Implementing a provisional overarching intervention for COVID-19 monitoring and control in the Brazil-Colombia-Peru frontier.

Introduction: he challenge was to provide comprehensive health resources to a remote and underserved population living in the Brazil-Colombia-Peru border, amid the most disruptive global crisis of the century. Methods: In August 2021, Fundação Oswaldo Cruz Amazonia (FIOCRUZ Amazônia) and partner collaborators implemented an overarching provisional program for SARS-CoV-2 detection and lineages characterization, training of laboratory personnel and healthcare providers, donation of diagnostic supplies and personal protective equipment, and COVID-19 vaccination. The expedition was conducted at the Port of Tabatinga, a busy terminal with an intense flux of people arriving and departing in boats of all sizes, located in the Amazon River basin. Local government, non-profit organizations, private companies, and other stakeholders supported the intervention. Results: The expedition was accomplished in a convergence point, where migrant workers, traders, army personnel, people living in urban areas, and people from small villages living in riversides and indigenous territories are in close and frequent contact, with widespread cross-border movement. Using a boat as a provisional lab and storage facility, the intervention provided clinical and laboratory monitoring for 891 participants; vaccination for 536 individuals; personal protective equipment for 200 healthcare providers; diagnostic supplies for 1,000 COVID-19 rapid tests; training for 42 community health agents on personal protection, rapid test execution, and pulse oximeter management; and hands-on training for four lab technicians on molecular diagnosis. Discussion: Our experience demonstrates that multilateral initiatives can counterweigh the scarcity of health resources in underserved regions. Moreover, provisional programs can have a long-lasting effect if investments are also provided for local capacity building.

Molecular detection of Mansonella mariae incriminates Simulium oyapockense as a potentially important bridge vector for Amazon-region zoonoses.

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.

Correction: SARS-CoV-2 in the Amazon region: A harbinger of doom for Amerindians.

[This corrects the article DOI: 10.1371/journal.pntd.0008686.].

Distribution and diversity of mosquitoes and Oropouche-like virus infection rates in an Amazonian rural settlement.

Mosquito diversity and disease transmission are influenced by landscape modifications, i.e., vectors and pathogens previously found only in forests are now found close to human environments due to anthropic changes. This study determined the diversity and distribution of mosquitoes in forest environments in order to analyze the potential vectors of Amazonian forest arboviruses. Mosquitoes were collected by 1) vertical stratification from forest canopy and ground areas using Hooper Pugedo (HP) light traps and human attraction and 2) horizontal stratification using HP light traps in peridomicile, forest edge, and forest environments near the Rio Pardo rural settlement, Amazonas, Brazil. A total of 3,750 mosquitoes were collected, representing 46 species. 3,139 individuals representing 46 species were sampled by vertical stratification. Both the Shannon-Weaver diversity index (H') and equitability (J') were higher in the canopy than on the ground. 611 individuals representing 13 species were sampled by horizontal stratification. H' decreased in the following order: forest edge > forest > peridomicile, and J' was greater at the forest edge and smaller in the peridomicile environment. Moreover, H' was higher for the human attraction collection method than the HP traps. A total of 671 pools were analyzed by RT-qPCR; three species were positive for Oropouche-like viruses (Ochlerotatus serratus, Psorophora cingulata, and Haemagogus tropicalis) and the minimum infection rate was 0.8%. The composition of mosquito species did not differ significantly between anthropic and forest environments in Rio Pardo. Some mosquito species, due to their abundance, dispersion in the three environments, and record of natural infection, were hypothesized to participate in the arbovirus transmission cycle in this Amazonian rural settlement.

Genomic and phylogenetic characterisation of an imported case of SARS-CoV-2 in Amazonas State, Brazil.

A new coronavirus [severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)] is currently causing a life-threatening pandemic. In this study, we report the complete genome sequencing and genetic characterisation of a SARS-CoV-2 detected in Manaus, Amazonas, Brazil, and the protocol we designed to generate high-quality SARS-CoV-2 full genome data. The isolate was obtained from an asymptomatic carrier returning from Madrid, Spain. Nucleotide sequence analysis showed a total of nine mutations in comparison with the original human case in Wuhan, China, and support this case as belonging to the recently proposed lineage A.2. Phylogeographic analysis further confirmed the likely European origin of this case. To our knowledge, this is the first SARS-CoV-2 genome obtained from the North Brazilian Region. We believe that the information generated in this study may contribute to the ongoing efforts toward the SARS-CoV-2 emergence.

Diversity of biting midges Culicoides (Diptera: Ceratopogonidae), potential vectors of disease, in different environments in an Amazonian rural settlement, Brazil

Abstract INTRODUCTION: The Culicoides transmit a variety of pathogens. Our aim was to survey the Culicoides species occurring in an Amazonian rural settlement, comparing abundance, richness, and diversity in different environments. METHODS: Culicoides were captured using CDC light traps. The Shannon-Wiener (H') and Rényi indices were used to compare species diversity and evenness between environments, the equitability (J') index was used to calculate the uniformity of distribution among species, and similarity was estimated using the Jaccard similarity index. A permutational multivariate analysis of variance was applied to assess the influence of environment on species composition. A non-metric dimensional scale was used to represent the diversity profiles of each environment in a multidimensional space. RESULTS: 6.078 Culicoides were captured, representing 84 species (45 valid species/39 morphotypes). H' values showed the following gradient: forest > capoeira > peridomicile > forest edge. The equitability J' was greater in capoeira and forests compared to peridomiciles and the forest edge. The population compositions of each environment differed statistically, but rarefaction estimates indicate that environments of the same type possessed similar levels of richness. Species of medical and veterinary importance were found primarily in peridomiciles: C. paraensis, vector of Oropouche virus; C. insignis and C. pusillus, vectors of Bluetongue virus; C. filariferus, C. flavivenula, C. foxi, and C. ignacioi, found carrying Leishmania DNA. CONCLUSIONS: This study indicates that diversity was higher in natural environments than in anthropized environments, while abundance and richness were highest in the most anthropized environment. These findings suggest that strictly wild Culicoides can adapt to anthropized environments.

Genomic and phylogenetic characterisation of an imported case of SARS-CoV-2 in Amazonas State, Brazil

A new coronavirus [severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)] is currently causing a life-threatening pandemic. In this study, we report the complete genome sequencing and genetic characterisation of a SARS-CoV-2 detected in Manaus, Amazonas, Brazil, and the protocol we designed to generate high-quality SARS-CoV-2 full genome data. The isolate was obtained from an asymptomatic carrier returning from Madrid, Spain. Nucleotide sequence analysis showed a total of nine mutations in comparison with the original human case in Wuhan, China, and support this case as belonging to the recently proposed lineage A.2. Phylogeographic analysis further confirmed the likely European origin of this case. To our knowledge, this is the first SARS-CoV-2 genome obtained from the North Brazilian Region. We believe that the information generated in this study may contribute to the ongoing efforts toward the SARS-CoV-2 emergence.

Ecological niche modelling for predicting the risk of cutaneous leishmaniasis in the Neotropical moist forest biome.

A major challenge of eco-epidemiology is to determine which factors promote the transmission of infectious diseases and to establish risk maps that can be used by public health authorities. The geographic predictions resulting from ecological niche modelling have been widely used for modelling the future dispersion of vectors based on the occurrence records and the potential prevalence of the disease. The establishment of risk maps for disease systems with complex cycles such as cutaneous leishmaniasis (CL) can be very challenging due to the many inference networks between large sets of host and vector species, with considerable heterogeneity in disease patterns in space and time. One novelty in the present study is the use of human CL cases to predict the risk of leishmaniasis occurrence in response to anthropogenic, climatic and environmental factors at two different scales, in the Neotropical moist forest biome (Amazonian basin and surrounding forest ecosystems) and in the surrounding region of French Guiana. With a consistent data set never used before and a conceptual and methodological framework for interpreting data cases, we obtained risk maps with high statistical support. The predominantly identified human CL risk areas are those where the human impact on the environment is significant, associated with less contributory climatic and ecological factors. For both models this study highlights the importance of considering the anthropogenic drivers for disease risk assessment in human, although CL is mainly linked to the sylvatic and peri-urban cycle in Meso and South America.

Genomic, epidemiological and digital surveillance of Chikungunya virus in the Brazilian Amazon.

BACKGROUND: Since its first detection in the Caribbean in late 2013, chikungunya virus (CHIKV) has affected 51 countries in the Americas. The CHIKV epidemic in the Americas was caused by the CHIKV-Asian genotype. In August 2014, local transmission of the CHIKV-Asian genotype was detected in the Brazilian Amazon region. However, a distinct lineage, the CHIKV-East-Central-South-America (ECSA)-genotype, was detected nearly simultaneously in Feira de Santana, Bahia state, northeast Brazil. The genomic diversity and the dynamics of CHIKV in the Brazilian Amazon region remains poorly understood despite its importance to better understand the epidemiological spread and public health impact of CHIKV in the country. METHODOLOGY/PRINCIPAL FINDINGS: We report a large CHIKV outbreak (5,928 notified cases between August 2014 and August 2018) in Boa vista municipality, capital city of Roraima's state, located in the Brazilian Amazon region. We generated 20 novel CHIKV-ECSA genomes from the Brazilian Amazon region using MinION portable genome sequencing. Phylogenetic analyses revealed that despite an early introduction of the Asian genotype in 2015 in Roraima, the large CHIKV outbreak in 2017 in Boa Vista was caused by an ECSA-lineage most likely introduced from northeastern Brazil. Epidemiological analyses suggest a basic reproductive number of R0 of 1.66, which translates in an estimated 39 (95% CI: 36 to 45) % of Roraima's population infected with CHIKV-ECSA. Finally, we find a strong association between Google search activity and the local laboratory-confirmed CHIKV cases in Roraima. CONCLUSIONS/SIGNIFICANCE: This study highlights the potential of combining traditional surveillance with portable genome sequencing technologies and digital epidemiology to inform public health surveillance in the Amazon region. Our data reveal a large CHIKV-ECSA outbreak in Boa Vista, limited potential for future CHIKV outbreaks, and indicate a replacement of the Asian genotype by the ECSA genotype in the Amazon region.

Potential of Aedes albopictus as a bridge vector for enzootic pathogens at the urban-forest interface in Brazil.

The invasive species Aedes albopictus is present in 60% of Brazilian municipalities, including at the interfaces between urban settings and forests that are zoonotic arbovirus hotspots. We investigated Ae. albopictus colonization, adult dispersal and host feeding patterns in the anthropic-natural interface of three forested sites covering three biomes in Brazil in 2016. To evaluate whether an ecological overlap exists between Ae. albopictus and sylvatic yellow fever virus (YFV) in forests, we performed similar investigations in seven additional urban-forest interfaces where YFV circulated in 2017. We found Ae. albopictus in all forested sites. We detected eggs and adults up to 300 and 500 m into the forest, respectively, demonstrating that Ae. albopictus forest colonization and dispersal decrease with distance from the forest edge. Analysis of the host identity in blood-engorged females indicated that they fed mainly on humans and domestic mammals, suggesting rare contact with wildlife at the forest edge. Our results show that Ae. albopictus frequency declines as it penetrates into the forest and highlight its potential role as a bridge vector of zoonotic diseases at the edge of the Brazilian forests studied.

Mansonella ozzardi mitogenome and pseudogene characterisation provides new perspectives on filarial parasite systematics and CO-1 barcoding.

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.

The Impact of Global Environmental Changes on Infectious Disease Emergence with a Focus on Risks for Brazil.

Environmental changes have a huge impact on the emergence and reemergence of certain infectious diseases, mostly in countries with high biodiversity and serious unresolved environmental, social, and economic issues. This article summarizes the most important findings with special attention to Brazil and diseases of present public health importance in the country such as Chikungunya, dengue fever, yellow fever, Zika, hantavirus pulmonary syndrome, leptospirosis, leishmaniasis, and Chagas disease. An extensive literature review revealed a relationship between infectious diseases outbreaks and climate change events (El Niño, La Niña, heatwaves, droughts, floods, increased temperature, higher rainfall, and others) or environmental changes (habitat fragmentation, deforestation, urbanization, bushmeat consumption, and others). To avoid or control outbreaks, integrated surveillance systems and effective outreach programs are essential. Due to strong global and local influence on emergence of infectious diseases, a more holistic approach is necessary to mitigate or control them in low-income nations.

The Impact of the Extreme Amazonian Flood Season on the Incidence of Viral Gastroenteritis Cases.

During the Amazonian flood season in 2012, the Negro River reached its highest level in 110 years, submerging residential and commercial areas which appeared associated with an elevation in the observed gastroenteritis cases in the city of Manaus. The aim of this study was to evaluate the microbiological water quality of the Negro River basin during this extreme flood to investigate this apparent association between the illness cases and the population exposed to the contaminated waters. Forty water samples were collected and analysed for classic and emerging enteric viruses. Human adenoviruses, group A rotaviruses and genogroup II noroviruses were detected in 100, 77.5 and 27.5% of the samples, respectively, in concentrations of 103-106 GC/L. All samples were compliant with local bacteriological standards. HAdV2 and 41 and RVA G2, P[6], and P[8] were characterised. Astroviruses, sapoviruses, genogroup IV noroviruses, klasseviruses, bocaviruses and aichiviruses were not detected. Statistical analyses showed correlations between river stage level and reported gastroenteritis cases and, also, significant differences between virus concentrations during this extreme event when compared with normal dry seasons and previous flood seasons of the Negro River. These findings suggest an association between the extreme flood experienced and gastrointestinal cases in the affected areas providing circumstantial evidence of causality between the elevations in enteric viruses in surface waters and reported illness.

Molecular Verification of New World Mansonella perstans Parasitemias.

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.

Evaluation of simple diffusion chlorinators for decontamination of wells in a rural settlement in Amazonia, Brazil.

While the Amazon region has the world's largest reserve of fresh water, the lack of water services and water treatment, especially in non-urban regions, causes environmental and health problems. In isolated rural settlements supply is usually by shallow wells, and the quality of water is a concern for residents. These are situations where there are restricted options for water treatment. This study aimed to assess the use of simplified diffusion chlorinators as an alternative water treatment method. Bacteriological analyses were made of 100 samples of water from the wells, before and after application of the chlorinators, in the Rural Settlement of Rio Pardo, Presidente Figueiredo in the Brazilian State of Amazonas. The sources that were analyzed were considered inappropriate for consumption without prior treatment, and the use of the chlorinators eliminated all contamination by thermotolerant coliforms in the great majority of cases. Also, the method was well received by residents, because it does not leave a taste in the water, is relatively low-cost and handling is easy. We discuss the advantages and limitations of the use of this method of treatment for this social-environmental context and present suggestions for improvement and adaptation, for application of this methodology in other settlements.

Historic accounts of Mansonella parasitaemias in the South Pacific and their relevance to lymphatic filariasis elimination efforts today.

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.

Avaliação de cloradores simplificados por difusão para descontaminação de água de poços em assentamento rural na Amazônia, Brasil / Evaluation of simple diffusion chlorinators for decontamination of wells in a rural settlement in Amazonia, Brazil

Resumo Apesar da região amazônica abrigar a maior reserva de água doce do planeta, a falta de saneamento e de tratamento de água, sobretudo na zona rural, causa problemas ambientais e de saúde. Em assentamentos rurais isolados, o abastecimento geralmente é feito por poços rasos (cacimbões) e a qualidade da água é uma preocupação dos moradores. Nestes casos, as opções de tratamento de água são restritas. O objetivo deste estudo foi avaliar o uso de cloradores simplificados por difusão como método alternativo de tratamento de água. Foram realizadas análises bacteriológicas de 100 amostras de água dos poços, antes e após a aplicação dos cloradores, no Assentamento Rural do Rio Pardo, Presidente Figueiredo (AM). As fontes analisadas foram consideradas inadequadas para consumo sem tratamento prévio, e o uso dos cloradores zerou a contaminação por coliformes termotolerantes, na grande maioria dos casos. Além disso, o método teve boa receptividade pelos moradores por não conferir sabor à água de consumo, por ter relativo baixo custo e ser de fácil manuseio. Discutimos as vantagens e as limitações do uso deste método de tratamento para esse contexto socioambiental e apresentamos sugestões de melhoria e adaptação para a aplicação desta metodologia em outros assentamentos.

Abstract While the Amazon region has the world’s largest reserve of fresh water, the lack of water services and water treatment, especially in non-urban regions, causes environmental and health problems. In isolated rural settlements supply is usually by shallow wells, and the quality of water is a concern for residents. These are situations where there are restricted options for water treatment. This study aimed to assess the use of simplified diffusion chlorinators as an alternative water treatment method. Bacteriological analyses were made of 100 samples of water from the wells, before and after application of the chlorinators, in the Rural Settlement of Rio Pardo, Presidente Figueiredo in the Brazilian State of Amazonas. The sources that were analyzed were considered inappropriate for consumption without prior treatment, and the use of the chlorinators eliminated all contamination by thermotolerant coliforms in the great majority of cases. Also, the method was well received by residents, because it does not leave a taste in the water, is relatively low-cost and handling is easy. We discuss the advantages and limitations of the use of this method of treatment for this social-environmental context and present suggestions for improvement and adaptation, for application of this methodology in other settlements.

Viruses Surveillance Under Different Season Scenarios of the Negro River Basin, Amazonia, Brazil.

The Negro River is located in the Amazon basin, the largest hydrological catchment in the world. Its water is used for drinking, domestic activities, recreation and transportation and water quality is significantly affected by anthropogenic impacts. The goals of this study were to determine the presence and concentrations of the main viral etiological agents of acute gastroenteritis, such as group A rotavirus (RVA) and genogroup II norovirus (NoV GII), and to assess the use of human adenovirus (HAdV) and JC polyomavirus (JCPyV) as viral indicators of human faecal contamination in the aquatic environment of Manaus under different hydrological scenarios. Water samples were collected along Negro River and in small streams known as igarapés. Viruses were concentrated by an organic flocculation method and detected by quantitative PCR. From 272 samples analysed, HAdV was detected in 91.9%, followed by JCPyV (69.5%), RVA (23.9%) and NoV GII (7.4%). Viral concentrations ranged from 10(2) to 10(6) GC L(-1) and viruses were more likely to be detected during the flood season, with the exception of NoV GII, which was detected only during the dry season. Statistically significant differences on virus concentrations between dry and flood seasons were observed only for RVA. The HAdV data provides a useful complement to faecal indicator bacteria in the monitoring of aquatic environments. Overall results demonstrated that the hydrological cycle of the Negro River in the Amazon Basin affects the dynamics of viruses in aquatic environments and, consequently, the exposure of citizens to these waterborne pathogens.

A field trial of a PCR-based Mansonella ozzardi diagnosis assay detects high-levels of submicroscopic M. ozzardi infections in both venous blood samples and FTA card dried blood spots.

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.