The leptospiral LigA protein consists of 13 bacterial immunoglobulin-like (Big) domains and is the only purified recombinant subunit vaccine that has been demonstrated to protect against lethal challenge by a clinical isolate of Leptospira interrogans in the hamster model of leptospirosis. We determined the minimum number and location of LigA domains required for immunoprotection. Immunization with domains 11 and 12 was found to be required but insufficient for protection. Inclusion of a third domain, either 10 or 13, was required for 100% survival after intraperitoneal challenge with Leptospira interrogans serovar Copenhageni strain Fiocruz L1-130. As in previous studies, survivors had renal colonization; here, we quantitated the leptospiral burden by qPCR to be 1.2×103 to 8×105 copies of leptospiral DNA per microgram of kidney DNA. Although renal histopathology in survivors revealed tubulointerstitial changes indicating an inflammatory response to the infection, blood chemistry analysis indicated that renal function was normal. These studies define the Big domains of LigA that account for its vaccine efficacy and highlight the need for additional strategies to achieve sterilizing immunity to protect the mammalian host from leptospiral infection and its consequences.

BackgroundLeptospirosis is a zoonosis caused by highly motile, helically shaped bacteria that penetrate the skin and mucous membranes through lesions or abrasions, and rapidly disseminate throughout the body. Although the intraperitoneal route of infection is widely used to experimentally inoculate hamsters, this challenge route does not represent a natural route of infection.Methodology/Principal FindingsHere we describe the kinetics of disease and infection in hamster model of leptospirosis after subcutaneous and intradermal inoculation of Leptospira interrogans serovar Copenhageni, strain Fiocruz L1-130. Histopathologic changes in and around the kidney, including glomerular and tubular damage and interstitial inflammatory changes, began on day 5, and preceded deterioration in renal function as measured by serum creatinine. Weight loss, hemoconcentration, increased absolute neutrophil counts (ANC) in the blood and hepatic dysfunction were first noted on day 6. Vascular endothelial growth factor, a serum marker of sepsis severity, became elevated during the later stages of infection. The burden of infection, as measured by quantitative PCR, was highest in the kidney and peaked on day 5 after intradermal challenge and on day 6 after subcutaneous challenge. Compared to subcutaneous challenge, intradermal challenge resulted in a lower burden of infection in both the kidney and liver on day 6, lower ANC and less weight loss on day 7.Conclusions/SignificanceThe intradermal and subcutaneous challenge routes result in significant differences in the kinetics of dissemination and disease after challenge with L. interrogans serovar Copenhageni strain Fiocruz L1-130 at an experimental dose of 2×106 leptospires. These results provide new information regarding infection kinetics in the hamster model of leptospirosis.

-INTRODUÇÃOO preparo e comércio de alimentos por ambulantes nas ruas das grandes cidades é um fenômeno mundial e tem especial importância nos países em desenvolvimento, onde constitui uma atividade econômica alternativa para os desempregados [4]. Devido a problemas sócio-econômicos de vários países, este setor da economia tem crescido bastante nas últimas décadas. Este fato, junto com a urbanização e o crescimento da população, faz com que se espere um crescimento ainda maior deste tipo de comércio [26].Estudos realizados na América Latina, estimam que 25 a 30% do gasto familiar nos grandes centros urbanos se destinam ao consumo de alimentos comercializados por vendedores de alimentos ambulantes [12]. Geralmente os alimentos vendidos por ambulantes são produtos prontos para o consumo, preparados no próprio local de comercialização que está situado em regiões de grande afluência de público [7]. Os produtos comercializados diferem entre os diversos países e culturas e também apresentam grande importância do ponto de vista turís-tico, pois muitos são produtos típicos de uma região, sendo muito apreciados pelos turistas.Em contraponto este tipo de comércio pode constituir um risco à saúde da população, pois os alimentos podem ser facilmente contaminados com microrganismos patogênicos, devido as condições inadequadas do local de preparo e a falta de conhecimentos de técnicas de manipulação higiênica por parte dos comerciantes. Além disso, muitos estabelecimentos de comércio ambulante não contam com sistema de abastecimento de água tratada, o que dificulta a higienização correta dos utensílios utilizados no preparo das refeições. Utensíli-os, superfícies e equipamentos insuficientemente limpos representam um risco de contaminação, especialmente para alimentos cozidos que não serão consumidos imediatamente [24].Nos países em desenvolvimento, onde é comum este tipo de comércio, produtos prontos para o consumo não RESUMOO comércio de alimentos prontos para o consumo por vendedores ambulantes pode constituir um alto risco para a saúde dos consumidores, visto que as pessoas envolvidas nesta atividade geralmente não tem preparo para a manipulação correta de alimentos. Neste trabalho foram investigadas as condições de preparo e a qualidade higiênico-sanitária de lanches comercializados por vendedores ambulantes em Pelotas. Foram colhidas de 60 estabelecimentos, uma amostra de água, uma da superfície de manuseio e um lanche (cachorroquente). Nos cachorros-quentes foram realizadas contagens de bactérias aeróbias mesófilas (BAM), Staphylococcus coagulase positiva (STA), coliformes totais (CT), coliformes a 45ºC (CF), e investigada a presença de Salmonella sp. Na água e superfície foram realizadas contagens de BAM, CT e CF. Entre as 60 amostras de cachorros-quentes analisadas, 53%, 48%, 37% e 25% estavam fora do padrão para CT, BAM, STA e CF, respectivamente. Em nenhuma amostra foi detectada a presença de Salmonella. As amostras de água apresentaram apenas 3 (5%) amostras fora do padrão para BAM, e 27% e 23% fora do padrão para...

BackgroundImmunomagnetic separation (IMS) and immunoassays are widely used for pathogen detection. However, novel technology platforms with highly selective antibodies are essential to improve detection sensitivity, specificity and performance. In this study, monoclonal antibodies (MAbs) against Internalin A (InlA) and p30 were generated and used on paramagnetic beads of varying diameters for concentration, as well as on fiber-optic sensor for detection.ResultsAnti-InlA MAb-2D12 (IgG2a subclass) was specific for Listeria monocytogenes and L. ivanovii, and p30-specific MAb-3F8 (IgM) was specific for the genus Listeria. At all bacterial concentrations (103–108 CFU/mL) tested in the IMS assay; the 1-μm diameter MyOne beads had significantly higher capture efficiency (P < 0.05) than the 2.8-μm diameter M-280 beads with both antibodies. The highest capture efficiency for MyOne-2D12 (49.2% for 105 CFU/mL) was significantly higher (P < 0.05) than that of MyOne-3F8 (16.6 %) and Dynabeads anti-Listeria antibody (9 %). Furthermore, capture efficiency for MyOne-2D12 was highly specific for L. monocytogenes and L. ivanovii. Subsequently, we captured L. monocytogenes by MyOne-2D12 and MyOne-3F8 from hotdogs inoculated with mono- or co-cultures of L. monocytogenes and L. innocua (10–40 CFU/g), enriched for 18 h and detected by fiber-optic sensor and confirmed by plating, light-scattering, and qPCR assays. The detection limit for L. monocytogenes and L. ivanovii by the fiber-optic immunosensor was 3 × 102 CFU/mL using MAb-2D12 as capture and reporter antibody. Selective media plating, light-scattering, and qPCR assays confirmed the IMS and fiber-optic results.ConclusionsIMS coupled with a fiber-optic sensor using anti-InlA MAb is highly specific for L. monocytogenes and L. ivanovii and enabled detection of these pathogens at low levels from buffer or food.

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