Wide distribution of Trypanosoma cruzi-infected triatomines in the State of Bahia, Brazil.

BACKGROUND: The identification of Trypanosoma cruzi and blood-meal sources in synanthropic triatomines is important to assess the potential risk of Chagas disease transmission. We identified T. cruzi infection and blood-meal sources of triatomines caught in and around houses in the state of Bahia, northeastern Brazil, and mapped the occurrence of infected triatomines that fed on humans and domestic animals. METHODS: Triatominae bugs were manually captured by trained agents from the Epidemiologic Surveillance team of Bahia State Health Service between 2013 and 2014. We applied conventional PCR to detect T. cruzi and blood-meal sources (dog, cat, human and bird) in a randomized sample of triatomines. We mapped triatomine distribution and analyzed vector hotspots with kernel density spatial analysis. RESULTS: In total, 5906 triatomines comprising 15 species were collected from 127 out of 417 municipalities in Bahia. The molecular analyses of 695 triatomines revealed a ~10% T. cruzi infection rate, which was highest in the T. brasiliensis species complex. Most bugs were found to have fed on birds (74.2%), and other blood-meal sources included dogs (6%), cats (0.6%) and humans (1%). Trypanosoma cruzi-infected triatomines that fed on humans were detected inside houses. Spatial analysis showed a wide distribution of T. cruzi-infected triatomines throughout Bahia; triatomines that fed on dogs, humans, and cats were observed mainly in the northeast region. CONCLUSIONS: Synanthropic triatomines have a wide distribution and maintain the potential risk of T. cruzi transmission to humans and domestic animals in Bahia. Ten species were recorded inside houses, mainly Triatoma sordida, T. pseudomaculata, and the T. brasiliensis species complex. Molecular and spatial analysis are useful to reveal T. cruzi infection and blood-meal sources in synanthropic triatomines, identifying areas with ongoing threat for parasite transmission and improving entomological surveillance strategies.

MCM3: A Novel Proliferation Marker in Oral Squamous Cell Carcinoma.

The present study sought to evaluate and compare the immunoexpression of proteins minichromosome maintenance (MCM) 3 and Ki-67 in oral squamous cell carcinoma (OSCC) to assess the potential of these proteins as markers of cellular proliferation. Twenty-eight cases of OSCC, 9 of tumor-free resection margins (TM), and 4 of non-neoplastic oral mucosa (NNM) were subjected to immunohistochemistry to detect the expression of proteins MCM3 and Ki-67. All OSCCs demonstrated positivity for both proteins. In these tumors, greater MCM3 immunoreactivity was observed in comparison with Ki-67, whereas TMs and NNMs exhibited greater Ki-67 expression compared with MCM3. The immunoexpression of Ki-67 seemed to be influenced by the inflammatory process, particularly in TM and NNM. Our findings indicate that although both MCM3 and Ki-67 represent reliable markers of cellular proliferation in OSCC, as MCM3 expression does not appear to be influenced by external factors, this protein may emerge as a novel marker of cellular proliferation in these types of tumors.

Real-Time PCR Reveals Rapid Dissemination of Leptospira interrogans after Intraperitoneal and Conjunctival Inoculation of Hamsters.

The pathogen Leptospira interrogans is a highly motile spirochete that causes acute and fulminant infections in humans and other accidental hosts. Hematogenous dissemination is important for infection by the pathogen but remains poorly understood because few animal model studies have used sensitive tools to quantify the bacteria. We evaluated the kinetics of leptospiral infection in Golden Syrian hamsters by a sensitive quantitative real-time PCR (TaqMan) with lipl32 as the target gene. The dissemination and bacterial burden were measured after intraperitoneal infection with a high dose (10(8)) or low dose (2.5 × 10(2)) of leptospires. We also examined the conjunctival challenge route to mimic the natural history of infection. Quantification of leptospires in perfused animals revealed that pathogens were detected in all organs of intraperitoneally infected hamsters, including the eye and brain, within 1 h after inoculation of 10(8) virulent L. interrogans bacteria. Peaks of 10(5) to 10(8) leptospires per gram or per milliliter were achieved in blood and all tissues between day 4 and day 8 after intraperitoneal inoculation of high- and low-dose challenges, respectively, coinciding with macroscopic and histological changes. The conjunctival route resulted in a delay in the time to peak organ burden in comparison to intraperitoneal infection, indicating that although infection could be established, penetration efficiency was low across this epithelial barrier. Surprisingly, infection with a large inoculum of high-passage-number attenuated L. interrogans strains resulted in dissemination to all organs in the first 4 days postinfection, albeit with a lower burden, followed by clearance from the blood and organs 7 days postinfection and survival of all animals. These results demonstrate that leptospiral dissemination and tissue invasion occur. In contrast, development of a critical level of tissue burden and pathology are dependent on the virulence of the infecting strain.

Detection and quantification of Leptospira interrogans in hamster and rat kidney samples: immunofluorescent imprints versus real-time PCR.

A major limitation in the clinical management and experimental research of leptospirosis is the poor performance of the available methods for the direct detection of leptospires. In this study, we compared real-time PCR (qPCR), targeting the lipL32 gene, with the immunofluorescent imprint method (IM) for the detection and quantification of leptospires in kidney samples from the rat and hamster experimental models of leptospirosis. Using a virulent strain of Leptospira interrogans serovar Copenhageni, a chronic infection was established in the rat model, which were euthanized 28 days post-infection, while the hamster model simulated an acute infection and the hamsters were euthanized eight days after inoculation. Leptospires in the kidney samples were detected using culture isolation, qPCR and the IM, and quantified using qPCR and the IM. In both the acute and chronic infection models, the correlation between quantification by qPCR and the IM was found to be positive and statistically significant (P<0.05). Therefore, this study demonstrates that the IM is a viable alternative for not only the detection but also the quantification of leptospires, particularly when the use of qPCR is not feasible.