A real-time reverse transcriptase polymerase chain reaction for detection and quantification of Vesiculovirus.

Vesiculoviruses (VSV) are zoonotic viruses that cause vesicular stomatitis disease in cattle, horses and pigs, as well as sporadic human cases of acute febrile illness. Therefore, diagnosis of VSV infections by reliable laboratory techniques is important to allow a proper case management and implementation of strategies for the containment of virus spread. We show here a sensitive and reproducible real-time reverse transcriptase polymerase chain reaction (RT-PCR) for detection and quantification of VSV. The assay was evaluated with arthropods and serum samples obtained from horses, cattle and patients with acute febrile disease. The real-time RT-PCR amplified the Piry, Carajas, Alagoas and Indiana Vesiculovirus at a melting temperature 81.02 ± 0.8ºC, and the sensitivity of assay was estimated in 10 RNA copies/mL to the Piry Vesiculovirus. The viral genome has been detected in samples of horses and cattle, but not detected in human sera or arthropods. Thus, this assay allows a preliminary differential diagnosis of VSV infections.

A tale of 141 municipalities: the spatial distribution of dengue in Mato Grosso, Brazil.

BACKGROUND: In recent years, the state of Mato Grosso has presented one of the highest dengue incidence rates in Brazil. The meeting of the Amazon, Cerrado and Pantanal biomes results in a large variation of rainfall and temperature across different regions of the state. In addition, Mato Grosso has been undergoing intense urban growth since the 1970s, mainly due to the colonization of the Mid-North and North regions. We analyzed factors involved in dengue incidence in Mato Grosso from 2008 to 2019. METHODS: The Moran Global Index was used to assess spatial autocorrelation of dengue incidence using explanatory variables such as temperature, precipitation, deforestation, population density and municipal development index. Areas at risk of dengue were grouped by the Local Moran Indicator. RESULTS: We noticed that areas at risk of dengue expanded from the Mid-North region to the North; the same pattern occurred from the Southeast to the Northeast; the South region remained at low-risk levels. The increase in incidence was influenced by precipitation, deforestation and the municipal development index. CONCLUSIONS: The identification of risk areas for dengue in space and time enables public health authorities to focus their control and prevention efforts, reducing infestation and the potential impact of dengue in the human population.

Invertebrates for vertebrate biodiversity monitoring: Comparisons using three insect taxa as iDNA samplers.

Metabarcoding of environmental DNA (eDNA) is now widely used to build diversity profiles from DNA that has been shed by species into the environment. There is substantial interest in the expansion of eDNA approaches for improved detection of terrestrial vertebrates using invertebrate-derived DNA (iDNA) in which hematophagous, sarcophagous, and coprophagous invertebrates sample vertebrate blood, carrion, or faeces. Here, we used metabarcoding and multiple iDNA samplers (carrion flies, sandflies, and mosquitos) collected from 39 forested sites in the southern Amazon to profile gamma and alpha diversity. Our main objectives were to (1) compare diversity found with iDNA to camera trapping, which is the conventional method of vertebrate diversity surveillance; and (2) compare each of the iDNA samplers to assess the effectiveness, efficiency, and potential biases associated with each sampler. In total, we collected and analysed 1759 carrion flies, 48,686 sandflies, and 4776 mosquitos. Carrion flies revealed the greatest total vertebrate species richness at the landscape level, despite the least amount of sampling effort and the fewest number of individuals captured for metabarcoding, followed by sandflies. Camera traps had the highest median species richness at the site-level but showed strong bias towards carnivore and ungulate species and missed much of the diversity described by iDNA methods. Mosquitos showed a strong feeding preference for humans as did sandflies for armadillos, thus presenting potential utility to further study related to host-vector interactions.

ZIKV B-cell epitopes for immunodiagnostic tests.

The use of serological tests is valuable to diagnose Zika virus (ZIKV) infection and carry out epidemiological surveillance. However, ZIKV serological tests may result in false positives due to cross-reactivity between antibodies against other Flavivirus, especially dengue virus that worldwide disseminated. We used three online tools to predict amino acid sequences of B-cell epitopes. We selected and synthetized two epitopes that showed appropriate features in the molecular dynamic simulation and demonstrated to be suitable for serological assays.

Ecological aspects of potential arbovirus vectors (Diptera: Culicidae) in an urban landscape of Southern Amazon, Brazil.

Amazon has been under intense human pressure, especially in the so-called "Arc of Deforestation" in the Eastern and Southern regions. Changes in biodiversity due to landscape disturbance in municipalities at the Arc of Deforestation are likely to impact mosquito species leading to the potential for emergence of arboviruses. Our aim was to describe the composition and structure of culicids in Sinop, a municipality located in the Southern Amazon at the State of Mato Grosso, as well as their presence in urban habitats, analyzing climate variables and potential risk of arbovirus transmission. Mosquitoes were collected in four habitats located in urban environments for 12 months. We recovered 5,210 adult mosquitoes from mosquito traps. Among the 33 species identified, 19 are associated with arbovirus transmission. Higher abundance, richness, diversity, and equitability of mosquito species were observed in urban forest parks and in new neighborhoods located in the outskirts of the city, nearby forest fragments and agricultural areas. As the environment consolidates as urban, both abundance and richness decrease. Highly urbanized neighborhoods had higher dominance of Culex quinquefasciatus, a mosquito associated with different arboviruses, including West Nile and Saint Louis encephalitis virus. The medically important species Cx. declarator, Cx. (Melanoconion) ssp., Aedeomyia squamipennis and Aedes scapularis were found in the four habitats. It is possible that these mosquitoes are adapting to different habitats and as a consequence, they are reaching urbanized areas. Mosquito abundance was higher in the days of high temperature and low precipitation in urban forest parks and in highly urbanized neighborhoods. These results suggest that Sinop region is a key area for surveillance of arbovirus vectors.

Detection of Ilheus virus in mosquitoes from southeast Amazon, Brazil.

BACKGROUND: Arbovirus surveillance in field-collected mosquitoes is essential in monitoring virus activity to avoid emergence and outbreaks of arboviruses. METHODS: We used reverse transcription polymerase chain reaction methods to search for arbovirus in mosquitoes collected in Brazil's southeast Amazon forest remnants during 2015-2016. RESULTS: We detected Iheus virus (ILHV) RNA in Culex declarator, Culex (Melanoconion) and Ochlerotatus serratus mosquitoes. CONCLUSIONS: These results indicate ILHV circulation in the studied area, highlighting its potential emergence in human population. More studies are necessary to confirm the roles of these three species in ILHV maintenance.

Aspects on the ecology of phlebotomine sand flies and natural infection by Leishmania hertigi in the Southeastern Amazon Basin of Brazil.

The medical and veterinary importance of sand flies (Diptera: Psychodidae) follow as a result of some species ability to transmit the zoonotic protozoa of the genus Leishmania. Of all municipalities in the state of Mato Grosso, Sinop ranks first in reported cases of American Cutaneous Leishmaniasis (ACL). Sinop urban zone encompasses three permanent forest preservation areas (APPs) that provide refuge for insects and other vertebrate hosts. We assessed ecological parameters and investigated the natural infection by Leishmania spp. of the phlebotomine fauna from four ecotypes with different levels of urbanization in the urban area of Sinop. A total of 62,745 sand flies were collected, of which 52.34% female. Out of 37 species in this study, nine were found to be constant. Sand flies frequency and diversity were highest in APPs (96.85%; 33 species). Lutzomyia dasypodogeton was the most frequent species and exhibited the greatest abundance (SISA=0.977). The neighborhoods around APPs and completely urbanized neighborhoods presented noteworthy ecological similarity. Moreover, eight vector sand fly species with medicalwere identified, and one L. antunesi sample pool was found to be naturally infected with Le. hertigi. We observed a high frequency and diversity of sand flies, including some species that are known to be major vectors of ACL. Further studies are needed on the natural rates of infection in humans, domestic animals, and sylvatic hosts to better comprehend the leishmaniases dynamics.

Serological evidence of hantavirus infection in an urban area in Mato Grosso State, Brazil.

INTRODUCTION: In Brazil, Mato Grosso (MT) has the highest number of hantavirus cardiopulmonary syndrome cases. Our study aimed to identify anti-hantavirus antibodies in the sera of patients from Sinop, MT, presenting with acute febrile illness. METHODS: A retrospective analysis of data for 198 sera samples assessed using enzyme-linked immunosorbent assay (ELISA) was conducted. RESULTS: Immunoglobulins G (IgGs) against the hantavirus nucleoprotein were found in 13.6% of the tested sera. No sample had immunoglobulin M (IgM) antibodies to hantavirus. Seropositivity occurred mainly in female residents in urban areas who worked around the household. CONCLUSIONS: Our findings suggest circulation of hantavirus in Sinop.

Frequency and diversity of phlebotomine sand flies (Diptera: Psychodidae) in Sinop, State of Mato Grosso, Brazil.

INTRODUCTION:: Understanding the diversity of sand flies is important for the epidemiology and control of leishmaniasis. This study aimed to understand the frequency, diversity, and seasonality of medically important sand flies in the municipality of Sinop, State of Mato Grosso, Brazil. METHODS:: The study was conducted in an urban area, including four ecotypes with different levels of urbanization. The sand flies were collected using light traps for three nights per month, from May 2014 to April 2015. RESULTS:: A total of 62,745 sand flies was collected, 52.34% of which were female. The frequency and diversity of sand flies was the highest in areas of permanent preservation (APPs) (96.85%), and was lower in more urbanized areas. Lutzomyia dasypodogeton was the most frequent species in the APPs. Lutzomyia antunesi was the most frequent in neighborhoods with forest fragments and neighborhoods around APPs, and L. aragaoi was the most frequent in completely urbanized neighborhoods. A higher frequency and diversity of sand flies was observed in the rainy season (87.92%) than in the dry season (12.08%). Eight medically important species were captured, and Lutzomyia antunesi, which is associated with American cutaneous leishmaniasis and visceral leishmaniasis, was observed in all ecotypes throughout the year. CONCLUSIONS:: We observed a high frequency and diversity of sand flies in all urban areas, and some species collected were major vectors of leishmaniasis. These results support the need for further studies of the natural rates of infection of these insects and the circulation of the disease in hosts and vectors.

Detection of Mayaro virus infections during a dengue outbreak in Mato Grosso, Brazil.

Arboviruses are common agents of human febrile illness worldwide. In dengue-endemic areas illness due to other arboviruses have been misdiagnosed as dengue based only on clinical-epidemiological data. In this study we investigated the presence of Brazilian arboviruses in sera of 200 patients presenting acute febrile illness, during a dengue outbreak in Sinop, MT, Brazil. The results showed that 38 samples were positive to Dengue virus (DENV) type 1, two samples to DENV type 4, and six to Mayaro virus. These results indicate that arboviruses others than DENV are circulating in Sinop and the surrounding region, which are going undiagnosed. In addition, molecular and evolutionary analyses indicate that two MAYV genotypes are co-circulating in Mato Grosso, Brazil. Thus, a strong surveillance program must be implemented to evaluate and monitor the distribution and the true importance of non-dengue arboviruses in the etiology of acute febrile illnesses.

Dengue-4 false negative results by Panbio® Dengue Early ELISA assay in Brazil.

BACKGROUND: Dengue is a serious public health problem in numerous countries. The ability to rapidly diagnosis dengue is important for patient triage and management. Detection of dengue viral protein, NS1, represents a new approach to dengue diagnosis. OBJECTIVE: The present study aims to evaluate if there are false negative results using the NS1 Ag rapid assay (Panbio(®) Dengue Early ELISA) in two different epidemiological situations (epidemic and non-epidemic). STUDY DESIGN: 220 serum samples from patients with clinical symptoms of classical dengue fever were tested by NS1 antigen capture ELISA and Multiplex-Nested-PCR. RESULTS: In samples collected in a non-epidemic period we found a 100% agreement of ELISA and RT-PCR in dengue negative samples and 85% agreement of ELISA and RT-PCR in dengue positive samples. But when we tested samples during an epidemic period (large DENV-4 outbreak) we found 15% false negative results (p<0.05) in dengue negative samples. CONCLUSIONS: Due to false negative results for DENV-4, the sole use of the Panbio(®) Dengue Early ELISA assay as a screening method for monitoring circulating dengue serotypes must be reevaluated.

Frequency and diversity of phlebotomine sand flies (diptera: psychodidae) in sinop, state of Mato Grosso, Brazil

Abstract INTRODUCTION: Understanding the diversity of sand flies is important for the epidemiology and control of leishmaniasis. This study aimed to understand the frequency, diversity, and seasonality of medically important sand flies in the municipality of Sinop, State of Mato Grosso, Brazil. METHODS: The study was conducted in an urban area, including four ecotypes with different levels of urbanization. The sand flies were collected using light traps for three nights per month, from May 2014 to April 2015. RESULTS: A total of 62,745 sand flies was collected, 52.34% of which were female. The frequency and diversity of sand flies was the highest in areas of permanent preservation (APPs) (96.85%), and was lower in more urbanized areas. Lutzomyia dasypodogeton was the most frequent species in the APPs. Lutzomyia antunesi was the most frequent in neighborhoods with forest fragments and neighborhoods around APPs, and L. aragaoi was the most frequent in completely urbanized neighborhoods. A higher frequency and diversity of sand flies was observed in the rainy season (87.92%) than in the dry season (12.08%). Eight medically important species were captured, and Lutzomyia antunesi, which is associated with American cutaneous leishmaniasis and visceral leishmaniasis, was observed in all ecotypes throughout the year. CONCLUSIONS: We observed a high frequency and diversity of sand flies in all urban areas, and some species collected were major vectors of leishmaniasis. These results support the need for further studies of the natural rates of infection of these insects and the circulation of the disease in hosts and vectors.

A real-time reverse transcriptase polymerase chain reaction for detection and quantification of Vesiculovirus

Vesiculoviruses (VSV) are zoonotic viruses that cause vesicular stomatitis disease in cattle, horses and pigs, as well as sporadic human cases of acute febrile illness. Therefore, diagnosis of VSV infections by reliable laboratory techniques is important to allow a proper case management and implementation of strategies for the containment of virus spread. We show here a sensitive and reproducible real-time reverse transcriptase polymerase chain reaction (RT-PCR) for detection and quantification of VSV. The assay was evaluated with arthropods and serum samples obtained from horses, cattle and patients with acute febrile disease. The real-time RT-PCR amplified the Piry, Carajas, Alagoas and Indiana Vesiculovirus at a melting temperature 81.02 ± 0.8ºC, and the sensitivity of assay was estimated in 10 RNA copies/mL to the Piry Vesiculovirus. The viral genome has been detected in samples of horses and cattle, but not detected in human sera or arthropods. Thus, this assay allows a preliminary differential diagnosis of VSV infections.

Co-infection ofddengue virus by serotypes 1 and4 in patient from medium sized city from Brazil / Co-infeccao por virus dengue, sorotipos 1 e 4, em paciente de cidade de porte medio no Brasil

SUMMARY The natural co-infection with dengue virus can occur in highly endemic areas where different serotypes have been observed for many years. We report one case of DENV-1/DENV-4 co-infection in human serum detected by molecular tests. Phylogenetic analysis of the sequences obtained indicated the presence of genotype V and II for DENV-1 and DENV-4, respectively. .

RESUMO A co-infecção por dengue vírus pode ocorrer em áreas com circulação endêmica, nas quais diferentes sorotipos vêm circulando durante muitos anos. Neste trabalho relatamos um caso de co-infecção por DENV-1/DENV-4 em soro humano, detectado por testes moleculares. Análises filogenéticas das sequências obtidas indicaram a presença do genótipo V e II de DENV-1 e DENV-4, respectivamente. .

Viral infections of the central nervous system in Brazil.

OBJECTIVE: Epidemiological studies have shown that most central nervous system (CNS) infections are viral. The objective of this study is to contribute to the knowledge base concerning viral CNS infections in Ribeirão Preto, Brazil. METHODS: Two hundred cerebrospinal fluid (CSF) samples were taken from patients with clinically suspected viral CNS infection and analyzed for herpesvirus, enterovirus, alphavirus and flavivirus using PCR assays. RESULTS: Viral genome was detected in 43 CSF samples (23.34%): 6% cytomegalovirus (CMV), 5% herpes simplex virus type 1 (HSV-1), 0.5% each varicella-zoster virus (VZV) and Epstein-Barr virus (EBV), and 11.34% enterovirus. Co-infections (CMV-enterovirus and CMV-HSV-1) were found in 3 patients. CSF parameters such as cytology and protein level were normal in many patients with viral genome-positive CSF. CONCLUSIONS: Data obtained in this study contribute to the knowledge base concerning viral CNS infections in Brazil. This information will have a major impact on the clinical management of patients with CNS disease.