Search

Thuy-Huong Ta-Tang,1 Sergio LB Luz,2 James L Crainey,2 José M Rubio3 1Malaria and NTDs Laboratory, National Centre of Tropical Medicine, Instituto de Salud Carlos III, Madrid, Spain; 2Laboratório de Ecologia de Doenças Transmissíveis na Amazônia, Instituto Leônidas e Maria Deane, Fundação Oswaldo Cruz, Manaus, Amazonas State, Brazil; 3Malaria & Emerging Parasitic Diseases Laboratory, National Microbiology Center, Instituto de Salud Carlos III, Madrid, SpainCorrespondence: José M RubioMalaria & Emerging Parasitic Diseases Laboratory, National Microbiology Center, Instituto de Salud Carlos III, Cra. Majadahonda-Pozuelo Km 2,5, Majadahonda, Madrid, 28220, SpainTel +34918223420Fax +34915097034Email jmrubio@isciii.esJames L CraineyLaboratório de Ecologia de Doenças Transmissíveis na Amazônia, Instituto Leônidas e Maria Deane, Fundação Oswaldo Cruz, 476 Rua Terezina, Adrianópolis, Manaus, Amazonas State, CEP 69057-070, BrazilEmail james.lee@fiocruz.brAbstract: Mansonellosis is caused by three filarial parasite species from the genus Mansonella that commonly produce chronic human microfilaraemias: M. ozzardi, M. perstans and M. streptocerca. The disease is widespread in Africa, the Caribbean and South and Central America, and although it is typically asymptomatic it has been associated with mild pathologies including leg-chills, joint-pains, headaches, fevers, and corneal lesions. No robust mansonellosis disease burden estimates have yet been made and the impact the disease has on blood bank stocks and the monitoring of other filarial diseases is not thought to be of sufficient public health importance to justify dedicated disease management interventions. Mansonellosis´s Ceratopogonidae and Simuliidae vectors are not targeted by other control programmes and because of their small size and out-door biting habits are unlikely to be affected by interventions targeting other disease vectors like mosquitoes. The ivermectin and mebendazole-based mass drug administration (iMDA and mMDA) treatment regimens deployed by the WHO´s Elimination of Neglected Tropical Diseases (ESPEN) programme and its forerunners have, however, likely impacted significantly on the mansonellosis disease burden, principally by reducing the transmission of M. streptocerca in Africa. The increasingly popular plan of using iMDA to control malaria could also affect M. ozzardi parasite prevalence and transmission in Latin America in the future. However, a potentially far greater mansonellosis disease burden impact is likely to come from short-course curative anti-Wolbachia therapeutics, which are presently being developed for onchocerciasis and lymphatic filariasis treatment. Even if the WHO´s ESPEN programme does not choose to deploy these drugs in MDA interventions, they have the potential to dramatically increase the financial and logistical feasibility of effective mansonellosis management. There is, thus, now a fresh and urgent need to better characterise the disease burden and eco-epidemiology of mansonellosis so that effective management programmes can be designed, advocated for and implemented.Keywords: Mansonellosis, Mansonella perstans, Mansonella ozzardi, Mansonella streptocerca, Wolbachia, doxycycline

Thuy-Huong Ta-Tang,1 James L Crainey,2 Rory J Post,3,4 Sergio LB Luz,2 José M Rubio1 1Malaria and Emerging Parasitic Diseases Laboratory, National Microbiology Center, Instituto de Salud Carlos III, Majadahonda, Spain; 2Laboratory of Infectious Disease Ecology in the Amazon, Oswaldo Cruz Foundation, Instituto Leônidas e Maria Deane, Manaus, Brazil; 3School of Natural Sciences and Psychology, John Moores University, Liverpool, 4Department of Disease Control, Faculty of Infectious Tropical Diseases, London School of Hygiene and Tropical Medicine, London, UK Abstract: Mansonellosis is a filarial disease caused by three species of filarial (nematode) parasites (Mansonella perstans, Mansonella streptocerca, and Mansonella ozzardi) that use humans as their main definitive hosts. These parasites are transmitted from person to person by bloodsucking females from two families of flies (Diptera). Biting midges (Ceratopogonidae) transmit all three species of Mansonella, but blackflies (Simuliidae) are also known to play a role in the transmission of M. ozzardi in parts of Latin America. M. perstans and M. streptocerca are endemic in western, eastern, and central Africa, and M. perstans is also present in the neotropical region from equatorial Brazil to the Caribbean coast. M. ozzardi has a patchy distribution in Latin America and the Caribbean. Mansonellosis infections are thought to have little pathogenicity and to be almost always asymptomatic, but occasionally causing itching, joint pains, enlarged lymph glands, and vague abdominal symptoms. In Brazil, M. ozzardi infections are also associated with corneal lesions. Diagnosis is usually performed by detecting microfilariae in peripheral blood or skin without any periodicity. There is no standard treatment at present for mansonellosis. The combination therapy of diethylcarbamazine plus mebendazole for M. perstans microfilaremia is presently one of the most widely used, but the use of ivermectin has also been proven to be very effective against microfilariae. Recently, doxycycline has shown excellent efficacy and safety when used as an antimicrobial against endosymbiotic Wolbachia bacteria harbored by some strains of M. perstans and M. ozzardi. Diethylcarbamazine and ivermectin have been used effectively to treat M. streptocerca infection. There are at present no estimates of the disease burden caused by mansonellosis, and thus its importance to many global health professionals and policy makers is presently limited to how it can interfere with diagnostic tools used in modern filarial disease control and elimination programs aimed at other species of filariae. Keywords: mansonellosis, filariasis, M. perstans, M. ozzardi, M. streptocerca, neglected disease

Competing Interests: The authors have declared that no competing interests exist.

Following the successful eradication of Wuchereria bancrofti , there are now just three species of conventional microfilaremic human filarial parasites endemic to the Brazilian Amazon region: Mansonella ozzardi , Mansonella perstans and Onchocerca volvulus. The zoonotic filarial parasite Dirofilaria immitis is also found in the Amazon region as are several sylvatic filarial parasites, some of which have been recorded causing zoonoses and some of which have never been recorded outside the region. Onchocerca volvulus is only found in the Amazonia onchocerciasis focus in the Brazilian state of Roraima where it affects the people of the Yanomami tribe living around the densely forested Venezuela border region. Mansonella ozzardi is by far the most common filarial parasite in Brazil and has a broad but patchy distribution throughout the western Amazon region. Recorded in the Brazilian states of Acre, Roraima, Matto Grosso, and within almost every municipality of Amazonas state, it is believed that pollution of the urban stream and river systems prevents the development of the simuliid vectors of M. ozzardi and explains the parasite's reduced distribution within urban areas and an absence of recent reports from the state capital Manaus. Decades of WHO-led periodic ivermectin treatment of Yanomami tribe's people have resulted in the partial suppression of O. volvulus transmission in this focus and has also probably affected the transmission of M. ozzardi in the region. Mansonella perstans , O. volvulus and very probably M. ozzardi infections can all be treated and most likely cured with a 4-6-week treatment course of doxycycline. The Brazilian Ministry of Health does not, however, presently recommend any treatment for mansonellosis infections and thus parasitic infections outside the Amazonia focus are typically left untreated. While the long treatment courses required for doxycycline-based mansonellosis therapies preclude their use in control programmes, new fast-acting filarial drug treatments are likely to soon become available for the treatment of both onchocerciasis and mansonellosis in the Amazon region. Filarial disease management in the Brazilian Amazon is thus likely to become dramatically more viable at a time when the public health importance of these diseases is increasingly being recognized., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2023 The Authors.)

Introduction: Four species of the Mansonella genus infect millions of people across sub-Saharan Africa and Central and South America. Most infections are asymptomatic, but mansonellosis can be associated with nonspecific clinical manifestations such as fever, headache, arthralgia, and ocular lesions ( M. ozzardi ); pruritus, arthralgia, abdominal pain, angioedema, skin rash, and fatigue ( M. perstans and perhaps Mansonella sp. 'DEUX'); and pruritic dermatitis and chronic lymphadenitis ( M. perstans )., Areas Covered: We searched the PubMed and SciELO databases for publications on mansonelliasis in English, Spanish, Portuguese, or French that appeared until 1 May 2023. Literature data show that anthelmintics - single-dose ivermectin for M. ozzardi , repeated doses of mebendazole alone or in combination with diethylcarbamazine (DEC) for M. perstans , and DEC alone for M. streptocerca - are effective against microfilariae. Antibiotics that target Wolbachia endosymbionts, such as doxycycline, are likely to kill adult worms of most, if not all, Mansonella species, but the currently recommended 6-week regimen is relatively impractical. New anthelmintics and shorter antibiotic regimens (e.g. with rifampin) have shown promise in experimental filarial infections and may proceed to clinical trials., Expert Opinion: We recommend that human infections with Mansonella species be treated, regardless of any apparent clinical manifestations. We argue that mansonellosis, despite being widely considered a benign infection, may represent a direct or indirect cause of significant morbidity that remains poorly characterized at present.

The canine filarial parasite Dirofilaria immitis has not been reported in Brazil´s Amazonas state capital, Manaus, for over a century. Here, we report one imported and 27 autochthonous D. immitis infections from a microfilarial survey of 766 domestic dog blood samples collected between 2017 and 2021 in Manaus. An Overall prevalence estimate of 15.44% (23/149) was calculated from our two rural collection sites; a prevalence of 1.22% (4/328) was estimated at our periurban collection site, and an overall prevalence of 0.35% (1/289) was calculated from our two urban clinic collections. Our data suggest that in the urban areas of Manaus, where the parasites are very likely vectored by the same species of mosquito that historically vectored Wuchereria bancrofti (Culex quinquefasciatus), prevalence levels are very low and possibly maintained by an influx from rural areas where sylvatic reservoirs and/or more favorable vector transmission dynamics maintain high prevalences.

Objective: To assess the emergent zoonotic disease risk posed by the voracious human-biting blackfly species Simulium oyapockense in the peripheral regions of an expanding urban centre situated deep in the Brazilian Amazon rainforest., Methods: We performed nine human landing catches at three periurban sites surrounding the Brazilian Amazon town of São Gabriel da Cachoeira. Using the detection of non-human primate filarial parasites as an indicator of the zoonotic disease threat posed by a biting insect, we screened 3328 S. oyapockense blackflies for the presence of zoonotic filarial DNA with an ITS-1 PCR assay and Sanger sequencing., Results: Between 98 and 100% of the biting insects captured during our nine collections were identified as S. oyapockense; at our three collection sites and during our three seasonally-distinct collections this species was captured at rates between 28 and 294 blackflies per hour. PCR screening of the march-collected S. oyapockense detected infectious-stage (L3) Mansonella mariae parasites (which are only known to infect non-human primates) in >0.15% of the tested head samples., Conclusions: Our results show that residents of the periurban regions of São Gabriel da Cachoeira are routinely exposed to the bites of S. oyapockense blackflies which have previously fed on non-human primates., (Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)

Mansonella ozzardi and Mansonella perstans infections both cause mansonellosis but are usually treated differently. Using a real-time polymerase chain reaction assay and deep sequencing, we reveal the presence of mansonellosis coinfections that were undetectable by standard diagnostic methods. Our results confirm mansonellosis coinfections and have important implications for the disease's treatment and diagnosis., (© The Author(s) 2020. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.)

As the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) pandemic continues to expand, healthcare resources globally have been spread thin. Now, the disease is rapidly spreading across South America, with deadly consequences in areas with already weakened public health systems. The Amazon region is particularly susceptible to the widespread devastation from Coronavirus disease 2019 (COVID-19) because of its immunologically fragile native Amerindian inhabitants and epidemiologic vulnerabilities. Herein, we discuss the current situation and potential impact of COVID-19 in the Amazon region and how further spread of the epidemic wave could prove devastating for many Amerindian people living in the Amazon rainforest., Competing Interests: The authors have declared that no competing interests exist.

In the past 5-10 years, Venezuela has faced a severe economic crisis, precipitated by political instability and declining oil revenue. Public health provision has been affected particularly. In this Review, we assess the impact of Venezuela's health-care crisis on vector-borne diseases, and the spillover into neighbouring countries. Between 2000 and 2015, Venezuela witnessed a 359% increase in malaria cases, followed by a 71% increase in 2017 (411 586 cases) compared with 2016 (240 613). Neighbouring countries, such as Brazil, have reported an escalating trend of imported malaria cases from Venezuela, from 1538 in 2014 to 3129 in 2017. In Venezuela, active Chagas disease transmission has been reported, with seroprevalence in children (<10 years), estimated to be as high as 12·5% in one community tested (n=64). Dengue incidence increased by more than four times between 1990 and 2016. The estimated incidence of chikungunya during its epidemic peak is 6975 cases per 100 000 people and that of Zika virus is 2057 cases per 100 000 people. The re-emergence of many vector-borne diseases represents a public health crisis in Venezuela and has the possibility of severely undermining regional disease elimination efforts. National, regional, and global authorities must take action to address these worsening epidemics and prevent their expansion beyond Venezuelan borders., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Background: Standard human landing catches (sHLCs) have historically been a key component of Onchocerca volvulus transmission monitoring, but expose health-workers to potentially hazardous vector bites. Novel human-bait-free trapping methods have been developed, but do not always work where they are needed and may not generate O. volvulus surveillance data that is directly comparable with historic data., Methodology: Simuliid sHLCs and mineral-oil protected HLCs (mopHLCs) were performed in a rural village of Amazonas state, Brazil. A four-hour direct comparisons of sHLCs and mopHLCs was carried-out using six vector collectors, each of whom used one leg for a sHLC and one for a mopHLC. Two-person collection teams then exclusively performed either mopHLCs or sHLCs for a further set of 12 four-hour collections. Following the completion of all collections, simuliid-bite mark estimates were made from legs used exclusively in sHLCs and legs used exclusively in mopHLCs., Principal Findings: All of the 1669 captured simuliids were identified as the O. volvulus vector Simulium oyapockense. Overall, mopHLC simuliids captured per hour (S/H) rates were lower than those obtained with sHLC trapping (15.5 S/H versus 20 S/H). Direct comparisons of simuliid capture rates found that vector-collectors captured simuliids significantly more efficiently ([Formula: see text]: 20.5 S/H) with mopHLC trapping than with sHLC trapping ([Formula: see text]: 16.4 S/H): P-value = 0.002. MopHLCs performed in isolation were, however, observed to capture vectors less efficiently ([Formula: see text]: 13.4 S/H) than sHLCs performed under similar conditions ([Formula: see text]: 19.98 S/H). All six vector collectors had significantly higher simuliid capture per counted bite mark (SC/CBM) rates using mopHLCs than they were observe to have using sHLCs ([Formula: see text]: 21 SC/CBM versus [Formula: see text]: 1 SC/CBM; p-value = 0.03125)., Conclusions: Vector collectors captured significantly more simuliids per counted bite mark with mopHLCs than with sHLCs. Further investigations into the utility of mopHLCs for onchocerciasis xenomonitoring and beyond are merited., Competing Interests: On behalf of all authors, I declare that none of us have no competing interests relating to the publication of this manscript.

Despite the broad distribution of M. ozzardi in Latin America and the Caribbean, there is still very little DNA sequence data available to study this neglected parasite's epidemiology. Mitochondrial DNA (mtDNA) sequences, especially the cytochrome oxidase (CO1) gene's barcoding region, have been targeted successfully for filarial diagnostics and for epidemiological, ecological and evolutionary studies. MtDNA-based studies can, however, be compromised by unrecognised mitochondrial pseudogenes, such as Numts. Here, we have used shot-gun Illumina-HiSeq sequencing to recover the first complete Mansonella genus mitogenome and to identify several mitochondrial-origin pseudogenes. Mitogenome phylogenetic analysis placed M. ozzardi in the Onchocercidae "ONC5" clade and suggested that Mansonella parasites are more closely related to Wuchereria and Brugia genera parasites than they are to Loa genus parasites. DNA sequence alignments, BLAST searches and conceptual translations have been used to compliment phylogenetic analysis showing that M. ozzardi from the Amazon and Caribbean regions are near-identical and that previously reported Peruvian M. ozzardi CO1 reference sequences are probably of pseudogene origin. In addition to adding a much-needed resource to the Mansonella genus's molecular tool-kit and providing evidence that some M. ozzardi CO1 sequence deposits are pseudogenes, our results suggest that all Neotropical M. ozzardi parasites are closely related.

Human filariae are vector-borne parasites and the causative agents of various diseases, including human onchocerciasis and lymphatic filariasis. Onchocerciasis causes a spectrum of cutaneous and ophthalmologic manifestations (including blindness) and has long been a major public health problem in Bioko Island (Equatorial Guinea). Bioko Island has been included in the WHO's Onchocerciasis Control Program since 1987. In Bioko Island, the specificity and sensitivity of clinical Onchocerca volvulus diagnosis is key. The objective of this work was to update onchocerciasis elimination progress in Bioko Island, after 18 years of mass ivermectin intervention, and the general filariasis situation through a rapid and accurate molecular method. A cross-sectional study was conducted in Bioko Island from mid-January to mid-February 2014. A total of 543 subjects were included in the study. Whole blood and one skin snip (from lumbar regions) were analysed with a real time PCR assay. Two other skin biopsies were analysed by an expert microscopist. All positive samples were confirmed by sequencing. Traditional microscopic examination of the skin biopsies failed to detect any microfilariae. However, 11 (2.03%) infections were detected using PCR assay, including one O. volvulus, two Mansonella streptocerca, seven Mansonella perstans and one Loa loa infections. PCR assays in blood detected 52 filariae-positive individuals (9.6%) which harboured M. perstans or L. loa. The low prevalence of O. volvulus confirms the success of the Onchocerciasis Control Programme and suggests that Mass Drug Administration in Bioko Island can be interrupted in the near future. The very high prevalence of M. perstans found in skin snips assays raises doubts about the reliability of microscope-based diagnosis of O. volvulus infections., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Research interest in Wolbachia is growing as new discoveries and technical advancements reveal the public health importance of both naturally occurring and artificial infections. Improved understanding of the Wolbachia bacteriophages (WOs) WOcauB2 and WOcauB3 [belonging to a sub-group of four WOs encoding serine recombinases group 1 (sr1WOs)], has enhanced the prospect of novel tools for the genetic manipulation of Wolbachia . The basic biology of sr1WOs, including host range and mode of genomic integration is, however, still poorly understood. Very few sr1WOs have been described, with two such elements putatively resulting from integrations at the same Wolbachia genome loci, about 2 kb downstream from the FtsZ cell-division gene. Here, we characterize the DNA sequence flanking the FtsZ gene of w Dam, a genetically distinct line of Wolbachia isolated from the West African onchocerciasis vector Simulium squamosum E. Using Roche 454 shot-gun and Sanger sequencing, we have resolved >32 kb of WO prophage sequence into three contigs representing three distinct prophage elements. Spanning ≥36 distinct WO open reading frame gene sequences, these prophage elements correspond roughly to three different WO modules: a serine recombinase and replication module (sr1RRM), a head and base-plate module and a tail module. The sr1RRM module contains replication genes and a Holliday junction recombinase and is unique to the sr1 group WOs. In the extreme terminal of the tail module there is a SpvB protein homolog-believed to have insecticidal properties and proposed to have a role in how Wolbachia parasitize their insect hosts. We propose that these w Dam prophage modules all derive from a single WO genome, which we have named here sr1WOdamA1. The best-match database sequence for all of our sr1WOdamA1-predicted gene sequences was annotated as of Wolbachia or Wolbachia phage sourced from an arthropod. Clear evidence of exchange between sr1WOdamA1 and other Wolbachia WO phage sequences was also detected. These findings provide insights into how Wolbachia could affect a medically important vector of onchocerciasis, with potential implications for future control methods, as well as supporting the hypothesis that Wolbachia phages do not follow the standard model of phage evolution.

We obtained ribosomal and mitochondrial DNA sequences from residents of Amazonas state, Brazil, with Mansonella parasitemias. Phylogenetic analysis of these sequences confirm that M. ozzardi and M. perstans parasites occur in sympatry and reveal the close relationship between M. perstans in Africa and Brazil, providing insights into the parasite's New World origins.

Background: Multi-drug resistant forms of Pseudomonas aeruginosa (MDRPA) are a major source of nosocomial infections and when discharged into streams and rivers from hospital wastewater treatment plants (HWWTP) they are known to be able to persist for extended periods. In the city of Manaus (Western Brazilian Amazon), the effluent of three HWWTPs feed into the urban Mindu stream which crosses the city from its rainforest source before draining into the Rio Negro. The stream is routinely used by Manaus residents for bathing and cleaning (of clothes as well as domestic utensils) and, during periods of flooding, can contaminate wells used for drinking water., Results: 16S rRNA metagenomic sequence analysis of 293 cloned PCR fragments, detected an abundance of Pseudomonas aeruginosa (P. aeruginosa) at the stream's Rio Negro drainage site, but failed to detect it at the stream's source. An array of antimicrobial resistance profiles and resistance to all 14 tested antimicrobials was detected among P. aeruginosa cultures prepared from wastewater samples taken from water entering and being discharged from a Manaus HWWTP. Just one P. aeruginosa antimicrobial resistance profile, however, was detected from cultures made from Mindu stream isolates. Comparisons made between P. aeruginosa isolates' genomic DNA restriction enzyme digest fingerprints, failed to determine if any of the P. aeruginosa found in the Mindu stream were of HWWTP origin, but suggested that Mindu stream P. aeruginosa are from diverse origins. Culturing experiments also showed that P. aeruginosa biofilm formation and the extent of biofilm formation produced were both significantly higher in multi drug resistant forms of P. aeruginosa., Conclusions: Our results show that a diverse range of MDRPA are being discharged in an urban stream from a HWWTP in Manaus and that P. aeruginosa strains with ampicillin and amikacin can persist well within it.

Background: Vector-biting behaviour is important for vector-borne disease (VBD) epidemiology. The proportion of blood meals taken on humans (the human blood index, HBI), is a component of the biting rate per vector on humans in VBD transmission models. Humans are the definitive host of Onchocerca volvulus, but the simuliid vectors feed on a range of animals and HBI is a key indicator of the potential for human onchocerciasis transmission. Ghana has a diversity of Simulium damnosum complex members, which are likely to vary in their HBIs, an important consideration for parameterization of onchocerciasis control and elimination models., Methods: Host-seeking and ovipositing S. damnosum (sensu lato) (s.l.) were collected from seven villages in four Ghanaian regions. Taxa were morphologically and molecularly identified. Blood meals from individually stored blackfly abdomens were used for DNA profiling, to identify previous host choice. Household, domestic animal, wild mammal and bird surveys were performed to estimate the density and diversity of potential blood hosts of blackflies., Results: A total of 11,107 abdomens of simuliid females (which would have obtained blood meal(s) previously) were tested, with blood meals successfully amplified in 3,772 (34 %). A single-host species was identified in 2,857 (75.7 %) of the blood meals, of which 2,162 (75.7 %) were human. Simulium soubrense Beffa form, S. squamosum C and S. sanctipauli Pra form were the most anthropophagic (HBI = 0.92, 0.86 and 0.70, respectively); S. squamosum E, S. yahense and S. damnosum (sensu stricto) (s.s.)/S. sirbanum were the most zoophagic (HBI = 0.44, 0.53 and 0.63, respectively). The degree of anthropophagy decreased (but not statistically significantly) with increasing ratio of non-human/human blood hosts. Vector to human ratios ranged from 139 to 1,198 blackflies/person., Conclusions: DNA profiling can successfully identify blood meals from host-seeking and ovipositing blackflies. Host choice varies according to sibling species, season and capture site/method. There was no evidence that HBI is vector and/or host density dependent. Transmission breakpoints will vary among locations due to differing cytospecies compositions and vector abundances.

There are two species of filarial parasites with sheathless microfilariae known to commonly cause parasitaemias in humans: Mansonella perstans and Mansonella ozzardi. In most contemporary accounts of the distribution of these parasites, neither is usually considered to occur anywhere in the Eastern Hemisphere. However, Sir Patrick Manson, who first described both parasite species, recorded the existence of sheathless sharp-tailed Mansonella ozzardi-like parasites occurring in the blood of natives from New Guinea in each and every version of his manual for tropical disease that he wrote before his death in 1922. Manson's reports were based on his own identifications and were made from at least two independent blood sample collections that were taken from the island. Pacific region Mansonella perstans parasitaemias were also later (in 1923) reported to occur in New Guinea and once before this (in 1905) in Fiji. Although Mansonella-parasitaemias are generally regarded as benign, they are thought to be of public health importance because they can affect the epidemiological monitoring of other filarial diseases. In this article, we reviewed the historic literature concerning Pacific-origin Mansonella-parasitaemias in an attempt to explain how, despite repeated reports of Pacific-region Mansonella-parasitaemias, by as early as the 1970s, the WHO had arrived at the present-day view that Wuchereria bancrofti is the only cause of filarial parasitaemias in Papua New Guinea. We have also evaluated the evidence supporting the contemporary existence of Pacific-area parasitaemia-causing Mansonella parasites and assessed the relevance such parasites could have for present-day lymphatic filariasis elimination efforts in the region., (Copyright © 2016 Hainan Medical College. Production and hosting by Elsevier B.V. All rights reserved.)

We report here the first complete mitochondria genome of Onchocerca volvulus from a focus outside of Africa. An O. volvulus mitogenome from the Brazilian Amazonia focus was obtained using a combination of high-throughput and Sanger sequencing technologies. Comparisons made between this mitochondrial genome and publicly available mitochondrial sequences identified 46 variant nucleotide positions and suggested that our Brazilian mitogenome is more closely related to Cameroon-origin mitochondria than West African-origin mitochondria. As well as providing insights into the origins of Latin American onchocerciasis, the Brazilian Amazonia focus mitogenome may also have value as an epidemiological resource.

Background: Chromobacterium violaceum (C. violaceum) occurs abundantly in a variety of ecosystems, including ecosystems that place the bacterium under stress. This study assessed the adaptability of C. violaceum by submitting it to nutritional and pH stresses and then analyzing protein expression using bi-dimensional electrophoresis (2-DE) and Maldi mass spectrometry., Results: Chromobacterium violaceum grew best in pH neutral, nutrient-rich medium (reference conditions); however, the total protein mass recovered from stressed bacteria cultures was always higher than the total protein mass recovered from our reference culture. The diversity of proteins expressed (repressed by the number of identifiable 2-DE spots) was seen to be highest in the reference cultures, suggesting that stress reduces the overall range of proteins expressed by C. violaceum. Database comparisons allowed 43 of the 55 spots subjected to Maldi mass spectrometry to be characterized as containing a single identifiable protein. Stress-related expression changes were noted for C. violaceum proteins related to the previously characterized bacterial proteins: DnaK, GroEL-2, Rhs, EF-Tu, EF-P; MCP, homogentisate 1,2-dioxygenase, Arginine deiminase and the ATP synthase β-subunit protein as well as for the ribosomal protein subunits L1, L3, L5 and L6. The ability of C. violaceum to adapt its cellular mechanics to sub-optimal growth and protein production conditions was well illustrated by its regulation of ribosomal protein subunits. With the exception of the ribosomal subunit L3, which plays a role in protein folding and maybe therefore be more useful in stressful conditions, all the other ribosomal subunit proteins were seen to have reduced expression in stressed cultures. Curiously, C. violeaceum cultures were also observed to lose their violet color under stress, which suggests that the violacein pigment biosynthetic pathway is affected by stress., Conclusions: Analysis of the proteomic signatures of stressed C. violaceum indicates that nutrient-starvation and pH stress can cause changes in the expression of the C. violaceum receptors, transporters, and proteins involved with biosynthetic pathways, molecule recycling, energy production. Our findings complement the recent publication of the C. violeaceum genome sequence and could help with the future commercial exploitation of C. violeaceum.

Background: Mansonella ozzardi is a poorly understood human filarial parasite with a broad distribution throughout Latin America. Most of what is known about its parasitism has come from epidemiological studies that have estimated parasite incidence using light microscopy. Light microscopy can, however, miss lighter, submicroscopic, infections. In this study we have compared M. ozzardi incidence estimates made using light microscopy, with estimates made using PCR., Methods: 214 DNA extracts made from Large Volume Venous Blood Samples (LVVBS) were taken from volunteers from two study sites in the Rio Solimões region: Codajás [n = 109] and Tefé [n = 105] and were subsequently assayed for M. ozzardi parasitism using a diagnostic PCR (Mo-dPCR). Peripheral finger-prick blood samples were taken from the same individuals and used for microscopic examination. Finger-prick blood, taken from individuals from Tefé, was also used for the creation of FTAcard dried blood spots (DBS) that were subsequently subjected to Mo-dPCR., Results: Overall M. ozzardi incidence estimates made with LVVBS PCRs were 1.8 times higher than those made using microscopy (44.9% [96/214] compared with 24.3% [52/214]) and 1.5 times higher than the PCR estimates made from FTAcard DBS (48/105 versus 31/105). PCR-based detection of FTAcard DBS proved 1.3 times more sensitive at diagnosing infections from peripheral blood samples than light microscopy did: detecting 24/105 compared with 31/105. PCR of LVVBS reported the fewest number of false negatives, detecting: 44 of 52 (84.6%) individuals diagnosed by microscopy; 27 of 31 (87.1%) of those diagnosed positive from DBSs and 17 out of 18 (94.4%) of those diagnosed as positive by both alternative methodologies., Conclusions: In this study, Mo-dPCR of LVVBS was by far the most sensitive method of detecting M. ozzardi infections and detected submicroscopic infections. Mo-dPCR FTAcard DBS also provided a more sensitive test for M. ozzardi diagnosis than light microscopy based diagnosis did and thus in settings where only finger-prick assays can be carried-out, it may be a more reliable method of detection. Most existing M. ozzardi incidence estimates, which are often based on light microscope diagnosis, are likely to dramatically underestimate true M. ozzardi parasitism incidence levels.

In a recent issue of Memórias do Instituto Oswaldo Cruz, published in Rio de Janeiro in February 2014 (109: 87-92), Adami et al. have published a survey reporting Mansonella parasite prevalence in the Amazon Region. This report makes a useful contribution to the existing knowledge of filarial parasite distribution within the Amazon area, parasite prevalence rates in relation to age and occupation and provides observations on the possible clinical impact of Mansonella ozzardi. Their publication also provides an account of what appears to be a novel ELISA that has recently been used in the Simuliidae and Onchocerciasis Laboratory of the Oswaldo Cruz Institute, Rio de Janeiro, Brazil. We are concerned that the publication of this ELISA may have created an excessively positive impression of the effectiveness of the onchocerciasis recrudescence serological surveillance tools that are presently available for use in the Amazonia onchocerciasis focus. In this letter we have, thus, sought to highlight some of the limitations of this ELISA and suggest how continuing insecurities concerning the detection of antibodies to Onchocerca volvulus within the Amazonia onchocerciasis focus might be minimised.

Following the success of the Onchocerciasis Elimination Programme for the Americas (OEPA), there is now just one Latin American onchocerciasis focus where onchocerciasis transmission is described as 'on-going:' the Amazonia Onchocerciasis focus. In the hyperendemic highland areas of the Amazonia focus, Simulium guianense s.l. Wise are the most important vectors of the disease. Populations of S. guianense s.l. are, however, known to vary in their cytogenetics and in a range of behaviours, including in their biting habits. In the hypoendemic lowland areas of the Amazonia focus, for example, S. guianense s.l. are generally regarded as zoophilic and consequently unimportant to disease transmission. Robust tools, to discriminate among various populations of S. guianense s.l. have, however, not yet been developed. In the work reported here, we have assessed the utility of a ribosomal DNA sequence fragment spanning the nuclear ribosomal ITS-1, ITS-2 and 5.8S sequence regions and a ∼850 nucleotide portion of the mitochondrial cytochrome oxidase gene (CO1) for species-level identification and for resolving the within species substructuring. We report here how we have generated 78 CO1 sequences from a rich set of both zoophilic and anthropophilic populations of S. guianense s.l. that were collected from eight sites that are broadly distributed across Brazil. Consistent with previous findings, our analysis supports the genetic isolation of Simulium litobranchium from S. guianense s.l. In contrast with previous findings, however, our results did not provide support for the divergence of the two species prior to the radiation of S. guianense s.l. In our analysis of the S. guianense s.l. ribosomal DNA sequence trace files we generated, we provide clear evidence of multiple within-specimen single nucleotide polymorphisms and indels suggesting that S. guianense s.l. ribosomal DNA is not a good target for conventional DNA barcoding. This is the first report of S. guianense s.l. within individual ribosomal DNA variation and thus the first evidence that the species is not subject to the normal effects of concerted evolution. Collectively, these data illustrate the need for diverse sampling in the development of robust molecular tools for vector identification and suggest that ribosomal DNA might be able to assist with resolving S. guianense s.l. species substructuring that C01 barcoding has hitherto failed to., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Background: Immunoassays for Plasmodium detection are, presently, most frequently based on monoclonal antibodies (MAbs); Polyclonal antibodies (PAbs), which are cheaper to develop and manufacture, are much less frequently used. In the present study we describe a sandwich ELISA assay which is capable of detecting P. vivax Lactate Dehydrogenase (LDH) in clinical blood samples, without cross reacting with those infected with P. falciparum., Methods: Two recombinant proteins were produced from different regions of the P. vivax LDH gene. Two sandwich ELISA assay were then designed: One which uses mouse anti-LDH 1-43aa PAbs as primary antibodies ("Test 1") and another which uses anti-LDH 35-305aa PAbs ("Test 2") as the primary antibodies. Rabbit anti-LDH 1-43aa PAbs were used as capture antibodies in both ELISA assays. Blood samples taken from P. vivax and P. falciparum infected patients (confirmed by light microscopy) were analysed using both tests., Results: "Test 2" performed better at detecting microscopy-positive blood samples when compared to "Test 1", identifying 131 of 154 positive samples (85%); 85 positives (55%) were identified using "test 1". "Test 1" produced one false positive sample (from the 20 malaria-free control) blood samples; "test 2" produced none. Kappa coefficient analysis of the results produced a value of 0.267 when microscope-positive blood smears were compared with "test 1", but 0.734 when microscope-positive blood smears were compared with the results from "test 2". Positive predictive value (PPV) and negative predictive value (NPV) were observed to be 98% and 22% respectively, for "Test 1", and 99% and 45%, for "test 2". No cross reactivity was detected with P. falciparum positive blood samples (n = 15) with either test assay., Conclusion: Both tests detected P. vivax infected blood and showed no evidence of cross-reacting with P. falciparum. Further studies will need to be conducted to establish the full potential of this technique for malaria diagnostics. As well as representing a promising new cost-effective novel technique for P. vivax diagnosis and research, the method for developing this assay also highlights the potential for PAb-based strategies for diagnostics in general.

The Amazonia onchocerciasis focus of southern Venezuela and northern Brazil is the larger of the two remaining Latin American onchocerciasis foci where disease transmission still occurs and is often regarded as the most challenging of all the Latin American foci to eliminate onchocerciasis. The site is home to a population of over 20,000 semi-nomadic, hunter-gatherer Yanomami people and is made-up of a mosaic of rainforest and savannah ecologies, which are influenced by the area's undulating terrain and rich geological diversity. At least six blackfly vectors have been implicated in onchocerciasis transmission in this focus; however, because of the difficulty in their routine identification the relative importance of each has been obscured. Simulium limbatum and Simulium incrustatum s.l. have both been recorded as vectors in the Amazonia focus, but they are difficult to discriminate morphologically and thus the ecological range of these species, and indeed the presence of S. limbatum in the Amazonia focus at all, have remained controversial. In the work described here, we report 15 S. incrustatum s.l. CO1 sequences and 27 S. limbatum sequences obtained from field-caught adult female blackflies collected from forest and savannah localities, inside and just outside the Amazonia focus. Phylogenetic analysis with the sequences generated in this study, showed that both the S. limbatum and the S. incrustatum s.l. CO1 sequences obtained (even from specimens living in sympatry) all fell into discrete species-specific bootstrap-supported monophyletic groups and thus confirmed the utility of the CO1 gene for identifying both these species inside the Amazonia focus. As the S. limbatum-exclusive cluster included CO1 sequences obtained from forest-caught and morphologically identified specimens these results provide the clearest evidence yet of the presence of S. limbatum inside the Amazonia focus. The question, however, of whether S. limbatum is actually a vector in the focus still remains unanswered as the data presented here also suggest that S. limbatum found in the savannahs adjacent to, but outside the Amazonia focus (and which represent the only S. limbatum population to be unambiguously incriminated as a host of Onchocerca volvulus), are genetically distinct from those living inside the focus. These findings highlight the need for a clearer picture of the vector taxonomy inside the Amazonia onchocerciasis focus., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Background: Simulium damnosum s.l., the most important vector of onchocerciasis in Africa, is a complex of sibling species that have been described on the basis of differences in their larval polytene chromosomes. These (cyto) species differ in their geographical distributions, ecologies and epidemiological roles. In Ghana, distributional changes have been recorded as a consequence of vector control and environmental change (e.g. deforestation), with potential disease consequences. We review the distribution of cytospecies in southern Ghana and report changes observed with reference to historical data collated from 1971 to 2005 and new identifications made between 2006 and 2011., Methods/results: Larvae were collected from riverine breeding sites, fixed in Carnoy's solution and chromosome preparations made. Cytotaxonomic identifications from 1,232 samples (including 49 new samples) were analysed. We report long-term stability in cytospecies distribution in the rivers Afram, Akrum, Pawnpawn and Pru. For the rivers Oda, Ofin and Tano we describe (for the first time) patterns of distribution. We could not detect cytospecies composition changes in the upper Pra, and the lower Pra seems to have been stable. The elimination of the Djodji form of S. sanctipauli in the Volta Region seems to have had no long-term effects on the distribution of the other cytospecies, despite an initial surge by S. yahense. There has been a recent increase in the occurrence of savannah cytospecies in the river Asukawkaw, and this might be related to continuing deforestation., Conclusions: Cytospecies' distributions have not been stable from 1971 to 2011. Although there are no obvious causes for the temporary appearance and subsequent disappearance of cytospecies in a particular location, a major influence has been vector control and migration patterns, probably explaining observed changes on the Black Volta and lower Volta rivers. Deforestation was previously implicated in an increase of savannah cytospecies in southern Ghana (1975-1997). Our data had little power to support (or refute) suggestions of a continuing increase, except in the Asukawkaw river basin.

Although approximately 40% of all the people blinded by Onchocerca volvulus are Nigerians, almost nothing was known about the various cytospecies of the blackfly vectors present in Nigeria until 1981. The activation of the Nigerian National Onchocerciasis Control Programme in 1986 (and that programme's initiation of mass distributions of ivermectin in 1991) provided a significant stimulus to understand the biology of the Nigerian vectors but the exploration of any possible differences between the cytospecies has been hampered by a lack of accessible taxonomic information. This review attempts to satisfy that need. There are nine different cytoforms reliably recorded from Nigeria (Simulium damnosum s.s. Nile form, S. damnosum s.s. Volta form, S. sirbanum Sirba form, S. sirbanum Sudanense form, S. soubrense Beffa form, S. squamosum A, S. squamosum B, S. squamosum C and S. yahense typical form), and three more are known from surrounding countries and might be reasonably expected to occur in Nigeria. All of these cytospecies are presumed to be vectors, although there have been almost no identifications of the vectors of O. volvulus in Nigeria. The biogeographical distribution of the cytoforms is broadly similar to that known in other parts of West Africa (although many of the cytoforms remain insufficiently studied). The physico-chemical hydrology of the Nigerian breeding sites of the cytospecies does not, however, correspond to that seen elsewhere in West Africa, and it is not clear whether this might be related to differences in the cytoforms. An illustrated cytotaxonomic key is presented to facilitate and encourage future studies.

A Bacterial Artificial Chromosome (BAC) library was made from wild-caught Simulium squamosum, which is an important vector of human onchocerciasis. The library is composed of 12,288 BACs, with an average insert size of 128 kb, and is expected to contain ~1.54 GB of cloned DNA. Random BAC-end sequencing generated over 95 kb of DNA sequence data from which putative S. squamosum gene sequences and novel repetitive DNA families were identified, including DNA transposons, retrotransposons and simple sequence repeats (SSRs). The sequence survey also provided evidence of DNA of microbial origin, and dissection of sample blackflies indicated that some of those used to prepare the library were likely to be parasitized by the mermithid Isomermis lairdi. Hybridisations with a set of three independent blackfly single-copy genes and two Wolbachia genes suggest that the library provides around 13-fold coverage of the S. squamosum genome and about 12-fold coverage of its Wolbachia endosymbiont., (Crown Copyright © 2010. Published by Elsevier Inc. All rights reserved.)

Wolbachia are intracellular bacteria mostly found in a diverse range of arthropods and filarial nematodes. They have been classified into seven distinct 'supergroups' and other lineages on the basis of molecular phylogenetics. The arthropod-infecting Wolbachia are usually regarded as reproductive parasites because they manipulate their host species' sexing system to enhance their own spread, and this has led to their investigation as potential agents of genetic control in medical entomology. We report 12 partial Wolbachia gene sequences from: aspC, aspS, dnaA, fbpA, ftsZ, GroEL, hcpA, IDA, rpoB, rpe, TopI and wsp as well as a single ftsZ pseudogene sequence, which have all been PCR-amplified from Simulium squamosum (Diptera: Simuliidae). To our knowledge this is the first such report from Simuliidae. Uninterrupted open-reading frame sequences were obtained from all 12 genes, covering approximately 6.2kb of unique DNA sequence. Phylogenetic analyses with the different coding genes gave consistent results suggesting that the Wolbachia sequences obtained here do not derive from any of the known Wolbachia supergroups or lineages. Consistent with a unique genetic status for the S. squamosumWolbachia, the hypervariable regions of the Wolbachia-specific wsp gene were distinct from all previous records in both sequence and length. As well as potential implications for newly emerging Wolbachia-based disease control methods, the results may be relevant to some problems experienced in the laboratory colonisation of Simulium damnosum sensu lato and why it is such a diverse species complex., (2009. Published by Elsevier Ltd. All rights reserved.)

The mermithid parasite, Isomermis lairdi Mondet, Poinar & Bernadou (Nematoda: Mermithidae), is known to have a major impact on populations of Simulium damnosum s.l. Theobald (Diptera: Simuliidae) and on their efficiency as vectors of Onchocerca volvulus (Leuckart) (Nematoda: Filarioidea). However, the value of I. lairdi and other mermithid parasites as potential means of integrated vector control has not been fully realized. This is partly because traditional taxonomic approaches have been insufficient for describing and analysing important aspects of their biology and host range. In total, rDNA barcode sequences have been obtained from over 70 I. lairdi mermithids found parasitizing S. damnosum s.l. larvae in three different rivers. No two sequences were found to vary by more than 0.5%, and cytospecies identification of mermithid hosts revealed that I. lairdi with identical rDNA barcodes can parasitize multiple cytoforms of the S. damnosum complex, including S. squamosum (Enderlein). Phylogenetic analysis using a partial sequence from the 18S ribosomal DNA barcode, grouped I. lairdi in a monophyletic group with Gastromermis viridis Welch (Nematoda: Mermithidae) and Isomermis wisconsinensis Welch (Nematoda: Mermithidae).

The population biology of internal parasites is difficult to study because the adult parasites are often inaccessible, deep within the host's body. Developing stages, such as eggs in the faeces or larvae in the skin are more easily obtained, but are difficult to handle because they are often very small and with a tough cuticle. This has limited their use in molecular ecology for estimating population biology parameters of the adults (their parents). We have used Onchocerca ochengi (a filarial nematode parasite of cattle) to describe a novel and generally applicable method of easily and conveniently isolating individual larvae (microfilariae) from the host using laser-assisted microdissection. Furthermore, we have been able to improve the isolation of DNA by using the laser to bisect the larva to release DNA from the tissues enclosed within the parasite cuticle, and in this way we have achieved amplification of fragments over 1400 bp, and routinely PCR-amplified single-copy sequences from 5% of the DNA from a single larva (the equivalent of approximately 15 nuclei), and regularly from 0.5%., (© 2009 The Authors. Journal compilation © 2009 Blackwell Publishing Ltd.)

The detection of horizontal transfer is important to understanding the origin and spread of transposable elements and in assessing their impact on genetic diversity. The occurrence of the phenomenon is not in doubt for two of the three major groups of elements, but is disputed for retroposons, largely on the grounds of data paucity and overreliance on divergence estimates between host species. We present here the most wide-ranging retroposon data set assembled to date for a species group, the mosquitoes. The results provide no evidence for horizontal transfer events and show conclusively that four previously reported events, involving Juan-A, Juan-C, T1, and Q, did not occur. We propose that the origin of all known mosquito retroposons can be attributed to vertical inheritance and that retroposons have therefore been a persistent source of genetic diversity in mosquito genomes since the emergence of the taxon. Furthermore, the data confirm that the unprecedented levels of retroposon diversity previously reported in Anopheles gambiae extends to at least seven other species representing five genera and all three mosquito subfamilies. Most notably, this included the L1 elements, which are not known in other insects. A number of novel well-defined monophyletic groups were also identified, particularly, JM2 and JM3 within the Jockey clade, which included sequences from seven and five mosquito species, respectively. As JM3 does not contain an Anopheles element, this represents a good example of stochastic loss and the best out of at least four found in this study. This exceptionally diverse data set when compared with the wealth of data available for the many unrelated species with which mosquitoes have intimate contact through blood feeding ought to be fertile ground for the discovery of horizontal transfer events. The absence of positive results therefore supports the view that retroposon horizontal transfer does not occur or is far more exceptional than for other types of transposable elements.