Experimental leptospirosis in capybaras (Hydrochaeris hydrochaeris) infected with Leptospira interrogans serovar Pomona.

Capybara (Hydrochaeris hydrochaeris), the largest rodent in the world, is widely distributed in South America. These animals live in areas with abundant water, which makes them a potential reservoir for Leptospira. The objective of this study was to investigate seroconversion, leptospiremia, and leptospiruria in capybaras experimentally infected with a virulent strain of Leptospira interrogans serovar Pomona. Seven capybaras were used: one control and six infected. Agglutinins against serovar Pomona were initially detected in serum 6 or 7 day after innoculation with Leptospira (10(9)-10(11) organisms, given i.v.), peaked (titer, approximately 3,200) between 9 and 27 day, and were still present at 83 day (end of study). The earliest and latest isolation of leptospires from the blood was from 2-12 day and from urine, 9-19 day after exposure. However, polymerase chain reaction and isolation results from kidney and liver samples were negative for leptospires. The control animal tested negative on all diagnostic tests. Hence, the capybara can serve as a host for Leptospira.

Evaluation of protein spectra cluster analysis for Streptococcus spp. identification from various swine clinical samples.

Traditional microbiological methods enable genus-level identification of Streptococcus spp. isolates. However, as the species of this genus show broad phenotypic variation, species-level identification or even differentiation within the genus is difficult. Herein we report the evaluation of protein spectra cluster analysis for the identification of Streptococcus species associated with disease in swine by means of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). A total of 250 S. suis-like isolates obtained from pigs with clinical signs of encephalitis, arthritis, pneumonia, metritis, and urinary or septicemic infection were studied. The isolates came from pigs in different Brazilian states from 2001 to 2014. The MALDI-TOF MS analysis identified 86% (215 of 250) as S. suis and 14% (35 of 250) as S. alactolyticus, S. dysgalactiae, S. gallinaceus, S. gallolyticus, S. gordonii, S. henryi, S. hyointestinalis, S. hyovaginalis, S. mitis, S. oralis, S. pluranimalium, and S. sanguinis. The MALDI-TOF MS identification was confirmed in 99.2% of the isolates by 16S rDNA sequencing, with MALDI-TOF MS misidentifying 2 S. pluranimalium as S. hyovaginalis. Isolates were also tested by a biochemical automated system that correctly identified all isolates of 8 of the 10 species in the database. Neither the isolates of the 3 species not in the database ( S. gallinaceus, S. henryi, and S. hyovaginalis) nor the isolates of 2 species that were in the database ( S. oralis and S. pluranimalium) could be identified. The topology of the protein spectra cluster analysis appears to sustain the species phylogenetic similarities, further supporting identification by MALDI-TOF MS examination as a rapid and accurate alternative to 16S rDNA sequencing.

Towards multilocus sequence typing of the Leishmania donovani complex: resolving genotypes and haplotypes for five polymorphic metabolic enzymes (ASAT, GPI, NH1, NH2, PGD).

Multilocus enzyme electrophoresis is the gold standard for identification of Leishmania species and strains. Drawbacks include: only amino acid polymorphisms affecting electrophoretic mobility are detected; distinct allozymes can have coincident mobilities; few characters are available; and parasites must be cultured in bulk. So far, thousands of Leishmania strains have been phenotyped by multilocus enzyme electrophoresis. Here, we sequence enzyme-coding genes to provide a PCR-based higher resolution equivalent of multilocus enzyme electrophoresis, particularly for Leishmania infantum. Of 15 enzymes used for multilocus enzyme electrophoresis (MON typing) we have sequenced aspartate aminotransferase, glucose-6-phosphate isomerase, nucleoside hydrolase 1, nucleoside hydrolase 2 and 6-phosphogluconate dehydrogenase. Heterozygous alleles were common, with multiple heterozygous sites within a single locus for several of the genes. Haplotypes were resolved by allele-specific PCR and allele-specific sequencing. Heterozygous haplotypes conformed to the haplotypes of putative parents. One strain appeared to be hybrid across two genetic groups of the Leishmania donovani complex. In most cases, a single amino acid polymorphism was responsible for change in enzyme mobility. Some indistinguishable phenotypes were produced by distinct genotypes. Silent genetic polymorphisms provided enhanced discrimination over multilocus enzyme electrophoresis, for example, by subdividing the zymodeme MON-1. The PCR-based genotyping that we describe could be applied directly to clinical samples or to small volume cultures and in a multilocus sequence typing format. Furthermore, it can be used to detect recombination indirectly and for population genetics studies.

Comparative efficacy of two single-dose bacterins in the control of Mycoplasma hyopneumoniae in swine raised under commercial conditions in Brazil.

A field study was conducted in Brazil to evaluate the efficacy of single vaccination of pigs with two bacterins to prevent Mycoplasma hyopneumoniae lung lesions. The first (T1) treatment group (174 pigs) was injected with 2 mL of saline solution; group T2 (177 pigs) with 2 mL of bacterin A, and group T3 (174 pigs) with 2 mL of bacterin B. On days-on-test (DOT) 0, 35, 66, 97 and 125, blood samples and tonsil swabs were collected from selected pigs for antibody determination (indirect ELISA) and PCR assay for the presence of M. hyopneumoniae. Pigs were slaughtered on DOT 126-129 and lung lesions were scored blindly. Bacterin A vaccinated pigs had significantly (P < or = 0.05) lower lung lesion scores (0.2%) than bacterin B (0.4%) or saline-treated pigs (1.2%); there was also a significantly lower (P < or = 0.05) number of pigs with lung lesions (27.1%), than bacterin B (38.2%) or saline-treated (55.4%) pigs. The two vaccines had similar (P>0.05) results in terms of mean weight gain, average daily weight gain, feed efficiency, frequency of PCR positives, and there was similar antibody conversion (ELISA). It was concluded that although the productivity parameters and antibody conversions were similar, bacterin A was more effective in preventing and reducing the severity of lung lesions than bacterin B.

Genetic diversity of Streptococcus suis serotype 2 isolated from pigs in Brazil.

Streptococcus suis is an emerging zoonotic pathogen that causes septicemia, meningitis, arthritis, and pneumonia in swine and humans. The present study aimed to characterize the genetic diversity of S. suis serotype 2 isolated from pigs showing signs of illness in Brazil using pulsed-field gel electrophoresis (PFGE), single-enzyme amplified fragment length polymorphism (SE-AFLP), and profiling of virulence-associated markers. A total of 110 isolates were studied, 62.7% of which were isolated from the central nervous system and 19.1% from the respiratory tract. Eight genotypes were obtained from the combination of virulence genes, with 43.6% and 5.5% frequencies for the mrp (+) /epf (+) /sly (+) and mrp (-) /epf (-) /sly (-) genotypes, respectively. The presence of isolates with epf gene variation with higher molecular weight also appears to be a characteristic of Brazilian S. suis serotype 2. The PFGE and SE-AFLP were able to type all isolates and, although they presented a slight tendency to cluster according to state and year of isolation, it was also evident the grouping of different herds in the same PFGE subtype and the existence of isolates originated from the same herd classified into distinct subtypes. No further correlation between the isolation sites and mrp/epf/sly genotypes was observed.

Streptococcus suis est un agent pathogène zoonotique en émergence responsable de septicémies, des méningites, d'arthrites, et de pneumonies chez les porcs et les humains. La présente étude visait à caractériser la diversité génétique de souches de S. suis sérotype 2 isolées au Brésil de porcs montrant des signes de maladie à l'aide des techniques suivantes : électrophorèse en champs pulsés (PFGE), polymorphisme des fragments amplifiés par un enzyme unique (SE-AFLP), et profilage des marqueurs de virulence. Un total de 110 isolats a été étudié, 62,7 % isolats provenant du système nerveux central et 19,1 % du tractus respiratoire. Huit génotypes furent obtenus de la combinaison de gènes de virulence, avec des fréquences de 43,6 % et 5,5 % pour les génotypes mrp+/epf+/sly+ et mrp−/epf−/sly−, respectivement. La présence d'isolats avec la variation du gène epf et un poids moléculaire plus élevé semble être également une caractéristique de S. suis sérotype 2 d'origine brésilienne. Les méthodes PFGE et SE-AFLP ont été en mesure de permettre le typage de tous les isolats et, bien qu'il y avait une légère tendance à se regrouper selon l'état et l'année d'isolement, il était également évident qu'il y avait du regroupement d'isolats provenant de différents troupeaux dans le même sous-type de PFGE et de l'existence d'isolats provenant du même troupeau classifiés dans des sous-types différents. Aucune autre corrélation entre le site d'isolement et les génotypes de mrp/epf/sly ne fut observée.(Traduit par Docteur Serge Messier).

Characterization of Streptococcus suis through serotyping, SE-AFLP and virulence profile / Caracterização de Streptococcus suis por sorotipagem, SE-AFLP e perfil de virulência

Streptococcus suis is one of most important pathogens in the swine industry worldwide. Despite its importance, studies of S. suis characterization in South America are still rare. This study evaluates S. suis isolates from distinct Brazilian states, from 1999 to 2004, and its molecular and serological characterization. A total of 174 isolates were studied. S. suis identification was confirmed by PCR and isolates were further serotyped and genotyped by SE-AFLP and amplification of virulence markers. Serotype 1, 2, 3, 4, 7, 18, 22 and 32 were identified among the studied isolates, and only 4% were characterized as non-typeable. The mrp+/epf+/sly+ genotype was the most frequent. The SE-AFLP analysis resulted in 29 patterns distributed in three main clusters with over 65% of genetic similarity. Isolates presented a slight tendency to cluster according to serotype and origin; however, no further correlation with virulence genotypes was observed.(AU)

Streptococcus suis é um dos patógenos de maior importância para indústria suinícola mundial. Apesar de sua importância, a caracterização de isolados de S. suis na América do Sul ainda é pouco descrita. O presente estudo descreve a avaliação de isolados de S. suis provenientes de diferentes Estados brasileiros, e sua caracterização sorológica e molecular. Foram avaliados 174 isolados de S. suis e os mesmos foram submetidos a SE-AFLP e pesquisa de marcadores de virulência. Os sorotipos 1, 2, 3, 4, 7, 18, 22 e 32 foram identificados dentre os isolados estudados e apenas 4% foram caracterizados como não tipáveis. O perfil de virulência mrp+/epf+/sly+ foi o mais frequente. A análise do SE-AFLP resultou em 29 perfis distribuídos em três grupos principais com mais de 65% de similaridade genética. Os isolados apresentaram tendência de se agrupar segundo origem e sorotipo; no entanto, não foi observada correlação entre os grupamentos e os perfis de virulência.(AU)