Resumo: A distribuição das células envolvidas na resposta imune em glândulas sexuais acessórias de carneiros experimentalmente infectados com Actinobacillus seminis foi estudada. Doze carneiros de um ano de idade foram experimentalmente infectados via intrauretral (IU) (n=4) e via intraepididimal (IE) (n=4) e quatro animais controles (CON) foram utilizados. Os animais foram abatidos 35 dias após a inoculação, amostras foram retiradas das glândulas sexuais acessórias e testes bacteriológicos e histopatológicos foram realizados. A presença de linfócitos CD4, CD8 e TCRγδ (WC1), células CD45RO, macrófagos (CD14), células dendríticas (CD1b) e células contendo IgA, IgG and IgM (IgCC) foi determinada. Os animais do grupo IE desenvolveram epididimite clínica. Não foram visualizadas lesões nos carneiros do grupo IU, dois dos CON inoculados intraepididimalmente desenvolveram pequenas lesões no epidídimo. Isolados de A. seminis foram obtidos de 6:16 (37,5%) nas glândulas sexuais acessórias no grupo IE mas não nos grupos IU e CON. No grupo CON células contendo IgA and IgM predominaram nas glândulas bulbouretrais e na próstata e foram escassas ou ausentes nas vesículas e na ampola. Um incremento significativo de células contendo IgA, IgG and IgM foi confirmado nas vesículas seminais, na ampola e nas glândulas bulbouretrais no grupo IE. Nos grupos IE e IU foi evidenciado um aumento em CD4, CD8, WC1, CD45RO e CD14 nas vesículas e ampola. As células dendríticas CD1b estavam presentes na ampola e nas vesículas com processo inflamatório. A. seminis induziu uma resposta imune local nos grupos IE e IU. Estes resultados indicam um padrão diferente de células imunes infiltrantes nas glândulas sexuais acessórias de carneiros infectados por A. seminis.

Abstract: The distribution of cells involved in the immune response in accessory sex glands of rams experimentally infected with Actinobacillus seminis was studied. Twelve one-year old rams were experimentally infected by intraurethral (IU) (n=4) and intraepididymal (IE) (n=4) route, and four control (CON) animals were used. The animals were slaughtered 35 days post-inoculation, samples were taken from accessory sex glands, and bacteriology and histopathology tests were performed. The presence of CD4, CD8 and TCRγδ (WC1) lymphocytes, CD45RO cells, macrophages (CD14), dendritic cells (CD1b), IgA-, IgG- and IgM-containing cells (IgCC) was determined. Animals of the IE group developed clinical epididymitis. No lesions were seen in rams of the IU group; two of the intraepididymal inoculated CON developed small lesions in the epididymis. A. seminis isolates were achieved from 6:16 (37.5%) accessory sex glands in the IE group, but not in the IU and CON groups. In the CON group, IgA- and IgM- containing cells predominated in the bulbourethral glands and the disseminated prostate, and they were scarce or null in the vesicles and ampullae. A significant increase of IgA-, IgG- and IgM- containing cells was confirmed in the seminal vesicles, the ampullae and the bulbourethral glands in the IE group. In the IE and IU groups, an increase in CD4, CD8, WC1, CD45RO and CD14 was evidenced in the vesicles and ampullae. CD1b dendritic cells were present in the ampullae and vesicles with inflammatory processes. A. seminis triggered a local immune response in the IE and IU groups. These results indicate a different pattern of infiltrating immune cells in the accessory sex glands of infected A. seminis rams.

The virulence of 12 Listeria monocytogenes strains from goat and goat by-products was evaluated. Variable virulence levels were found when compared the relative virulence (mortality, %) and the ability to grow infect the spleen of BALB/c mice inoculated intravenously or intragastrically. Two strains from feed produced 100 % mortality, by either inoculation route and seven strains, including those from feed, infected the spleen with both inoculation routes. A strain inoculated intravenously without causing death in the mice produced spleen counts similar to those obtained in dead mice, this could indicate that a high count of the microorganism in the spleen not necessarily ensures its virulence. We also tested the capacity to infect J774 macrophages and Caco-2 epithelial cells, finding higher invasion and intracellular growth in Caco-2 with respect the macrophages and showing quadratic growth kinetics in both cell lines. Only the two feed strains did not show a significant difference from the control strain (P>0.05) regarding kinetic parameters and invasion percentage in both cell lines. Although all isolates were able to invade Caco-2 cells, not all isolates were able to invade the spleen after the intragastrically inoculation. Two goats feed strains were the most virulent, representing a risk for animal and human health.

Se evaluó la virulencia de cepas de Listeria monocytogenes todas de serotipo 4b procedentes de cabras y sus derivados. Se observaron niveles de virulencia variables cuando se comparó la virulencia relativa (porcentaje de letalidad) en ratones BALB/c inoculados vía intravenosa o intragástrica y su capacidad para infectar macrófagos J774A.1, y células epiteliales Caco-2. Dos cepas obtenidas de alimento de cabras produjeron 100 % de letalidad por ambas vías de inoculación y no mostraron diferencia significativa con la cepa testigo (P>0.05) respecto al porcentaje de invasión y a los parámetros de la cinética de crecimiento cuadrática observada en ambas líneas celulares. Si bien todas las cepas lograron invadir las células Caco-2, solamente algunas consiguieron invadir el bazo después de la inoculación por vía intragástrica. Las dos cepas provenientes de alimento de cabras fueron las más virulentas, representando un riesgo para la salud humana y animal, ya que pueden ser diseminadas en el hato y de este a otras explotaciones o a las instalaciones donde se elaboran alimentos.

Paratuberculosis is a chronic granulomatous enteritis caused by Mycobacterium avium subspecies paratuberculosis (Map), which affects wild and domestic ruminants. Map is shed in feces from infected animals. Transmission of the infection takes place by oral ingestion of the bacterium from contaminated food and water with feces. With the objective to establish a paratuberculosis diagnosis in ovine by nested-PCR from fecal samples, 204 fecal and serum ovine samples were studied. Feces were evaluated by nested-PCR and bacterial culture, serum samples were analyzed by agar gel immunodiffusion (AGID). Nested-PCR yielded a 210 bp amplification product that corresponds to Map-IS900, in 61 out of 204 samples. From these, 43 were from AGID positive animals and 18 from negative animals. Seventeen Map strains were isolated by bacterial culture and AGID detected 91 positive animals. Nested-PCR allowed to detect, sooner, greater number of animals shedding bacillus, even when they had resulted negative to the serological test. This result is considered important because generally these animals, while remaining in the farm, constitute the main source of infection for the herd. Nested-PCR should be considered as an alternative, when a prompt result is required to know the health status of the herd with respect to paratuberculosis.

La paratuberculosis es una enteritis granulomatosa de curso crónico ocasionada por Mycobacterium avium subespecie paratuberculosis (Map), afecta a rumiantes domésticos y silvestres. Map es excretada en las heces de animales que desarrollan la enfermedad, y la transmisión de la infección se da mediante la ingestión de alimentos y agua contaminados por heces de animales infectados. Con el objetivo de establecer el diagnóstico de paratuberculosis en ovinos por medio de la PCR-anidada a partir de muestras de heces, se trabajaron 204 muestras de heces y sueros de ovinos; las heces se evaluaron por PCR-anidada y cultivo bacteriológico, las muestras de sueros fueron analizadas por medio de inmunodifusión en agar gel (lDGA). Con la PCR-anidada se obtuvo un producto de amplificación 210 pb que corresponde a la IS900 de Map, en 61 de las 204 muestras. De éstas, 43 eran de animales positivos a IDGA y 18 negativos. Mediante cultivo bacteriológico se aislaron 17 cepas de Map; en este contexto, la IDGA detectó a 91 animales como positivos. La PCR-anidada permitió detectar en menor tiempo a mayor cantidad de animales que estaban eliminando al bacilo, aun cuando habían resultado negativos a la prueba serológica; este resultado se considera importante, ya que generalmente estos animales, al permanecer dentro de la granja, constituyen la principal fuente de infección para el rebaño. Se debe considerar a la PCR-anidada como alternativa, cuando se requiera el diagnóstico en breve tiempo, para conocer el estado sanitario del rebaño con respecto a paratuberculosis.