Geographic analysis on the occurrence of human and canine leptospirosis in the city of Maringá, state of Paraná, Brazil

INTRODUCTION: Leptospirosis is a zoonotic disease, the primary hosts of which are wild, synanthropic, and household animals. Humans behave as terminal and accidental hosts. The prevalence of leptospirosis depends on carrier animals that disseminate the agent, on the environmental survival of this agent, and on the contact of susceptible individuals. Each serovar has one or more hosts with different adaptation levels. The focuses of leptospirosis are infected, sick, and asymptomatic animals, which are considered to be sources of environmental infection. This study aimed to determine the risk areas for leptospiral infection in stray dogs and patients diagnosed with leptospirosis from 2006 to 2008 in Maringá, State of Paraná, Brazil. METHODS: Three hundred and thirty-five stray dogs and 25 patients were studied. Serum from both animals and patients was examined by the microscopic serum agglutination test to study anti-leptospiral antibodies. To determine the risk areas and the spatial distribution of the disease, thematic maps were designed. RESULTS: Forty-one (12.2%) dogs positive for one or more leptospire serovars were observed, the most frequent serovars being Pyrogenes (43.9%), Canícola (21.9%), and Copennhageni (19.5%). Among the humans, 2 (8%) were positive for serovars Pyrogenes and Hardjo Prajitno and for Pyrogenes and Cynopteri. CONCLUSIONS: Spatial analysis showed that the risk for dogs and humans in the City of Maringá to become infected with leptospires exists in both the central and the peripheral areas, a fact that reinforces the relevance of this study and of continuous epidemiological and environmental surveillance actions to control the disease in animals and in humans.

Factors associated with Leishmania chagasi infection in domestic dogs from Teresina, State of Piauí, Brazil

INTRODUCTION: Many studies have evaluated risk factors for human visceral leishmaniasis, but few have focused on the infection among dogs. The objective of this study was to assess the association between peridomestic socioeconomic and environmental factors and the presence of dogs seropositive for Leishmania chagasi in the City of Teresina, Brazil. METHODS: This case-control study was based on the results of a routine seroepidemiological survey among domestic dogs carried out in 2007. Serological tests were performed by means of indirect immunofluorescence antibody test. All dwellings in which at least one seropositive dog was detected were considered cases, and controls were a random sample of dwellings in which only seronegative dogs were identified. Associations between variables were expressed as odds ratios (OR) and their respective 95% confidence intervals (95%CI) estimated using multivariate logistic regression. RESULTS: Dwellings with a history of dogs removed by the visceral leishmaniasis control program in the last 12 months had five-fold higher odds of having at least one seropositive dog as compared with dwellings having no history of dog removal (OR = 5.19; 95%CI = 3.20-8.42). Dwellings with cats had 58% increased odds of dog infection as compared with those having no cats (OR = 1.58; 95%CI = 1.01-2.47). CONCLUSIONS: Identification of factors associated with canine visceral leishmaniasis might be used for the delimitation of areas of higher risk for human visceral leishmaniasis, since infection in dogs generally precedes the appearance of human cases.

Antigens of worms and eggs showed a differentiated detection of specific IgG according to the time of Schistosoma mansoni infection in mice

INTRODUCTION: The correlation between the immunological assay and the antibody titer can offer a tool for the experimental analysis of different phases of the disease. METHODS: Two simple immunological assays for Schistosoma mansoni in mice sera samples based on specific IgG detection for worms soluble antigens and eggs soluble antigens were standardized and evaluated in our laboratory. Fifty mice were used in negative and positive groups and the results obtained by enzyme-linked immunosorbent assays (ELISA) assays were compared with the number of worms counted and the IgG titers at different times of infection. RESULTS: Data showed that ELISA using adult worm antigens (ELISA-SWAP) presented a satisfactory correlation between the absorbance value of IgG titers and the individual number of worms counted after perfusion technique (R²=0.62). In addition, ELISA-SWAP differentially detected positive samples with 30 and 60 days post infection (p=0.011 and 0.003, respectively), whereas ELISA using egg antigens (ELISA-SEA) detected samples after 140 days (p=0.03). CONCLUSIONS: These data show that the use of different antigens in immunological methods can be used as potential tools for the analysis of the chronological evolution of S. mansoni infection in murine schistosomiasis. Correlations with human schistosomiasis are discussed.

Susceptibility of Argentinean Biomphalaria tenagophila and Biomphalaria straminea to infection by Schistosoma mansoni and the possibility of geographic expansion of mansoni schistosomiasis

Introduction Human migration and the presence of natural vectors (mollusks) of Schistosoma mansoni are the primary causes of the expansion of mansoni schistosomiasis into southern areas of South America. Water conditions are favorable for the expansion of this disease because of the extensive hydrographic network, which includes the basins of the Paraná and Uruguay rivers and favors mollusk reproduction. These rivers also aid agriculture and tourism in the area. Despite these favorable conditions, natural infection by S. mansoni has not yet been reported in Argentina, Uruguay, or Paraguay. Methods Two species of planorbid from Argentina, Biomphalaria straminea and B. tenagophila, were exposed to the miracidia of five Brazilian strains of S. mansoni. Results Biomphalaria tenagophila (Atalaya, Buenos Aires province) was infected with the SJS strain (infection rate 3.3%), confirming the experimental susceptibility of this Argentinian species. Biomphalaria straminea (Rio Santa Lucía, Corrientes province) was susceptible to two Brazilian strains: SJS (infection rate 6.7%) and Sergipe (infection rate 6.7%). Conclusions These results demonstrate that species from Argentina have the potential to be natural hosts of S. mansoni and that the appearance of foci of mansoni schistosomiasis in Argentina is possible.

Cutaneous and visceral leishmaniasis co-infection in dogs from Rio de Janeiro, Brazil: evaluation by specific PCR and RFLP-PCR assays

Introduction During a diagnostic evaluation of canine visceral leishmaniasis (VL), two of seventeen dogs were found to be co-infected by Leishmania (Viannia) braziliensis and Leishmania (Leishmania) chagasi. Methods Specific polymerase chain reaction (PCR) and restriction fragment length polymorphism-PCR (RFLP-PCR) assays were performed. Results PCR assays for Leishmania subgenus identification followed by RFLP-PCR analysis in biopsies from cutaneous lesions and the spleen confirmed the presence of Leishmania (Viannia) braziliensis and Leishmania (Leishmania) chagasi in those fragments. Conclusions This report reinforces the importance of using serological and molecular techniques in the epidemiological surveillance of canine populations in endemic areas in which both diseases are known to co-exist. In such cases, a reassessment of the control measures is required.

Seroprevalence and seroincidence of Leptospira infection in dogs during a one-year period in an endemic urban area in Southern Brazil

INTRODUCTION: Leptospirosis is a zoonosis that affects both humans and animals. Dogs may serve as sentinels and indicators of environmental contamination as well as potential carriers for Leptospira. This study aimed to evaluate the seroprevalence and seroincidence of leptospirosis infection in dogs in an urban low-income community in southern Brazil where human leptospirosis is endemic. METHODS: A prospective cohort study was designed that consisted of sampling at recruitment and four consecutive trimestral follow-up sampling trials. All households in the area were visited, and those that owned dogs were invited to participate in the study. The seroprevalence (MAT titers ≥100) of Leptospira infection in dogs was calculated for each visit, the seroincidence (seroconversion or four-fold increase in serogroup-specific MAT titer) density rate was calculated for each follow-up, and a global seroincidence density rate was calculated for the overall period. RESULTS: A total of 378 dogs and 902.7 dog-trimesters were recruited and followed, respectively. The seroprevalence of infection ranged from 9.3% (95% CI; 6.7 - 12.6) to 19% (14.1 - 25.2), the seroincidence density rate of infection ranged from 6% (3.3 - 10.6) to 15.3% (10.8 - 21.2), and the global seroincidence density rate of infection was 11% (9.1 - 13.2) per dog-trimester. Canicola and Icterohaemorraghiae were the most frequent incident serogroups observed in all follow-ups. CONCLUSIONS: Follow-ups with mean trimester intervals were incapable of detecting any increase in seroprevalence due to seroincident cases of canine leptospirosis, suggesting that antibody titers may fall within three months. Further studies on incident infections, disease burden or risk factors for incident Leptospira cases should take into account the detectable lifespan of the antibody.

Leishmania, Babesia and Ehrlichia in urban pet dogs: co-infection or cross-reaction in serological methods?

INTRODUCTION: The present study was designed to assess the occurrence of co-infection or cross-reaction in the serological techniques used for detecting the anti-Leishmania spp., -Babesia canis vogeli and -Ehrlichia canis antibodies in urban dogs from an area endemic to these parasites. METHODS: The serum samples from dogs were tested for the Babesia canis vogeli strain Belo Horizonte antigen and Ehrlichia canis strain São Paulo by immunofluorescence antibody test (IFAT) and by anti-Leishmania immunoglobulin G (IgG) antibody detection to assess Leishmania infection. We used the following four commercial kits for canine visceral leishmaniasis: ELISA, IFAT, Dual Path Platform (DPP) (Bio Manguinhos(r)/FIOCRUZ/MS) and a rK39 RDT (Kalazar Detect Canine Rapid Test; Inbios). RESULTS : Of 96 serum samples submitted to serological assays, 4 (4.2%) were positive for Leishmania as determined by ELISA; 12 (12.5%), by IFAT; 14 (14.6%) by rK39 RDT; and 20 (20.8%), by DPP. Antibodies against Ehrlichia and Babesia were detected in 23/96 (23.9%) and 30/96 (31.2%) samples, respectively. No significant association was identified between the results of tests for detecting Babesia or Ehrlichia and those for detecting Leishmania (p-value>0.05). CONCLUSIONS: In the present study, we demonstrated co-infection with Ehrlichia or Babesia and Leishmania in dogs from Minas Gerais (Brazil); we also found that the serological tests that were used did not cross-react.

Spatial analysis of visceral leishmaniasis in the municipality of Rondonópolis, in the Brazilian State of Mato Grosso, from 2003 to 2012: human, canine and vector distribution in areas of disease transmission

INTRODUCTION: Visceral leishmaniasis (VL) is a zoonosis of great importance to public health and is considered a neglected disease by the World Health Organization. The disease has expanded and become more prevalent in urban areas in Brazil. METHODS: Geospatial analyses were performed and thematic maps of the triad of the disease were produced for the study period (2003-2012) in the urban area of the municipality of Rondonópolis in the midwestern State of Mato Grosso (MT), Brazil, TerraView 4.2.2 software was used for the analyses. RESULTS: A total of 87.9% of the 186 confirmed human cases of VL were cured. Children between the ages of 1 and 4 were the most affected. Registered deaths were predominant among adults aged 60 years or older. The urban area of the municipality consists of eight strata and 12 census districts include 237 neighborhoods. All sectors had confirmed cases of VL. During the study period, human cases of the disease were recorded in 90 neighborhoods. The 23 deaths from the disease were distributed in 21 neighborhoods. Sandflies carrying the parasite were captured in 192 out of 200 neighborhoods evaluated for the presence of the VL vector. The presence of dogs carrying the parasite was confirmed in, 140 out of 154 surveyed neighborhoods. CONCLUSIONS: The data demonstrated the endemic nature of VL, with a high percentage of infected children, a high distribution of canine infection, and a wide adaptation and dispersal of the vectors in the urban environment. These results, illustrate the process of urbanization of VL in the municipality of Rondonópolis, MT, Brazil.

Predictive factors for Leishmania infantum infection in dogs examined at a veterinary teaching hospital in Teresina, State of Piauí, Brazil

Abstract: INTRODUCTION: In Brazil, culling of seropositive dogs is one of the recommended strategies to control visceral leishmaniasis. Since infectiousness is correlated with clinical signs, control measures targeting symptomatic dogs could be more effective. METHODS: A cross-sectional study was carried out among 1,410 dogs, predictive models were developed based on clinical signs and an indirect immunofluorescence antibody test. RESULTS: The validated predictive model showed sensitivity and specificity of 86.5% and 70.0%, respectively. CONCLUSIONS: Predictive models could be used as tools to aid control programs in focusing on a smaller fraction of dogs contributing more to infection dissemination.

Pancreatic lesions in acute experimental Chagas' disease

BACKGROUND: Chagas' disease is an endemic tropical affliction found from southern United States to Argentina. The acute phase of this disease is difficult to study in man because the symptoms are non-specific and most cases require no medical assistance. Experimental models have been developed for sequential studies, and intense parasitism in all organs and tissues, including the pancreas, have been detected in the acute phase. PURPOSE: To evaluate the involvement of the pancreas in acute experimental Chagas' disease in a mouse model by histopathological characterization. CASUISTIC AND METHODS: Ten BALBc mice, about 20 g, injected i.p. with 100 000 forms of the Y strain of Trypanosoma cruzi were used. The animals were sacrificed after 14 days of infection. Fragments of pancreas were processed by conventional paraffin embedding and hematoxylin-eosin staining. RESULTS: Ruptured pseudocysts and release of parasites to the extracellular medium caused by necrosis of acinar and duct cells and foci of fat were the most striking histopathological features of acute Chagasic pancreatitis. CONCLUSION: Parasitism is the main cause of acute pancreatitis in Chagas' disease.

Insights into bacterial colony morphology evolution and diversification during infection development in cystic fibrosis

Tese de Doutoramento em Engenharia Biomédica.

Participación de las células supresoras mieloides (CSM) y T regulatorias (Treg) en el control de la infección con Trypanosoma cruzi y el desarrollo de la patología inflamatoria asociada a la infeccion. Role of Myeloid-Derived Suppressor Cells and regulatory T cells in the control of T. cruzi infection and the infection-associated pathology.

La infección de mamíferos con el T. cruzi resulta en diferentes alteraciones inmunológicas que permiten la persistencia crónica del parásito y destrucción inflamatoria progresiva del tejido cardiaco, nervioso y hepático. Los mecanismos responsables de la patología de la enfermedad de Chagas han sido materia de intensa investigación habiéndose propuesto que el daño producido en esta enfermedad puede ser consecuencia de la respuesta inflamatoria del individuo infectado y/o de una acción directa del parásito sobre los tejidos del hospedador. El propósito del presente proyecto es estudiar comparativamente, en dos cepas de ratones con diferente susceptibilidad a la infección y desarrollo de patología, la participación y los mecanismos efectores de las células supresoras mieloides (CSM) y las celulas T regulatorias inducidas por la infección experimental con Trypanosoma cruzi en el control de la infección con este protozoario y en el desarrollo de la patología hepática siendo los objetivos especificos desarrolar: - Investigar la generación y/o reclutamiento de células de CSM en bazo e hígado de ratones infectados con Trypanosoma cruzi y su contribución a la desigual susceptibilidad a la infección y respuesta inmune desarrollada en las cepas de ratones BALB/c y C57BL/6; - Investigar la capacidad de las CSM inducidas por la infección con T. cruzi en bazo e hígado de ratones de ambas cepas para suprimir la respuesta de células T in vitro e indagar sobre los mecanismos de supresión utilizados; - Investigar la generación y/o reclutamiento de células Treg durante la infección experimental con Trypanosoma cruzi, su participación en la desigual susceptibilidad a la infección y respuesta inmune desarrollada en ambas cepas de ratones y los mecanismos de supresión utilizados. - Analizar en tejido hepático o leucocitos infiltrantes la presencia de COX2, PGE2, MMP2 y 9, IL1b, IL6, IDO, IL10 y GM-CSF capaces de inducir la expansión de las CSM; - Dilucidar si la administración del ligando para TLR2 (Pam3CyS) previo a la infección de ratones C57BL/6 (en los cuales se detecta un menor número de CSM) es capaz de modular la respuesta inflamatoria y el daño hepático a través de la inducción de CSM y/o T reg en hígado y bazo. La comprension de los eventos celulares y moleculares que regulan la producción de citoquinas pro- y anti-inflamatorias y otros mediadores, así como el papel de los receptores de la inmunidad innata durante la infección con T. cruzi contribuirá a responder interrogantes que son claves para el diseño de nuevas estrategias de intervención inmune tendientes a preservar los mecanismos de defensa del huésped. Two nonexclusive mechanisms have been proposed to explain the Chagas’s disease pathology: 1) The pathology of the disease seems to be consequence of the inflammatory response triggered for the parasite; or 2) The damage is produced by the parasite direct effect. Recently, we reported that TLR2, TLR4 and TLR9 (innate immune response receptors) are differentially modulated in injured livers from BALB/c (lesser liver pathology) and C57BL/6 (elevated liver pathology) mice during Trypanosoma cruzi infection. The aim of our proposal is the study of role of Myeloid-Derived Suppressor Cells (MDSC) and regulatory T cells in the control of T. cruzi infection and the infection-associated pathology. Our specific aims are: -To study the induction or recruitment of MDSC in splenn and liver of BALB/c and C57BL/6 mice and their relationship with the differential susceptibility and immune response observed in these both mice strains; - To determine the ability and the mechanisms used by the T. cruzi-induced MDSC to suppress the T cell proliferative response; -To study the induction or recruitment of Treg in liver of BALB/c and C57BL/6 mice and their relationship with the differential susceptibility and immune response observed in these both mice strains; -To analize in liver tissue or tissue infiltrating lymphocytes the activation of COX2, PGE2, MMP2 y 9, IL1b, IL6, IDO, IL10 y GM-CSF known to promote the development of MDSC; -To determine whether the treatment with Pam3CyS (TLR2 ligand) is able to modulate the liver inflammatory respose and damage througth the induction of MDSC or Treg.

The interaction of Gram negative bacteria and S. mansoni in mice with experimental Schistosomiasis

Animals (122 mice) were infected each with eighty cercariae of S. mansoni and subsequently challenged intravenously eight weeks later with the following gram-negative organisms. S. typhi, E. coli, Klebsiella-enterobacter species, Proteus mirabilis and Pseudomonas aeruginosa. Enumeration of bacteria in the liver, spleen and blood and S. mansoni from the portal sistem was performed from one to four weeks later in infected animals. A significant difference between infection produced by S. typhi and other gram negative organisms was observed: S. typhi persisted longer in the spleen and liver and could be recovered from S. mansoni worms up to three weeks following bacterial infection. Other gram negative bacteria disappeared from S. mansoni worms after two weeks of initial challenge. Additional animals (51 mice) infected with S. mansoni were given S. typhi, E. coli or sterile saline. After two weeks, animals were sacrificed and the recovery rate of worms from the portal system, and the mesenteric and hepatic oogram were determined. in animals infected with E. coli a significant decrease in the number of worms was observed compared to the saline control group; thirty worms were recovered in the control group compared to two worms in e. coli infected animals. In addition, the patterns of oviposition was significantly different in these latter animals suggesting complete inhibition of this process. Following S. typhi infection the difference in recovery of worms and pattern of oviposition was minimal. These findings suggest a difference in the interaction of various gram negative bacteria and S. mansoni and are consistent with the clinical observation of prolonged salmonella bacteremia in patients with schistosomiasis.

Studies in search of a suitable experimental insect model for xenodiagnosis of hosts with Chagas' disease. 1 - Comparative xenodiagnosis with nine triatomine species of animals with acute infections by Trypanosoma cruzi

In search of a suitable vector species for xenodiagnosis of humans and animals with chronic Chagas' disease we first investigated the reactions of different vector species to acute infection with Trypanosoma cruzi. Vector species utilized in this study were: Triatoma infestans, Rhodnius prolixus and Triatoma dimidiata, all well adapted to human habitats; Triatoma rubrovaria and Rhodnius neglectus both considered totally wild species; Panstrongylus megistus, Triatoma sordida, Triatoma pseudomaculata and Triatoma brasiliensis, all essentially sylvatic but some with domiciliary tendencies and others restricted to peridomestic biotopes with incipient colonization of human houses after successful eradication of T. infestans. Results summarized in Table IV suggest the following order of infectivity among the 9 studied vector species: P. megistus with 97.8% of infected bugs, T. rubrovaria with 95% of positive bugs a close second followed by T. Pseudomaculata with 94.3% and R. neglectus with 93.8% of infected bugs, almost identical thirds. R. prolixus, T. infestans and T. dimidiata exhibited low infection rates of 53.1%, 51.6% and 38.2% respectively, coupled with sharp decreases occuring with aging of infection (Fig. 1). The situation was intermediate in T. brasiliensis and T. sordida infection rates being 76.9% and 80% respectively. Results also point to the existence of a close correlation between prevalence and intensity of infection in that, species with high infection rates ranging from 93.8% to 97.8% exhibited relatively large proportions of insects (27.3% - 33.5%) harbouring very dense populations of T. cruzi. In species with low infection rates ranging from 38.2% to 53.1% the proportion of bugs demonstrating comparable parasite densities was at most 6%. No differences attributable to blood-meal size or to greater susceptibility of indigenous vector species to parasites of their own geographical area, as suggested in earlier...

Studies in search of a suitable experimental insect model for xenodiagnosis of hosts with Chagas' disease: 2 - Attempts to upgrade the reliability and the efficacy of xenodiagnosis in chronic Chagas' disease

In order to upgrade the reliability of xenodiagnosis, attention has been directed towards population dynamics of the parasite, with particular interest for the following factors: 1. Parasite density which by itself is not a research objective, but by giving an accurate portrayal of parasite development and multiplication, has been incorporated in screening of bugs for xenodiagnosis. 2. On the assumption that food availability might increase parasite density, bugs from xenodiagnosis have been refed at biweekly intervals on chicken blood. 3. Infectivity rates and positives harbouring large parasite yields were based on gut infections, in which the parasite population comprised of all developmental forms was more abundant and easier to detect than in fecal infections, thus minimizing the probability of recording false negatives. 4. Since parasite density, low in the first 15 days of infection, increases rapidly in the following 30 days, the interval of 45 days has been adopted for routine examination of bugs from xenodiagnosis. By following the enumerated measures, all aiming to reduce false negative cases, we are getting closer to a reliable xenodiagnostic procedure. Upgrading the efficacy of xenodiagnosis is also dependent on the xenodiagnostic agent. Of 9 investigated vector species, Panstrongylus megistus deserves top priority as a xenodiagnostic agent. Its extraordinary capability to support fast development and vigorous multiplication of the few parasites, ingested from the host with chronic Chagas' disease, has been revealed by the strikingly close infectivity rates of 91.2% vs. 96.4% among bugs engorged from the same host in the chronic and acute phase of the disease respectively (Table V), the latter comporting an estimated number of 12.3 x 10[raised to the power of 3] parasites in the circulation at the time of xenodiagnosis, as reported previously by the authors (1982).