SAINT LOUIS ENCEPHALITIS VIRUS IN MATO GROSSO, CENTRAL-WESTERN BRAZIL

The dengue virus (DENV), which is frequently involved in large epidemics, and the yellow fever virus (YFV), which is responsible for sporadic sylvatic outbreaks, are considered the most important flaviviruses circulating in Brazil. Because of that, laboratorial diagnosis of acute undifferentiated febrile illness during epidemic periods is frequently directed towards these viruses, which may eventually hinder the detection of other circulating flaviviruses, including the Saint Louis encephalitis virus (SLEV), which is widely dispersed across the Americas. The aim of this study was to conduct a molecular investigation of 11 flaviviruses using 604 serum samples obtained from patients during a large dengue fever outbreak in the state of Mato Grosso (MT) between 2011 and 2012. Simultaneously, 3,433 female Culex spp. collected with Nasci aspirators in the city of Cuiabá, MT, in 2013, and allocated to 409 pools containing 1-10 mosquitoes, were also tested by multiplex semi-nested reverse transcription PCR for the same flaviviruses. SLEV was detected in three patients co-infected with DENV-4 from the cities of Cuiabá and Várzea Grande. One of them was a triple co-infection with DENV-1. None of them mentioned recent travel or access to sylvatic/rural regions, indicating that transmission might have occurred within the metropolitan area. Regarding mosquito samples, one pool containing one Culex quinquefasciatus female was positive for SLEV, with a minimum infection rate (MIR) of 0.29 per 1000 specimens of this species. Phylogenetic analysis indicates both human and mosquito SLEV cluster, with isolates from genotype V-A obtained from animals in the Amazon region, in the state of Pará. This is the first report of SLEV molecular identification in MT.

PARACOCCIDIOIDOMYCOSIS: CHALLENGES IN THE DEVELOPMENT OF A VACCINE AGAINST AN ENDEMIC MYCOSIS IN THE AMERICAS

SUMMARYParacoccidioidomycosis (PCM), caused by Paracoccidioides spp, is an important endemic mycosis in Latin America. There are two recognized Paracoccidioides species, P. brasiliensis and P. lutzii, based on phylogenetic differences; however, the pathogenesis and disease manifestations of both are indistinguishable at present. Approximately 1,853 (~51,2%) of 3,583 confirmed deaths in Brazil due to systemic mycoses from 1996-2006 were caused by PCM. Antifungal treatment is required for patients with PCM. The initial treatment lasts from two to six months and sulfa derivatives, amphotericin B, azoles and terbinafine are used in clinical practice; however, despite prolonged therapy, relapses are still a problem. An effective Th1-biased cellular immune response is essential to control the disease, which can be induced by exogenous antigens or modulated by prophylactic or therapeutic vaccines. Stimulation of B cells or passive transference of monoclonal antibodies are also important means that may be used to improve the efficacy of paracoccidioidomycosis treatment in the future. This review critically details major challenges facing the development of a vaccine to combat PCM.

Paracoccidioides brasiliensis AND Paracoccidioides lutzii, A SECRET LOVE AFFAIR

SUMMARYTo commemorate Prof. Carlos da Silva Lacaz's centennial anniversary, the authors have written a brief account of a few, out of hundreds, biological, ecological, molecular and phylogenetic studies that led to the arrival of Paracoccidioides lutzii, hidden for more than a century within Paracoccidioides brasiliensis. Lacaz's permanent interest in this fungus, and particularly his conviction on the benefits that research on paracoccidioidomycosis would bring to patients, were pivotal in the development of the field.

THE PROCESS OF Leishmania INFECTION - DISEASE AND NEW PERSPECTIVES OF PALEOPARASITOLOGY

Species of the genus Leishmania (Kinetoplastida, Trypanosomatidae) are causative agents of leishmaniasis, a complex disease with variable clinical spectrum and epidemiological diversity, constituting, in some countries, a serious public health problem. The origin and evolution of leishmaniasis has been under discussion regarding some clinical and parasitological aspects. After the introduction of paleoparasitology, molecular methods and immunodiagnostic techniques have been applied allowing the recovery of parasite remains, as well as the diagnosis of past infections in humans and other hosts. The dating of archaeological samples has allowed the parasitological analysis in time and space. This manuscript presents the state of the art of leishmaniasis and prospects related to paleoparasitology studies and their contribution to the evolutionary and phylogenetic clarification of parasites belonging to the genus Leishmania, and the leishmaniasis caused by them.

Molecular evidence of mother-to-child transmission of HTLV-IIc in the Kararao Village (Kayapo) in the Amazon Region of Brazil

Blood samples from native Indians in the Kararao village (Kayapo), were analysed using serological and molecular methods to characterize infection and analyse transmission of HTLV-II. Specific reactivity was observed in 3/26 individuals, of which two samples were from a mother and child. RFLP analysis of the pX and env regions confirmed HTLV-II infection. Nucleotide sequence of the 5' LTR segment and phylogenetic analysis showed a high similarity (98%) between the three samples and prototype HTLV-IIa (Mot), and confirmed the occurrence of the HTLV-IIc subtype. There was a high genetic similarity (99.9%) between the mother and child samples and the only difference was a deletion of two nucleotides (TC) in the mother sequence. Previous epidemiological studies among native Indians from Brazil have provided evidence of intrafamilial and vertical transmission of HTLV-IIc. The present study now provides molecular evidence of mother-to-child transmission of HTLV-IIc, a mechanism that is in large part responsible for the endemicity of HTLV in these relatively closed populations. Although the actual route of transmission is unknown, breast feeding would appear to be most likely.

Polymerase chain reaction and restriction fragment length polymorphism analysis of the ITS2 region for differatiation of Brazilian Biomphalaria intermediate hosts of the Schistosoma mansoni

We sequenced the internal transcribed spacer 2 of the ribosomal DNA (ITS2-DNAr) from the three Schistosoma mansoni intermediate hosts in Brazil: Biomphalaria glabrata, Biomphalaria tenagophila and Biomphalaria straminea. Analysis of a restriction map from those sequences allowed us to select putative restriction enzymes able to identify the snail species under study. Four restriction enzymes were used and HpaII provided simple species-specific profiles easily visualized in polyacrylamide gels. The use of ITS2 is advantageous as it provides a small fragment of 460 bp which may be easily amplified by PCR. In the current work, we showed that the amplification of ITS2-DNAr together with HpaII enzyme restriction is an auxiliary molecular tool for the morphological identification of such snails as well as for taxonomic and phylogenetic studies of neotropical planorbids.

Hepatitis B virus genotyping among chronic hepatitis B patients with resistance to treatment with lamivudine in the City of Ribeirão Preto, State of São Paulo

INTRODUCTION: Lamivudine is a nucleoside analogue that is used clinically for treating chronic hepatitis B infection. However, the main problem with prolonged use of lamivudine is the development of viral resistance to the treatment. Mutations in the YMDD motif of the hepatitis B virus DNA polymerase gene have been associated with resistance to drug therapy. So far, there have not been many studies in Brazil reporting on genotype-dependent development of resistance to lamivudine. Thus, the aim of the present study was to determine the possible correlation between a certain genotype and increased development of resistance to lamivudine among chronic hepatitis B patients. METHODS: HBV DNA in samples from 50 patients under lamivudine treatment was amplified by means of conventional PCR. Samples were collected at Hospital das Clínicas, FMRP-USP. The products were then sequenced and phylogenetic analysis was performed. RESULTS: Phylogenetic analysis revealed that 29 (58%) patients were infected with genotype D, 20 (40%) with genotype A and one (2%) with genotype F. Mutations in the YMDD motif occurred in 20% of the patients with genotype A and 27.6% of the patients with genotype D. CONCLUSIONS: Despite the small number of samples, our results indicated that mutations in the YMDD motif were 1.38 times more frequent in genotype D than in genotype A.

Distribution of hepatitis C virus genotypes among different exposure categories in the State of Pará, Brazilian Amazon

INTRODUCTION: Epidemiological studies concerning HCV genotypic distribution in the Brazilian Amazon are scarce. Thus, this study determined the patterns of distribution of HCV genotypes among different exposure categories in the State of Pará, Brazilian Amazon. METHODS: A cross-sectional study was conducted on 312 HCV-infected individuals belonging to different categories of exposure, who were attended at the HEMOPA, CENPREN and a private hemodialysis clinic in Belém. They were tested for HCV antibodies using an immunoenzymatic test, RNA-HCV, using real-time PCR and HCV genotyping through phylogenetic analysis of the 5' UTR. The population groups were epidemiologically characterized according to data collected in a brief interview or medical consultation. RESULTS: Genotype 1 predominated in all the different categories of HCV exposure. HCV genotypic distribution among blood donors comprised genotypes 1 (94%) and 3 (6%). All patients with chronic hematologic diseases had HCV genotype 1. The genotypic distribution in illicit-drug users comprised genotypes 1 (59.6%) and 3 (40.4%). In patients under hemodialysis, genotypes 1 (90.1%), 2 (3.3%), and 3 (6.6%) were detected. Finally, the frequency of genotypes 1 and 3 was significantly different between the groups: BD and DU, PUH and DU, PUH and PCHD and PCHD and DU. CONCLUSIONS: The genotypic frequency and distribution of HCV in different categories of exposure in the State of Pará showed a predominance of genotype 1, regardless of the possible risk of infection.

Viral etiology among the elderly presenting acute respiratory infection during the influenza season

INTRODUCTION: Acute respiratory tract infections are the most common illness in all individuals. Rhinoviruses have been reported as the etiology of more than 50% of respiratory tract infections worldwide. The study prospectively evaluated 47 elderly individuals from a group of 384 randomly assigned for acute respiratory viral infections (cold or flu) and assessed the occurrence of human rhinovirus (HRV), influenza A and B, respiratory syncytial virus and metapneumovirus (hMPV) in Botucatu, State of São Paulo, Brazil. METHODS: Forty-nine nasal swabs collected from 47 elderly individuals following inclusion visits from 2002 to 2003 were tested by GenScan RT-PCR. HRV-positive samples were sequenced for phylogenetic analysis. RESULTS: No sample was positive for influenza A/B or RSV. HRV was detected in 28.6% (14/47) and hMPV in 2% (1/47). Of 14 positive samples, 9 isolates were successfully sequenced, showing the follow group distribution: 6 group A, 1 group B and 2 group C HRVs. CONCLUSIONS: The high incidence of HRV during the months of the influenza season requires further study regarding HRV infection impact on respiratory complications among this population. Infection caused by HRV is very frequent and may contribute to increasing the already high demand for healthcare during the influenza season.

First report of rabies infection in bats, Molossus molossus, Molossops neglectus and Myotis riparius in the city of São Paulo, State of São Paulo, southeastern Brazil

INTRODUCTION: This paper presents the first report of rabies in three bat species, Molossus molossus, Molossops neglectus and Myotis riparius in the city of São Paulo, Brazil. METHODS: Bats were diagnosed as positive for rabies using the fluorescent antibody test and mouse inoculation test. The isolates were characterized antigenically using a panel of eight monoclonal antibodies. The samples were also genetically analyzed by partial sequencing of the portion of nucleoprotein gene between positions 1157 and 1445nt. RESULTS: Analysis of the results verified that the sample isolated from the species M. molossus presented antigenic variant 6, while the other two samples showed a different profile from that established in the panel, one not previously reported in the literature. The results of genetic analysis revealed that the M. molossus sample segregated with Lasiurus sp. isolates, M. neglectus segregated with a subgroup of Eptesicus furinalis isolates and the Myotis riparius sample segregated with Myotis sp. isolates. CONCLUSIONS: The cases reported in this paper emphasize the need for clarification of the circumstances in which cases of rabies in wildlife occur, principally in urban areas.

Molecular characterization of rabies virus isolated from non-haematophagous bats in Brazil

INTRODUCTION: Rabies is an important zoonosis that causes thousands of deaths worldwide each year. Although the terrestrial cycle, mainly transmitted by dogs, is controlled in Brazil, the aerial cycle remains a serious public health issue, besides the economic problem. In the aerial cycle, the haematophagous bat Desmodus rotundus is the main source of infection, where several different species of non-haematophagous bats can be infected and can transmit the virus. METHODS: The aim of this work was to study the epidemiological pattern of rabies using antigenic characterization with monoclonal antibodies and genetic characterization by reverse-transcriptase polymerase chain reaction followed by sequencing and phylogenetic analysis of non-haematophagous bats' and herbivorous animals' central nervous system samples from the western region of the State of São Paulo, Brazil. RESULTS: From 27 samples, 3 antigenic variants were identified: AgV-3, AgV-4, and AgV-6; and from 29 samples, 5 different clusters were identified, all belonging to the rabies virus species. CONCLUSIONS: Although only non-haematophagous bats were evaluated in the studied region, the majority of samples were from antigenic and genetic variants related to haematophagous bats Desmodus rotundus. Samples from the same antigenic variant were segregated in more than one genetic cluster. This study demonstrated the diversity of rabies virus genetic lineages presented and circulating in non-haematophagous bats in the studied region.