions between 1.7-1.9 Å, show that all three compounds bind at the ISG15/Ub-S2 allosteric binding site, preventing the essential ISG15-PLpro molecular interactions. All compounds demonstrate clear inhibition in a deISGylation assay, two exhibit distinct antiviral activity and one inhibited a cytopathic effect in a non-cytotoxic concentration range. These results highlight the druggability of the rarely explored ISG15/Ub-S2 PLpro allosteric binding site to identify new and effective antiviral compounds. Importantly, in the context of increasing PLpro mutations in the evolving new variants of SARS-CoV-2, the natural compounds we identified may also reinstate the antiviral immune response processes of the host that are down-regulated in COVID-19 infections.

Brazil presents one of the highest COVID-19 death tolls in the world. The initial SARS-CoV-2 epicenter was São Paulo city. As of 2019, the homeless population of São Paulo city was estimated at 24,344 individuals, the largest national homeless population. The present study aimed to concomitantly assess the molecular and serological prevalence and associated risk factors of SARS-CoV-2 infection in a homeless population and related shelter workers from a day-shelter. Serum samples, nasopharyngeal and oropharyngeal swabs of persons who are homeless and shelter workers collected from August 25th to 27th, 2020 were tested for the presence of anti-SARS-CoV-2 IgM and IgG antibodies by ELISA and SARS-CoV-2 RNA by RT-qPCR, respectively. All swab samples tested negative by RT-qPCR. Seropositivity of IgM and IgG was 5/203 (2.5%) and 111/203 (54.7%) in persons who are homeless, and 5/87 (5.7%) and 41/87 (47.1%) in shelter workers, respectively, with no statistical differences between groups. The high seroprevalence found herein indicates early environmental and urban spreading of SARS-CoV-2, associated with sociodemographic and economic vulnerability., Author summary Brazil is one of the world’s most social-economically unequal countries, with a rising homeless population potentialized by the SARS-CoV-2 pandemic, particularly in its largest city, São Paulo. While few studies, mostly in high-income countries, have addressed the impact of the coronavirus pandemic on homeless populations, none has been carried out in Brazil. Herein, we report a high SARS-CoV-2 IgG seroprevalence in a homeless population, with 111/203 (54.7%) seropositive individuals. At the time of the study (August 25th- 27th 2020), both homeless and social worker populations showed no active SARS-CoV-2 infection, indicating that they were likely exposed sometime within the pandemic’s first peak in the city. Our study has also shown significant risk and protective factors for SARS-CoV-2 infection, including that Black shelter workers were at higher risk of SARS-CoV-2 infection when compared with the white shelter workers.

BackgroundCoronaVac is an inactivated SARS-CoV-2 vaccine that has been rolled out in several low and middle-income countries including Brazil, where it was the mainstay of the first wave of immunization of health care workers and the elderly population. We aimed to assess the T cell and antibody responses of vaccinees as compared to convalescent subjects.MethodsWe detected IgG against SARS-CoV-2 antigens, neutralizing antibodies against the original SARS-CoV-2 strain, and used SARS-CoV-2 peptides to detect IFN-g and IL-2 specific T cell responses in a cohort of CoronaVac vaccinees (N=101) and convalescent (N=72) individuals.FindingsAmong vaccinees, 95% displayed T cell or antibody responses to SARS-CoV-2 as compared to 99% convalescent individuals. However, we observed that among vaccinees, males and individuals 55 years or older developed significantly lower anti-RBD, anti-NP and neutralizing antibody responses as well as antigen-induced IL-2 production by T cells.InterpretationEven though some studies indicated Coronavac helped reduce mortality among elderly people, considering the current dominance of the gamma variant of concern (VOC) and potential increase of the delta VOC, in Brazil, our data support that Coronavac vaccinees above 55 years old Coronavac vaccinees above 55 years old could benefit from a heterologous third dose/booster vaccine to improve immune response and protection.FundingBrazilian Ministry for Science, Technology and Innovation, Sao Paulo State Foundation for Scientific research (FAPESP), JBS S.A.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the agent of coronavirus disease 2019 (COVID-19), is responsible for the worst pandemic of the 21st century. Like all human coronaviruses, SARS-CoV-2 originated in a wildlife reservoir, most likely from bats. As SARS-CoV-2 has spread across the globe in humans, it has spilled over to infect a variety of non-human animal species in domestic, farm, and zoo settings. Additionally, a broad range of species, including one neotropical monkey, have proven to be susceptible to experimental infection with SARS-CoV-2. Together, these findings raise the specter of establishment of novel enzootic cycles of SARS-CoV-2. To assess the potential exposure of free-living non-human primates to SARS-CoV-2, we sampled 60 neotropical monkeys living in proximity to Manaus and São José do Rio Preto, two hotspots for COVID-19 in Brazil. Our molecular and serological tests detected no evidence of SAR-CoV-2 infection among these populations. While this result is reassuring, sustained surveillance efforts of wildlife living in close association with human populations is warranted, given the stochastic nature of spillover events and the enormous implications of SARS-CoV-2 spillover for human health.

Zika virus (ZIKV) is a mosquito-borne viral disease associated with fetal microcephaly and other central nervous system (CNS) symptomatology. It was first identified in a Rhesus macaque in Uganda in 1947 and later in humans (Zika fever). In 2015, ZIKV was notified in Northeast Brazil where it was associated with CNS alterations and with rapid epidemic spread. Considering that ZIKV infects Old World monkeys, the aim of this study was to follow its potential in neotropical primates. Here, we show the detection of ZIKV in marmosets and capuchin monkeys captured in Ceara state, Northeast Brazil. Nine (9/132) samples were positive by quantitative RT-PCR assay. Neutralizing antibodies in primates for ZIKV were also detected by PRNT. The ZIKV-positive samples were obtained from peridomestic animals captured in proximity to humans in areas with reports of ZIKV-associated microcephaly cases during the epidemic period. These results reiterate the molecular evidence of ZIKV infection in neotropical primates, and the temporal detection suggests that detection in primates occurred during the epidemic period in humans. However, a continuous surveillance is necessary to exclude the possibility of virus circulation and transmission in wild environments.

This study aims to describe the sociodemographic determinants associated with exposure to Zika Virus (ZIKV) in pregnant women during the 2015–2016 epidemic in Salvador, Brazil. Methods We recruited women who gave birth between October 2015 and January 2016 to a cross-sectional study at a referral maternity hospital in Salvador, Brazil. We collected information on their demographic, socioeconomic, and clinical characteristics, and evaluated their ZIKV exposure using a plaque reduction neutralization test. Logistic regression was then used to assess the relationship between these social determinants and ZIKV exposure status. Results We included 469 pregnant women, of whom 61% had a positive ZIKV result. Multivariate analysis found that lower education (adjusted Prevalence Rate [aPR] 1.21; 95%CI 1.04–1.35) and food insecurity (aPR 1.17; 95%CI 1.01–1.30) were positively associated with ZIKV exposure. Additionally, age was negatively associated with the infection risk (aPR 0.99; 95%CI 0.97–0.998). Conclusion Eve after controlling for age, differences in key social determinants, as education and food security, were associated with the risk of ZIKV infection among pregnant women in Brazil. Our findings elucidate risk factors that can be targeted by future interventions to reduce the impact of ZIKV infection in this vulnerable population., Author summary The Zika virus (ZIKV) epidemic in Brazil has intensified global concern about congenital defects associated with intrauterine exposure. Social determinants are factors that reinforce and contribute to the transmission and spread of ZIKV as well as other arboviruses like Dengue. We performed a cross-sectional study to describe the prevalence of ZIKV and the contribution of social determinants to transmission among pregnant women during the 2015–2016 ZIKV epidemic in Salvador, Brazil. We found that 61% of pregnant women were ZIKV seropositive. We also found that lower education level, food insecurity and lower maternal age were associated with higher ZIKV infection risk. These findings contribute to understanding the role of social determinants in ZIKV transmission, providing key social factors that can be combined with pre-existing tactics (vector control and environmental improvement) to create policies and interventions which reduce social inequalities and risk of infection in vulnerable populations like pregnant women.

An unprecedented global health crisis has been caused by a new virus called severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). We performed experiments to test if a hypertonic saline solution was capable of inhibiting virus replication. Our data show that 1.2% NaCl inhibited virus replication by 90%, achieving 100% of inhibition at 1.5% in the nonhuman primate kidney cell line Vero, and 1.1% of NaCl was sufficient to inhibit the virus replication by 88% in human epithelial lung cell line Calu-3. Furthermore, our results indicate that the inhibition is due to an intracellular mechanism and not to the dissociation of the spike SARS-CoV-2 protein and its human receptor. NaCl depolarizes the plasma membrane causing a low energy state (high ADP/ATP concentration ratio) without impairing mitochondrial function, supposedly associated with the inhibition of the SARS-CoV-2 life cycle. Membrane depolarization and intracellular energy deprivation are possible mechanisms by which the hypertonic saline solution efficiently prevents virus replication in vitro assays.

Background Brazil became the epicenter of the COVID-19 pandemic in Latin America since May 2020, reporting the highest number of cases and deaths in the region. Healthcare workers (HCWs) are at increased risk of SARS-CoV-2 infection, experiencing a significant burden from COVID-19. Identifying and understanding the clinical characteristics and risk factors associated with infection are of paramount importance to inform screening strategies and infection control practices in this scenario. The aims of this study were to investigate the prevalence and clinical characteristics of HCWs with COVID-19 symptoms. Methods Between March 21st and May 22nd, 2020 a cross-sectional study was performed in a tertiary university hospital in São Paulo. Prevalence of SARS-CoV-2 infection among HCWs with COVID-19 symptoms was determined by RT-PCR testing on nasopharyngeal and oropharyngeal samples. Participants were asked to complete an electronic structured questionnaire including clinical and demographic data. Results Overall, 125 (42.37%) of 295 symptomatic HCWs tested positive for SARS-CoV-2. Over the 10-week study period, positivity rates varied from 22.2% (95% CI 15.9–60.3%) in the second week to 55.9% (95% CI 43.2–68.6%) in the sixth week, reaching a plateau (38–46%) thereafter. Median (SD) age was 34.2 (9.9) years and 205 (69.5%) were female. We did not find significant differences in the prevalence of the most commonly reported underlying medical condition among healthcare workers that tested positive or negative for SARS-CoV-2 infection. After multivariable analysis, using logistic regression, anosmia (adjusted OR 4.4 95% CI 2.21–8.74) and ocular pain (adjusted OR 1.95 95% CI 1.14–3.33) were the only symptoms independently associated with positivity for SARS-CoV-2 infection. Follow-up information on clinical outcomes showed that 9 (7.2%) HCWs were hospitalized (seven were male) and 2 (1.6%) died. Conclusions The findings of this study confirmed the high burden of SARS-CoV-2 infection among healthcare workers in the hardest hit city by the pandemic in Latin America. Anosmia and ocular pain were symptoms independently associated with COVID-19 diagnosis. In low and middle-income countries, where limited availability of tests is frequent, these findings may contribute to optimize a targeted symptom-oriented screening strategy.

Background: Since aging affects the immune responses against vaccination, the present study evaluated the effects of L-glutamine (Gln) supplementation in the humoral and cellular immune responses in elderly subjects, practitioners or not, of physical exercise training. Methods: Eighty-four elderly people (aged 72.6 &plusmn, 6.1), non-practitioners (NP, n = 31), and practitioners of combined-exercise training (CET, n = 53) were submitted to Influenza virus vaccination and supplemented with Gln (0.3 g/kg of weight + 10 g of maltodextrin, groups: NP-Gln (n = 14), and CET-Gln (n = 26)), or placebo (10 g of maltodextrin, groups: NP-PL (n = 17), and CET-PL (n = 27)). Blood samples were collected pre (baseline) and 30 days post-vaccination and supplementation. Results: Comparing with the baseline values, whereas the NP-Gln and CET-PL groups showed higher specific-IgM levels, the CET-Gln group showed higher specific-IgM and IgA levels post-vaccination. The titer rate of hemagglutination inhibition was higher in the CET-Gln, NP-PL, and NP-Gln groups post-vaccination than baseline values. The absolute number of naive and effector CD4+ T cells was higher especially in the NP-Gln and CET-Gln groups, whilst activated CD4+ T cells were higher in CET subgroups post-vaccination. Conclusion: Our results showed that both l-glutamine supplementation and combined-exercise training can improve the immune responses to the Influenza virus vaccine in elderly subjects.

Although first considered a benign infection, recent studies have disclosed severe and potentially lethal inflammatory manifestations of COVID-19 in children. We report the case of a 4-year-old child with a post-infectious multisystem inflammatory syndrome associated with COVID-19, with a Kawasaki-like shock and prominent neurologic features, for whom a cytokine storm and reduced brain-derived neurotrophic factor were well documented.

In March 2020, WHO declared a pandemic state due to SARS-CoV-2 having spread. TaqMan-based real-time RT-qPCR is currently the gold standard for COVID-19 diagnosis. However, it is a high-cost assay, inaccessible for the majority of laboratories around the world, making it difficult to diagnose on a large scale. The objective of this study was to standardize lower cost molecular methods for SARS-CoV-2 identification. E gene primers previously determined for TaqMan assays by Colman et al. (2020) were adapted in SYBR Green assay and RT-PCR conventional. The cross-reactivity test was performed with 17 positive samples for other respiratory viruses, and the sensibility test was performed with 8 dilutions (10 based) of SARS-CoV-2 isolated and 63 SARS-CoV-2-positive samples. The SYBR Green assays and conventional RT-PCR have not shown amplification of the 17 respiratory samples positives for other viruses. The SYBR Green-based assay was able to detect all 8 dilutions of the isolate. The conventional PCR detected until 107 dilution, both assays detected the majority of the 63 samples, 98.42% of positivity in SYBR Green, and 93% in conventional PCR. The average Ct variation between SYBR Green and TaqMan was 1.92 and the highest Ct detected by conventional PCR was 35.98. Both of the proposed assays are less sensitive than the current gold standard; however, our data shows a low sensibility variation, suggesting that these methods could be used by laboratories as a lower cost molecular method for SARS-CoV-2 diagnosis. Electronic supplementary material The online version of this article (10.1007/s42770-020-00347-5) contains supplementary material, which is available to authorized users.

We are facing an unprecedented global health crisis caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). At this date more than 680 thousand people have died due to coronavirus disease 2019 (COVID-19). Unfortunately, until now no effective treatment to combat the virus and vaccine are available. We performed experiments to test if hypertonic saline solution is able to inhibit virus replication in vitro. Our data shows that 260 mM NaCl (1.5%) inhibits 100% SARS-CoV-2 replication in Vero cells. Furthermore, our results suggest that the virus replication inhibition is due to an intracellular mechanism and not due to the dissociation between spike SARS-CoV-2 protein and its human receptor angiotensin-converting enzyme 2 interaction. NaCl depolarizes the plasma membrane supposedly associated with the inhibition of the SARS-CoV-2 life cycle. This observation could lead to simple, safe and low cost interventions at various stages of COVID-19 treatment, improving the prognosis of infected patients, thereby mitigating the social and economic costs of the pandemic.

Human cytomegalovirus (HCMV) infections remain a neglected public health issue. The aim of the present study was to evaluate the frequency of HCMV congenital infections in newborns up to 1 month in the Sao Paulo State, from 2010 to 2018. The molecular characterization of HCMV-positive samples was also undertaken. Urine samples from 275 potential congenital HCMV-infected patients were tested by real-time Polymerase Chain Reaction (qPCR). HCMV-positive samples were amplified by conventional PCR targeting the UL89 gene, sequenced and searched for mutations. A total of 32 (11.6%) positive-HCMV cases were detected (mean Ct 30.59); mean and median age of 10.3 and 6 days old, respectively. Children aged between 0-3 weeks had higher HCMV detection rates (84.4%; 27/32). UL89 gene was successfully sequenced in two samples, both classified as the human betaherpesvirus 5. No described resistance-associated mutations were identified. A routine screening in newborns coupled with the genetic characterization of key viral genes is vital to decrease sequels associated with congenital HCMV infections.

Objective: Respiratory syncytial virus is a pathogen frequently involved in nosocomial outbreaks. Although several studies have reported nosocomial outbreaks in neonatal intensive care units, molecular epidemiology data are scarce. Here, the authors describe two consecutive respiratory syncytial virus outbreaks caused by genotypes ON-1 and NA-2 in a neonatal intensive care unit in São Paulo, Brazil. Methods: A prospective search for respiratory syncytial virus was performed after diagnosing the index case and four other symptomatic newborns in the neonatal intensive care unit. Nasopharyngeal aspirate samples of all patients in the neonatal intensive care unit were tested for 17 respiratory viruses using real-time reverse transcriptase polymerase chain reaction. Genotyping was performed using nucleotide sequencing. Results: From May to August 2013, two different outbreaks were detected in the neonatal intensive care unit. A total of 20 infants were infected with respiratory syncytial virus-A (ten and 14 with ON-1 and NA-2 genotypes, respectively). The mean age of the infants was 10 days, mean birth weight was 1,961 g, and the mean gestational age was 33 weeks. Risk factors (heart disease, lung disease, and prematurity) were present in 80% and 85.7% of infants in the ON-1 and NA-2 groups, respectively. In total, 45.8% of infants were asymptomatic and 20.8% required mechanical ventilation. Coinfections were not detected during the outbreaks. Conclusions: Infants in a neonatal intensive care unit who develop abrupt respiratory symptoms should be tested for respiratory viruses, especially respiratory syncytial virus. Even in the absence of severe symptoms, respiratory syncytial virus detection can prevent nosocomial transmission through infection control measures. A better understanding of respiratory syncytial virus molecular epidemiology is essential for developing new vaccines and antiviral drugs against respiratory syncytial virus. Resumo: Objetivo: O vírus sincicial respiratório é um patógeno frequentemente envolvido em surtos nosocomiais. Embora vários estudos tenham relatado tais surtos em unidades de terapia intensiva neonatal, os dados epidemiológicos moleculares são escassos. Neste artigo, descrevemos dois surtos consecutivos de vírus sincicial respiratório causados pelos genótipos ON-1 e NA-2 em uma unidade de terapia intensiva neonatal em São Paulo, Brasil. Métodos: Uma busca prospectiva por vírus sincicial respiratório foi realizada após o diagnóstico do caso índice e outros quatro recém-nascidos sintomáticos na unidade de terapia intensiva neonatal. Amostras de aspirado nasofaríngeo de todos os pacientes da unidade de terapia intensiva neonatal foram testadas para 17 vírus respiratórios com reação em cadeia da polimerase via transcriptase reversa em tempo real. A genotipagem foi realizada utilizando sequenciamento de nucleotídeos. Resultados: De maio a agosto de 2013, foram detectados dois surtos diferentes na unidade de terapia intensiva neonatal. Vinte e quatro crianças foram infectadas com vírus sincicial respiratório-A (10 e 14 com os genótipos ON-1 e NA-2, respectivamente). A média da idade dos lactentes era de 10 dias, o peso médio ao nascer foi de 1961 g e a idade gestacional média de 33 semanas. Fatores de risco (doença cardíaca, doença pulmonar e prematuridade) estavam presentes em 80% e 85,7% dos bebês nos grupos ON-1 e NA-2, respectivamente. No total, 45,8% dos lactentes eram assintomáticos e 20,8% necessitaram de ventilação mecânica. Não foram detectadas coinfecções durante os surtos. Conclusões: Bebês em unidade de terapia intensiva neonatal que desenvolvem sintomas respiratórios abruptos devem ser testados para vírus respiratórios, especialmente o vírus sincicial respiratório. Mesmo na ausência de sintomas graves, a detecção de vírus sincicial respiratório pode prevenir a transmissão nosocomial através de medidas de controle de infecção. Um melhor entendimento da epidemiologia molecular do vírus sincicial respiratório é essencial para o desenvolvimento de novas vacinas e drogas antivirais contra o vírus sincicial respiratório. Keywords: Respiratory syncytial virus, ON-1, NA-2, Outbreak, Neonatal intensive care unit, Palavras-chave: Vírus sincicial respiratório, ON-1, NA-2, Surto, Unidade de terapia intensiva neonatal

BACKGROUND Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) was confirmed in Brazil in February 2020, the first cases were followed by an increase in the number of cases throughout the country, resulting in an important public health crisis that requires fast and coordinated responses. OBJECTIVES The objective of this work is to describe the isolation and propagation properties of SARS-CoV-2 isolates from the first confirmed cases of coronavirus disease 2019 (COVID-19) in Brazil. METHODS After diagnosis in patients that returned from Italy to the São Paulo city in late February by RT-PCR, SARS-CoV-2 isolates were obtained in cell cultures and characterised by full genome sequencing, electron microscopy and in vitro replication properties. FINDINGS The virus isolate was recovered from nasopharyngeal specimen, propagated in Vero cells (E6, CCL-81 and hSLAM), with clear cytopathic effects, and characterised by full genome sequencing, electron microscopy and in vitro replication properties. Virus stocks - viable (titre 2.11 × 106 TCID50/mL, titre 1.5 × 106 PFUs/mL) and inactivated from isolate SARS.CoV2/SP02.2020.HIAE.Br were prepared and set available to the public health authorities and the scientific community in Brazil and abroad. MAIN CONCLUSION We believe that the protocols for virus growth and studies here described and the distribution initiative may constitute a viable model for other developing countries, not only to help a rapid effective pandemic response, but also to facilitate and support basic scientific research.

Background: Pseudomonas aeruginosa (PS) infection results in severe morbidity and mortality, especially in immune-deficient populations. Aerobic exercise (AE) modulates the immune system, but its effects on the outcomes of pulmonary PS infection in elderly mice are unknown. Methods: BALB/c mice (24 weeks old) were randomized to sedentary, exercise (EX), PS, and PS + EX groups for the acute experimental setting, and PS and PS + EX groups for the chronic setting. Low-intensity AE was performed for 5 weeks, 60 min/day; 24 h after the final AE session, mice were inoculated with 5 × 104 colony-forming units (CFU) of PS, and 24 h and 14 days after PS inoculation, mice were studied. Results: AE inhibited PS colonization (p < 0.001) and lung inflammation (total cells, neutrophils, lymphocytes [p < 0.01] in bronchoalveolar lavage [BAL]), with significant differences in BAL levels of IL-1β (p < 0.001), IL-6 (p < 0.01), CXCL1 (p < 0.001), and TNF-α (p < 0.001), as well as parenchymal neutrophils (p < 0.001). AE increased BAL levels of IL-10 and parenchymal (p < 0.001) and epithelial (p < 0.001) IL-10 expression, while epithelial (p < 0.001) and parenchymal (p < 0.001) NF-κB expression was decreased. AE diminished pulmonary lipid peroxidation (p < 0.001) and increased glutathione peroxidase (p < 0.01). Pre-incubation of BEAS-2B with IL-10 inhibited PS-induced epithelial cell expression of TNF-α (p < 0.05), CD40 (p < 0.01), and dichlorodihydrofluorescein diacetate (p < 0.05). Conclusions: AE inhibits PS-induced lung inflammation and bacterial colonization in elderly mice, involving IL-10/NF-κB, and redox signaling.

Objective To describe the neurological and neurodevelopmental outcomes of children with Congenital Zika Syndrome (CZS) associated microcephaly beyond 2 years of age. Method We followed children with CZS-associated microcephaly in an outpatient clinic in Salvador, Brazil. Neurological and neurodevelopmental assessments were performed using the Hammersmith Infant Neurological Examination (HINE) and Bayley Scales of Infant and Toddler Neurodevelopment (Bayley-III) respectively. Results Of the 42 children included, 19 were male (45.2%); median (interquartile range) age at neurological evaluation was 28 (25–32) months, and 36 (85.7%) had severe microcephaly. HINE and Bayley-III results were completed for 35/42 (83.3%) and 33/42 (78.5%) children respectively. Bayley-III identified a severe developmental delay in 32/33 (97.0%) children while 1/33 (3.0%) had only a mild delay. In the multivariable analysis, we found that Bayley-III and HINE scores were correlated. Better HINE scores were associated with higher Bayley-III cognitive raw scores (β = 0.29; CI 95% = 0.02–0.57) and motor raw scores (β = 0.43; CI 95% = 0.04–0.82) after adjusting for head circumference, prematurity, and age at neurodevelopmental evaluation. Furthermore, we found that greater head circumference at follow up was associated with higher cognitive (β = 1.27; CI 95% = 0.01–2.53) and motor raw scores (β = 2.03; CI 95% = 0.25–3.81). Conclusion Children with CZS-associated microcephaly demonstrate severe neurodevelopmental delays and slower growth rates than their peers over time. Still, they have remarkably heterogeneous neurodevelopmental profiles according to neurological exam scores which correlate with their long-term outcomes. We found that HINE scores effectively captured the heterogeneity of neurological capabilities among these children and could be predictive of cognitive and motor development progress.

Yellow fever virus RNA is usually detected in blood of infected humans. We detected virus RNA in urine and semen samples from a convalescent patient. A complete virus genome was sequenced for an isolate from a urine sample. This virus had a South American I genotype and unique synapomorphic changes.

Rationale The chikungunya virus (CHIKV) was first isolated in a Tanzanian epidemic area between 1952 and 1953. The best description of the CHIKV transmission during pregnancy can be found in a well-documented epidemic in 2005, in the "La Reunion" island, a French territory located in the Indian Ocean, in which about one-third of the population was infected. Reports of arbovirus infections in pregnancy are increasing over time, but the spectrum of clinical findings remains an incognita among researchers, including CHIKV. Patient concerns In this report, it was possible to verify 2 cases exposed to CHIKV during foetal period and the possible implications of the infection on gestational structures and exposed children after the birth. Diagnosis In both cases, the mothers were positive by laboratory tests in serologic analysis for CHIKV, as ezyme-linked immunossorbent assay (ELISA), plaque reduction neutralisation testing (PRNT) and immunofluorescence (IF); but there were no positive tests in quantitative polymerase chain reaction (qPCR) for mothers or children. Interventions The exposed children were followed up in a paediatrics clinic in order not only to provide the medical assistance, but also to verify child development and the possible implications and neurocognitive changes caused by gestational infection. Outcomes There were neurological and developmental changes in one of the children followed up on an outpatient basis. There was an improvement in the neurological situation and symptoms only 3 years and 1 month after birth. Lessons Based on the cases presented, we can conclude that clinical symptoms of CHIKV maternal infection may occur late in new-borns and can affect their development.

Background: Although glutamine is able to improve the immune response, its action in the upper airway immunity against the influenza virus vaccine remains unclear. Therefore, we aimed to evaluate the L-glutamine supplementation effect on the mucosal immune/inflammatory response of elderly subjects vaccinated against the influenza virus. Methods: Saliva sampling from 83 physically active elderly volunteers were collected pre- and 30 days after influenza virus vaccination and supplementation with L-glutamine (Gln, n = 42) or placebo (PL, n = 41). Results: Gln group showed higher salivary levels of interleukin (IL)-17, total secretory immunoglobulin A (SIgA), and specific-SIgA post-vaccination than values found pre-vaccination and in the PL group post-vaccination. Whereas higher salivary levels of IL-6 and IL-10 were observed post-vaccination in the Gln group, IL-37 levels were lower post-vaccination in both groups than the values pre-vaccination. Tumor necrosis factor (TNF)-α levels were unchanged. Positive correlations between IL-6 and IL-10 were found in all volunteer groups pre- and post-vaccination and also between IL-17 and IL-6 or IL-10 in the Gln group post-vaccination. A negative correlation between IL-37 and IL-10 was found pre- and post-vaccination in the PL group. Conclusion: Gln supplementation was able to modulate salivary cytokine profile and increase SIgA levels, both total and specific to the influenza virus vaccine, in physically active elderly subjects.

Objective: Characterize the role of human parainfluenza virus and its clinical features in Brazilian children under 2 years of age presenting with acute lower respiratory tract infections. Methods: Real-time assays were used to identify strains of human parainfluenza virus and other common respiratory viruses in nasopharyngeal aspirates. One thousand and two children presenting with acute lower respiratory tract illnesses were enrolled from February 2008 to August 2010. Results: One hundred and four (10.4%) patients were human parainfluenza virus positive, of whom 60 (57.7%) were positive for human parainfluenza virus-3, 30 (28.8%) for human parainfluenza virus-4, 12 (11.5%) for human parainfluenza virus-1, and two (1.9%) for human parainfluenza virus-2. Seven (6.7%) patients had more than one strain of human parainfluenza virus detected. The most frequent symptoms were tachypnea and cough, similar to other viral respiratory infections. Clinical manifestations did not differ significantly between human parainfluenza virus-1, -2, -3, and -4 infections. Human parainfluenza virus-1, -3, and -4 were present in the population studied throughout the three years of surveillance, with human parainfluenza virus-3 being the predominant type identified in the first two years. Conclusion: Human parainfluenza viruses contribute substantially to pediatric acute respiratory illness (ARI) in Brazil, with nearly 30% of this contribution attributable to human parainfluenza virus-4. Resumo: Objetivo: Caracterizar o papel do VPH-4 e suas características clínicas em crianças brasileiras com menos de dois anos de idade com infecções agudas do trato respiratório inferior. Métodos: Ensaios em tempo real foram utilizados para identificar tipos de VPH e outros vírus respiratórios comuns em aspirados nasofaríngeos. Mil e duas crianças com doença aguda do trato respiratório inferior foram inscritas para participar de fevereiro de 2008 a agosto de 2010. Resultados: 104 (10,4%) pacientes eram VPH positivos, dos quais 60 (57,7%) eram positivos para VPH-3, 30 (28,8%) para VPH-4, 12 (11,5%) para VPH-1 e dois (1,9%) para VPH-2. Sete (6,7%) pacientes apresentaram mais de um tipo de VPH detectado. Os sintomas mais frequentes foram tosse e taquipneia, semelhantes a outras infecções respiratórias virais. As manifestações clínicas não diferiram de forma significativa entre as infecções por VPH-1, -2, -3 e -4. Os VPH-1, -3 e -4 estavam presentes na população estudada ao longo dos três anos de vigilância, e o VPH-3 foi o tipo predominante identificado nos primeiros dois anos. Conclusão: Os VPHs contribuem substancialmente para a DRA pediátrica no Brasil com quase 30% dessa contribuição atribuível ao VPH-4. Keywords: Human parainfluenza virus, Respiratory virus, Pediatric patients, Acute respiratory illness, Human respirovirus 4, Palavras-chave: Vírus da parainfluenza humana, Vírus respiratório, Pacientes pediátricos, Doença respiratória aguda, Respirovírus humano 4