Genetic Analysis of LigA, LipL32, Loa22, and OmpL1 Antigens from Leptospira spp. Sejroe Serogroup: How Close We Are To a Vaccine Against Bovine Leptospirosis?

Bovine leptospirosis has as main causative agents Leptospira spp. from Sejroe serogroup. Vaccination is a crucial step to control this infection. The use of conserved proteins among Leptospira spp. is of great importance for a protective immune response. The aim of the present study is to genetically analyze antigens of Leptospira spp. from Sejroe serogroup strains isolated from cattle for a preliminary evaluation of vaccine candidates. Genes associated with antigenicity-LigA, LipL32, Loa22, and OmpL1-were analyzed through bioinformatic and immunoinformatic tools. Despite high diversity observed in strains, on an amino acid level, highly conserved regions were observed (> 90%), particularly in LipL32 gene. Moreover, highly conserved amino acid regions (> 30 aa) were observed in all genes, regardless of species, geographical origin or biological source of isolation. Superposed structures of protein fragments including all the predicted MHC-II and B-Cell epitopes were demonstrated. Results presented herein are preliminary, but a fundamental step towards the development of an efficient vaccine against bovine leptospirosis, a silent but enormously concerning disease.

Giardia lamblia-infected preschoolers present growth delays independent of the assemblage A, B or E.

BACKGROUND: Intestinal parasite Giardia can affect children's physical development mainly stunting even in asymptomatic cases. The protozoa G. lamblia is divided into assemblages A-H. However, it is still unclear whether clinical manifestations and pathogenesis may vary according to the infecting assemblage. OBJECTIVES: To investigate whether G. lamblia assemblages influence differently the physical development of preschoolers from a community of Rio de Janeiro, Brazil. METHODS: Anthropometric parameters were analysed from children attending a daycare centre and stool samples were obtained for the G. lamblia diagnosis. G. lamblia isolates from positive samples were genotyped. Data were analysed in order to verify whether there is a relationship between G. lamblia infection and the physical development of children according to the assemblage. FINDINGS: Herein we demonstrated that although eutrophic, G. lamblia-infected daycare preschoolers from a low-income community presented growth delay compared to non-infected ones. This effect was observed for the three assemblages (A, B or E) found infecting humans. MAIN CONCLUSION: G. lamblia causes growth delays on children independent of infecting assemblage (A, B or E).

The same strain leading to different clinical outcomes: The enigma behind the canine leptospirosis.

Canine leptospirosis is a worldwide zoonosis, varying from asymptomatic and chronic infections to clinical acute disease. In many parts of the world Leptospira interrogans serogroup Icterohaemorrhagiae strains have great epidemiological importance, being the most prevalent on dogs. The present study aims to characterize and compare strains/sequences belonging to the serogroup Icterohaemorrhagiae recovered from clinically ill and asymptomatic dogs. Based on secY gene sequences of L. interrogans serovar Icterohaemorrhagiae, we have studied genetic diversity of strains obtained from 13 dogs, including dogs with clinical signs of acute leptospirosis, asymptomatic dogs and animals with chronic kidney disease, all of them from the same geographical area, the state of Rio de Janeiro, Brazil. No genetic variations on secY gene were observed between strains/groups. No significant associations were observed between clinical status and age, sex or vaccinal status. The same strain leads to different clinical outcomes on canine leptospirosis. The answer for this will rise from deep studies regarding whole genomic sequencing of the strains, as well as proteomics. Those studies may provide key information for understanding of the clinical manifestation of the disease.

Leptospira spp. strains associated with Bovine Genital Leptospirosis (BGL).

Bovine Genital Leptospirosis (BGL) is an important syndrome that leads to reproductive failures. The present study aimed to perform a molecular analysis of Leptospira spp. identified from genital and urine samples from in vivo naturally infected cows with poor reproductive performance. A total of 48 cows destined for culling due to low reproductive efficiency were selected and submitted to sampling. Uterine fragments, cervicovaginal mucus (CVM), and urine were collected from all of the cows and processed for culturing and PCR. One isolate was recovered from the uterus of one cow. Other 25 animals were PCR-positive, totaling 26 positive cows. Of them, 18 animals were positive in lipL32-PCR to genital samples, while only seven animals were positive in urine. From those, sequencing of secY gene was performed. Of the 21 good sequences obtained, 16 were L. interrogans, two were L. noguchii, two were L. santarosai and one was L. borgpetersenii. In order to evaluate the genetic similarity of sequences found herein and other sequences from bovines worldwide, a phylogenetic analysis and haplotype networks were performed. Cows with reproductive failures had a significant association (p < 0.05) with positive PCR of genital samples when compared to PCR of urine. None of the animals were positive for genital samples and urine simultaneously. A high diversity of leptospiral strains were found, even in animals of the same epidemiological region. Haplotype networks of L. interrogans showed clusters of sequences from the uterus and CVM with high similarity to other genital sequences originating from previous studies. L. borgpetersenii haplotype networks presented two major clusters with high similarity, even from worldwide sequences, while L. santarosai showed clusters with high genetic distances, even with all the sequences being from Brazil. This study reinforces the theory that BGL and renal infection are distinct diseases, as well as, genital samples are crucial for the diagnosis of cows with reproductive failures caused by leptospires. In addition, haplotype networks confirmed a high genetic similarity between sequences from the present study and Sejroe strains, reinforcing Sejroe strains as the main BGL agents.

Feasibility of dried blood spot for hepatitis C diagnosis in vulnerable subjects and people living in remote areas from Brazil.

BACKGROUND: Agile, accessible and cheap diagnosis of hepatitis C virus (HCV) infection is essential to achieve the elimination of this infection, worldwide, as mandated by the World Health Organzation as part of its strategy for 2030. Dried blood spots (DBS) can be an attractive alternative for sample collection among people living in remote areas and vulnerable populations due to the less invasive collection, its biosafety, and storage & transportation of samples at room temperature. DESIGN: This study aims to estimate the usefulness of dried blood spot samples for the diagnosis and the assessment of HCV infection rates in three different settings in Brazil. Cross-sectional analysis of a sample collection from different populations, aiming to assess the performance of the testing algorithms and respective procedures among different populations with diverse background infection rates. METHODS: We reported the evaluation of DBS as alternative samples for detecting anti-HCV in different groups in real life conditions: (I) Vulnerable subjects living in remote areas of Southeast, North and Northeast Brazil (n = 1464); (II) Beauticians (n = 288); (III) People who use non-injectable drugs (n = 201); (IV) patients referred to outpatient care (n = 275). RESULTS: General assay accuracy was 99%, with a weighted kappa value of 0.9, showing an excellent performance. Sensitivities ranged from 87.5% to 100.0% between groups and specificities were above 99.2%. A total of 194 individuals had HCV RNA in serum and concordance of anti-HCV detection in DBS was 98.4%. CONCLUSIONS: DBS samples could be used for anti-HCV detection in different populations recruited in real life conditions and ambulatory settings, with a high overall sensitivity and specificity.

Evaluation of cytopathological screening results and risk factors of women who underwent Papanicolaou test in a maternity school in Fortaleza, Ceará, Brazil.

INTRODUCTION: Papanicolaou test (Pap smear) is the standard screening test of pre-neoplastic lesions and cervical cancer. This study aimed to investigate cervical cancer screening results and risk factors such as age, reason for the examination, the epithelia detected in the sample, microbiota and signs of sexually transmitted infection (STIs) of women in a maternity school in Fortaleza, Ceará, Brazil. METHODS: In this cross-sectional study, data were retrieved of 353 women who underwent Pap smear between April 2016 and January 2017 at the Federal University of Ceará. RESULTS: Of all Pap smear samples retrieved, 54.1% (191/353) had glandular epithelium and 40.2% (142/353) had metaplastic epithelium. After statistical analyses adjusted for the final model, age ≥51 years (odds ratio = 3.47) and signs of STIs (odds ratio = 4.95) remained as risk factors. CONCLUSIONS: The diagnosis of high-grade lesions and carcinomas in patients older than 50 years indicates a deficiency in cervical screening. Women with signs and symptoms of STIs and candidiasis sought medical services more frequently than asymptomatic women, and presence of these signs and symptoms contributes to the diagnosis of cervical cancer. We highlight the importance of obtaining a correct smear sampling to allow prompt detection of all preneoplastic lesions; moreover, the implementation of human papillomavirus vaccination and an efficient routine Pap screening are necessary in low-resource settings.

Occurrence of uterine carriers for Leptospira interrogans on slaughtered cows.

Reproductive tract is an important site of infection for chronic leptospirosis and cooperate in pathogenesis of reproductive failure, leading to economic losses. Since serology techniques cannot detect chronic carriers, the molecular analysis of clinical samples is an alternative to detect these animals on livestock. The aim of the present study was to perform a retrospective study in order to detect leptospiral uterine carriers in slaughtered cows. Tissue samples were collected from 50 post-pubertal, nonpregnant cows. These samples were fixed in 10% buffered formalin, paraffin-embedded and stored. PCR targeting lipL32 gene and molecular characterization by secY sequence was performed. Leptospiral DNA was identified in 18% (9/50) examined blocks. Two sequences were characterized as L. interrogans. These findings suggest that the presence of infectious leptospires in uterus is associated with the physiopathogenesis of the reproductive failure.

VP7 and VP8* genetic characterization of group A rotavirus genotype G12P[8]: Emergence and spreading in the Eastern Brazilian coast in 2014.

Group A rotavirus (RVA) genotype G12 is habitually associated with diarrhea disease (DD) in African children and recently its detection has increased worldwide. A total of 970 stool samples collected from individuals with DD in the Northeastern, Southeastern, and Southern Brazilian regions, Eastern coast, were analyzed and 321 (33%) were positive for RVA and of these, 241 (75%) genotyped as G12P[8]. The rate of RVA positivity was higher among children aged 5-10 years old (60%). All RVA infections observed in adults aged >21 years were G12P[8] (n = 27) showing that this genotype affected older age groups during the year of 2014 in Brazil. Phylogenetic analysis of VP7 and VP8* G12P[8] strains demonstrated an elevated similarity among Brazilian and G12-III prototypes strains circulating worldwide recently, suggesting that this lineage is associated with the global spread of the G12 genotype, considered as the 6th most prevalent human RVA genotype nowadays; while other G12 lineages remain sporadically detected and usually detected in association with other P genotypes. VP8* analysis revealed that Brazilian strains belong to P[8]-3 lineage, the single P[8] lineage presently detected in the country. No major nucleotide/amino acid disparities were observed among strains recovered from children and adults for VP7 and VP8* genes. These data are essential to support the surveillance studies, particularly in countries where the RVA vaccine was introduced in their National Immunization Program enabling identification of potential alterations in the epidemiological profile that can impact its efficacy in vaccination programs. J. Med. Virol. 89:64-70, 2017. © 2016 Wiley Periodicals, Inc.

Status of benzimidazole resistance in intestinal nematode populations of livestock in Brazil: a systematic review.

BACKGROUND: Benzimidazoles (BZ) are a class of drugs widely used in veterinary and human medicine, creating a great selection pressure and the emergence of BZ resistance. We conducted a systematic review to assess the status of resistance and/or effectiveness reduction of BZ drugs in animal nematodes in Brazil, and make information accessible to the scientific community, as many studies are published in Portuguese. PubMed, SciELO Brasil, LILACS/Bireme, GNTD database, and Google Scholar were searched with no language restrictions. RESULTS: A total of 40 studies met our eligibility criteria (from the year 1989 forward). Sheep was the host most frequently analysed, and albendazole was the most frequently drug studied. The majority of studies (75.7%) showed that BZ drugs are insufficiently active (FECRT <80%) against nematode parasites of livestock. The mean FECRT for fenbendazole, thiabendazole, albendazole, mebendazole, oxfendazole, and ricobendazole were 71.8%, 71.8%, 58.6%, 53.9%, 46.9%, and 41.5%, respectively. It was observed through linear regression that FECRT is significantly reduced over time between 2007 and 2014 (R = -0.653 p = 0.021) for the treatment of cattle with BZ, suggesting progressive loss of effectiveness and increased resistance for these hosts. CONCLUSIONS: The scenario of BZ resistance in nematode populations in Brazil is not favourable. Given the high cost of drug discovery and development, it is urgent to implement control measures and to monitor the effectiveness/resistance to nematodes in livestock in Brazil.

Frequency and molecular characterisation of Entamoeba histolytica, Entamoeba dispar, Entamoeba moshkovskii, and Entamoeba hartmanni in the context of water scarcity in northeastern Brazil.

This study aimed to estimate the frequency, associated factors, and molecular characterisation of Entamoeba histolytica, Entamoeba dispar, Entamoeba moshkovskii, andEntamoeba hartmanni infections. We performed a survey (n = 213 subjects) to obtain parasitological, sanitation, and sociodemographic data. Faecal samples were processed through flotation and centrifugation methods.E. histolytica, E. dispar, E. moshkovskii, and E. hartmanni were identified by nested-polymerase chain reaction (PCR). The overall prevalence of infection was 22/213 (10.3%). The infection rate among subjects who drink rainwater collected from roofs in tanks was higher than the rate in subjects who drink desalinated water pumped from wells; similarly, the infection rate among subjects who practice open defecation was significantly higher than that of subjects with latrines. Out of the 22 samples positive for morphologically indistinguishableEntamoeba species, the differentiation by PCR was successful for 21. The species distribution was as follows: 57.1% to E. dispar, 23.8% to E. histolytica, 14.3% toE. histolytica and E. dispar, and 4.8% E. dispar and E. hartmanni. These data suggest a high prevalence of asymptomatic infection by the group of morphologically indistinguishable Entamoeba histolytica/dispar/moshkovskiicomplex and E. hartmanni species. In this context of water scarcity, the sanitary and socioenvironmental characteristics of the region appear to favour transmission.

A non-enteric adenovirus A12 gastroenteritis outbreak in Rio de Janeiro, Brazil.

A gastroenteritis outbreak that occurred in 2013 in a low-income community in Rio de Janeiro was investigated for the presence of enteric viruses, including species A rotavirus (RVA), norovirus (NoV), astrovirus (HAstV), bocavirus (HBoV), aichivirus (AiV), and adenovirus (HAdV). Five of nine stool samples (83%) from patients were positive for HAdV, and no other enteric viruses were detected. Polymerase chain reaction products were sequenced and subjected to phylogenetic analysis, which revealed four strains and one strain of non-enteric HAdV-A12 and HAdV-F41, respectively. The HAdV-A12 nucleotide sequences shared 100% nucleotide similarity. Viral load was assessed using a TaqMan real-time PCR assay. Stool samples that were positive for HAdV-A12 had high viral loads (mean 1.9 X 107 DNA copies/g stool). All four patients with HAdV-A12 were < 25 months of age and had symptoms of fever and diarrhoea. Evaluation of enteric virus outbreaks allows the characterisation of novel or unique diarrhoea-associated viruses in regions where RVA vaccination is routinely performed.

Increasing incidence of pertussis in Brazil: a retrospective study using surveillance data.

BACKGROUND: Many countries have reported an increase in the incidence of pertussis, which has become a global public health concern. METHODS: In this study, the epidemiology of pertussis in Brazil was assessed retrospectively using surveillance data gathered from case notification forms from 2007 to 2014. RESULTS: From 2007 to 2014, 80,068 suspected cases of pertussis were reported in Brazil. Of these, 24,612 (32 %) were confirmed by various criteria. The annual distribution of confirmed cases demonstrated a significant increase in incidence rate since 2012. A seasonal pattern in which cases occur most frequently between the end of spring and midsummer has been identified. Among the confirmed cases, 34.5 % occurred in infants aged 0-2 months, 22.4 % occurred in infants aged 3-6 months, 21 % occurred in children aged 7 months to 4 years, and 8 % were reported in adults >21 years. Of the confirmed cases, 47.2 % met only clinical criteria, 15.5 % met clinical and epidemiological criteria, and 36.6 % were confirmed in a laboratory. The overall case fatality rate was 2.1 %, reaching 4.7 % among infants aged 0-2 months. The complications most commonly reported in the notification forms were pneumonia, encephalitis, dehydration, otitis, and malnutrition. Of the confirmed cases, 23.1 % occurred in subjects who received at least 3 doses of the pertussis vaccine. Within this group, there were 1098 infants aged 7 to 15 months and 2079 children aged 16 months to 4 years. In 2012, 18 states did not achieve 95 % immunization coverage, a number that dropped to 10 and 6 in 2013 and 2014, respectively. CONCLUSIONS: Brazil's main challenges in facing pertussis resurgence will be to offer the best quality medical attention to reduce mortality, to improve the infrastructure for laboratory diagnosis and to increase vaccination coverage. Additional studies to assess the effectiveness of the current vaccination schedule and basic research on the genetics and evolution of circulating B. pertussis strains are also needed.

Autochthonous transmission of Chagas disease in Rio de Janeiro State, Brazil: a clinical and eco-epidemiological study.

BACKGROUND: After the control of the main modes of Chagas disease (CD) transmission in most endemic countries, it is important to identify the participation of native sylvatic vectors in CD transmission. Although CD is not considered endemic in Rio de Janeiro State (RJ), Brazil, we identified patients with CD born in RJ and investigated the possible autochthonous transmission in the state. METHODS: Patients born in RJ and followed in our institution between 1986 and 2011 were retrospectively analyzed. The cases identified as autochthonous transmission were submitted to epidemiological, clinical, serological, parasitological and molecular studies. Sectional field study with serological survey, research of sylvatic reservoirs and vectors was conducted in rural areas where patients were born. RESULTS: Among 1963 patients, 69 (3.5%) were born in RJ. From these, 15 (21.7%) were considered to have acquired the infection by autochthonous transmission. Cardiac form was the commonest form of presentation (60%). In rural areas in RJ northern region, sylvatic cycles of Trypanosoma cruzi and domestic invasion by Triatoma vitticeps were identified, and CD prevalence among inhabitants was 0.74%.TcI genotype was identified in sylvatic reservoirs and vectors. The genotype (mixed infection TcI/TcVI) could be identified in one of the autochthonous cases. CONCLUSIONS: The autochthonous vectorial transmission of CD occurs in RJ, probably due to wild cycles of T. cruzi and sylvatic vectors, such as T. vitticeps. Therefore, the health authorities should evaluate if RJ should be included in the original endemic area of CD and CD should be included in the diagnostic work out of cardiomyopathy of patients born in RJ. Moreover, control and educational measures should be put into place in the risk areas.

Factors associated with mucosal involvement in tegumentary leishmaniasis: a nation-based study using surveillance data from Brazil.

This study aimed to assess the factors associated with mucosal leishmaniasis (ML) within the scope of tegumentary leishmaniasis (TL) cases reported in Brazil. Surveillance data were assessed, and comparisons were made between ML and cutaneous leishmaniasis (CL) cases. Additionally, ML incidence rates for municipalities were depicted through a geographic information system. From 2007 to 2017, 235,489 TL cases were reported, of which 235,232 were classified as follows: 14,204 (6%) were ML cases and 221,028 (94%) were CL cases. Multivariate analysis showed that the proportion of ML cases reached 16.8% among individuals >75 years (adjusted OR = 2.77; 95% CI = 2.41-3.19; p < 0.001), and ML was also more frequent among males (aOR = 1.28; 95% CI = 1.20-1.38; p < 0.001), HIV-positive patients (aOR = 2.15; 95% CI = 1.80-2.56; p < 0.001), patients residing in urban areas (aOR = 1.52; 95% CI = 1.43-1.62; p < 0.001), and imported cases (with respect to county) when compared to autochthonous cases (aOR = 1.84; 95% CI = 1.71-1.98; p < 0.001). A lower proportion of positive results in direct parasitological examinations was observed in ML cases (32.6% vs. 60.8%; p < 0.001). The leishmanin skin test results were more often positive in ML cases (41.7% vs. 25.9%; p < 0.001). In ML, compatible changes in histopathology were more frequent (14.6% vs. 3.9%; p < 0.001). A greater proportion of ML cases were treated with amphotericin B (6.9% vs. 0.9%; p < 0.001). The case-fatality rate was higher in ML (0.6% vs. 0.1%; p < 0.001). A higher incidence of ML was observed in a geographical band extending across the Amazon region from the southern Para State to the Acre State. ML exhibited varying frequencies within specific populations. The definition of predictable factors predisposing Leishmania-infected subjects to develop ML is important for defining strategies to mitigate the mucosal damage caused by leishmaniasis.

Circulation of Vaccine-derived Rotavirus G1P[8] in a Vulnerable Child Cohort in Rio de Janeiro.

BACKGROUND: The expansion of rotavirus (RV) immunization in several countries reduced the burden of acute diarrheal disease (ADD) and diarrhea-associated mortality. Although community transmission of live attenuated monovalent rotavirus vaccine (G1P[8] RV1) virus has been demonstrated in children and household contacts, fecal shedding of these strains in neonates and infants under six weeks of age has never been demonstrated. The objective of the study was to assess ADD and rotavirus vaccine strain shedding before and after immunization through 24 months of age. METHODS: This was a prospective cohort study in a low-resource community in which stool samples were collected from neonates from 15 to 45 days of age every 2 weeks, after both doses of G1P[8] RV1, and in subsequent ADD episodes until 2 years of age. RV was detected and genotyped in stool samples by RT-PCR. RESULTS: We enrolled 242 participants who were followed for an average of 23 months. The specific prevalence of G1P[8] RV1 virus was 3.3% in neonates and infants less than six weeks of age, 50% after the first dose, and 25.6% after the second dose. Among the 70 participants with ADD, G1P[8] RV1 virus was identified in only one participant (1.4% prevalence). CONCLUSIONS: In vaccinated children, there were no breakthrough infections with G1P[8] RV1 and ADD was rare supporting high vaccine effectiveness. We observed G1P[8] RV1 virus shedding among neonates and infants before the first vaccine dose, providing evidence of transmission of the vaccine strain from immunized children to those who are not yet vaccinated.

Successful treatment with streptomycin of genital leptospirosis in naturally infected cows under field conditions.

The single-dose protocol of streptomycin treatment has been recommended to treat renal leptospirosis in bovines. However, treating genital infection remains a challenge. Recently, a protocol using three doses of streptomycin demonstrated effectiveness in the genital clearance of experimentally infected ewes. Therefore, the present study aimed to apply this three-dose protocol for genital infection treatment in naturally infected cows under field conditions. Thirty beef cows were diagnosed as positive by lipL32-PCR in their genital samples. Nucleotide sequences (n = 10) characterized them as Leptospira interrogans sg Sejroe, genetically related to Hardjoprajitno strains. After molecular diagnosis, 13 cows received a single dose of 25 mg/kg streptomycin. The other 17 cows were submitted to the three-dose protocol. The successful treatment rate of genital infection on the single streptomycin dose was 7/13 (53.8%), while the cows that received the three doses 16/17 were negative (94.1% of efficacy). Based on those results, we conclude that the standard treatment preconized for renal infection is not adequate for genital infection, and the three-dose protocol was successful in eliminating the carrier status of genital leptospirosis.

Pinnipeds carriers of pathogenic Leptospira: New data based on molecular characterization.

Leptospirosis is a bacterial disease caused by the infection of pathogenic strains of the genus Leptospira, endemic in tropical and subtropical regions. Although well documented in terrestrial animals and humans, little information is available on its distribution and impact on marine animals. Despite clinical manifestations that may occur, the occurrence of carriers was suggested in some species. Nevertheless, there are few studies regarding the infection by Leptospira sp. in marine mammals. In this context, and considering the One Health approach, the present aimed to investigate pinnipeds' role as Leptospira sp. carriers. Kidneys of 47 pinnipeds of two species, Arctocephalus australis (n = 40) and Arctocephalus tropicalis (n = 7) were collected. DNA was extracted and the diagnosis was performed through LipL32-PCR and genetic characterization based on secY gene sequencing. Phylogenetic analysis and haplotype networks were constructed. Pathogenic Leptospira sp. DNA was detected in 31.9% (15/47) of the tested pinnipeds. It was possible to amplify and sequence eight strains (6 for A. australis, 2 for A. tropicalis), all identified as L. interrogans, with high similarity with sequences from Icterohaemorrhagiae serogroup. Phylogenetic analysis revealed sequences from the present study grouped in species-specific unique clusters, but very close to others from humans, wild animals, and domestic animals. We demonstrate that pinnipeds could act as carriers, and play an important role in leptospirosis dynamics.

Unconventional Sites for Diagnosis of Leptospirosis in Bovine Anicteric Fetuses.

BACKGROUND: Bovine leptospirosis is an important reproductive disease and abortion is a major sign, leading to economic impacts. Due to its multifactorial etiology, the proper diagnosis of the cause of the abortion is crucial. Necropsy of the fetuses followed by molecular analysis is recommended for diagnosis, and the investigation mainly occurs in the kidneys and liver. This study aimed to analyze unconventional sites for the presence of leptospiral DNA in bovine anicteric aborted fetuses. METHODS: Five fetuses of the same herd were received for necropsy and diagnosis. Conventional lipL32-PCR was performed in the fetuses' kidneys, livers, lungs, hearts, spleens, subcapsular kidney content, abomasal fluid, and in the cavity's hemorrhagic contents. To complete the investigation, the sera of 30 cows of the herd were collected to perform the serologic screening by Microscopic Agglutination Test. In addition, six subfertile non-pregnant cows from the same herd were selected due to their low reproductive performance, and genital samples (uterine fragment and cervicovaginal mucus) and urine were collected for lipL32-PCR. PCR-positive samples were submitted to a nested PCR of the secY gene and intended for sequencing. RESULTS: The herd presented seroreactive animals (11/30, 36.6%), all against the Sejroe serogroup, with titers between 200 and 1600. In necropsy, four fetuses showed hemorrhagic and anicteric lesions, while one fetus had no macroscopic lesions. Regarding molecular analysis, all the fetuses were positive in lipL32-PCR and the positive sites were the heart, lungs, subcapsular kidney content, thymus, kidneys, liver, and abomasal fluid. Only one fetus presented positive results in the kidney and liver, while three fetuses were positive in the abomasal fluid. Five of six cows were positive for lipL32-PCR, all being positive only in genital samples. Of the fetuses and the cows, seven sequences were obtained and all were identified as Leptospira interrogans serogroup Sejroe serovar Hardjoprajitno. CONCLUSIONS: In order to improve the diagnosis of leptospirosis in cows, it is recommended to perform a comprehensive analysis of the samples, beyond the kidneys and liver. Thus, we highly encourage testing multiple organs by PCR to investigate abortions suspected of bovine leptospirosis, particularly in anicteric fetuses.

A molecular and morphological study of Ascaris suum in a human-pig contact scenario in northeastern Brazil.

The aim of the present study was to assess morphologic and genetic data on ascariasis in swine (Sus scrofa domesticus) and humans in low-resource rural and periurban communities in the state of Piauí, Brazil. Our cross-sectional survey included 100 fecal samples obtained from swine and 682 samples from humans. Fifteen pigs were necropsied. Human and porcine fecal samples were examined to identify Ascaris eggs. Parasites obtained in the swine necropsies were studied using scanning electron microscopy (SEM), and the mitochondrial gene encoding the cytochrome oxidase 1 (cox1) enzyme was partially amplified and sequenced for molecular taxonomy and phylogenetic analyses. The overall prevalence of Ascaris eggs in the swine fecal samples was 16/100 (16%). No Ascaris eggs were identified in the human fecal samples. SEM of six worms recovered from pigs demonstrated morphological characteristics of A. suum. Cox1 sequences were compatible with A. suum reference sequences. Original and reference (GenBank) nucleotide sequences were organized into clusters that did not segregate the parasites by host species or and region. The largest haplogroups were dominated by haplotypes H01, H02 and H31. In the communities studied, there was no epidemiological evidence of the zoonotic transmission of ascariasis at the human-swine interface.

Eco-epidemiology of Chagas disease in northeastern Brazil: Triatoma brasiliensis, T. pseudomaculata and Rhodnius nasutus in the sylvatic, peridomestic and domestic environments.

An entomological survey was carried out in four rural localities situated in the state of Ceará, assessing Chagas disease seroprevalence in man, focusing on the presence of vectors in natural foci contiguous to the domestic and peridomestic environments. Fifty-three Triatoma brasiliensis, nine T. pseudomaculata and 71 Rhodnius nasutus were collected in their natural habitats as far as 10 m from the houses, and 663, 59 and 8 respectively were captured in peridomestic artificial structures, adjacent to the houses, including henhouses, pigpens, corrals, perches and piles of bricks, tiles and wood. Within the households, 37 T. brasiliensis, one specimen of T. pseudomaculata and one of R. nasutus were captured. Overall, Trypanosoma cruzi infection rates were 2.3% for T. brasiliensis and 11.3% for R. nasutus. Despite that the seroprevalence survey in man did not reveal positive results using two serological techniques, natural triatomine habitats are juxtaposed to man-made artificial ecotopes, resulting in overlapping habitats. The contiguity between natural ecotopes and human dwellings increases the interaction between vectors and humans, challenging continuous surveillance and vector control efforts.