Genetically Related Mycobacterium bovis Strains Displayed Differential Intracellular Growth in Bovine Macrophages.

Molecular typing of bacterial isolates provides a powerful approach for distinguishing Mycobacterium bovis (M. bovis) genotypes. It is known that M. bovis strain virulence plays a role in prevalence and spread of the disease, suggesting that strain virulence and prevailing genotypes are associated. However, it is not well understood whether strain virulence correlates with particular genotypes. In this study, we assessed the in vitro intracellular growth of 18 M. bovis isolates in bovine macrophages as an indicator of bacterial virulence and sought a relationship with the genotype identified by spoligotyping. We found 14 different spoligotypes-11 were already known and three spoligotypes had never been reported before. We identified 2 clusters that were phylogenetically related, containing 10 and 6 strains, respectively, and 2 orphan strains. Intracellular growth and phagocytic rates of 18 M. bovis strains were heterogeneous. Our results suggest that M. bovis intracellular growth and phagocytosis are independent of the bacterial lineage identified by spoligotyping.

Molecular epidemiology of human cases of tuberculosis by Mycobacterium bovis in Mexico.

The purpose of this study was to evaluate the role of Mycobacterium bovis in human cases of tuberculosis (TB) in an endemic area of the disease in cattle. Sputum, urine and other tissue samples were obtained from: (1) TB-symptomatic patients, (2) dairy farm workers and (3) abattoir workers. Samples of macroscopic lesions suspicious of TB were also obtained from cattle at slaughter in the same geographic area. A total of 562 human samples were collected: 255 from symptomatic patients, 218 from farm workers and 93 from abattoir workers. Samples were analysed by the bacillus acido-alcohol resitant (BAAR) and polymerase chain reaction (PCR) tests and cultured in Stonebrink and Löwenstein-Jensen. Spoligotyping was performed in all isolates obtained by culture and the DNA obtained by PCR. From the total number of human cases, 34 (6%) showed M. bovis spoligotype; eight spoligotypes from cattle showed an identical pattern to three spoligotypes from humans; a different set of spoligotypes from cattle (n = 8) had only one spacer difference to a set of spoligotypes from humans (n = 2). These results provide further evidence that infected cattle represent a risk to public health and support previous reports about the role of M. bovis in Mexican patients. There is no doubt that genotyping M. bovis isolates collected from cattle may have a substantial impact on our understanding of the epidemiology of TB.

[Molecular epidemiology of cattle and human tuberculosis in Mexico]. / Epidemiología molecular de las tuberculosis bovina y humana en una zona endémica de Querétaro, México.

OBJECTIVE: The purpose of the study was to determine the role of bovine TB in cases of human TB. MATERIAL AND METHODS: Two-hundred and fifty-five samples from symptomatic patients were included in the study. All samples were cultured in Stonebrink and Lowënstein-Jensen media and analyzed using a nested PCRMPB70. The molecular analysis was performed by spoligotyping. RESULTS: From 255 samples, 74 were PCR-positive and 20 were culture-positive. From 94 samples positive to PCR or to isolation, 66 (70%) showed a spoligotype compatible with M. tuberculosis, and 13 (13.8%) with M. bovis. Four fingerprints of M. bovis from humans were identical to the fingerprints of M. bovis from cattle in the same region. CONCLUSIONS: Our study shows that M. bovis plays an important role in the epidemiology of TB in humans and that TB in cattle represents a risk to public health.

Epidemiología molecular de las tuberculosis bovina y humana en una zona endémica de Querétaro, México / Molecular epidemiology of cattle and human tuberculosis in Mexico

OBJETIVO: Determinar el papel de la tuberculosis bovina en la tuberculosis humana. MATERIAL Y MÉTODOS: Se analizaron 255 muestras de pacientes sintomáticos, sembradas en medios de Stonebrink y Lõwenstein-Jensen y analizadas por PCRMPB70 anidada y luego por spoligotyping. RESULTADOS: De las 255 muestras, 74 fueron positivas a la PCR y 20 al aislamiento: de las primeras, 58 (78 por ciento) mostraron espoligotipo de M. tuberculosis y 5 (6.7 por ciento) de M. bovis; de las segundas, 8 (47 por ciento) revelaron espoligotipo de M. tuberculosis y 8 (47 por ciento) de M. bovis. De las 94 muestras positivas al aislamiento o PCR, 66 (70 por ciento) correspondieron a M. tuberculosis y 13 (13.8 por ciento) a M. bovis. Los patrones moleculares de cuatro muestras de M. bovis de seres humanos fueron idénticos a los de las cepas de M. bovis de ganado. CONCLUSIONES: Se demuestra que M. bovis juega un papel importante en la epidemiología de la tuberculosis humana y representa un riesgo para la salud pública.

OBJECTIVE: The purpose of the study was to determine the role of bovine TB in cases of human TB. MATERIAL AND METHODS: Two-hundred and fifty-five samples from symptomatic patients were included in the study. All samples were cultured in Stonebrink and Lowënstein-Jensen media and analyzed using a nested PCRMPB70. The molecular analysis was performed by spoligotyping. RESULTS: From 255 samples, 74 were PCR-positive and 20 were culture-positive. From 94 samples positive to PCR or to isolation, 66 (70 percent) showed a spoligotype compatible with M. tuberculosis, and 13 (13.8 percent) with M. bovis. Four fingerprints of M. bovis from humans were identical to the fingerprints of M. bovis from cattle in the same region. CONCLUSIONS: Our study shows that M. bovis plays an important role in the epidemiology of TB in humans and that TB in cattle represents a risk to public health.

Molecular epidemiology of Mycobacterium bovis: usefulness in international trade.

Tuberculosis (TB) represents a barrier for free trade of livestock between Mexico and the United States of America (US). In spite of efforts from Mexico to export TB-free animals, some of those found with TB lesions in slaughterhouses in the US are traced back to that country. Therefore, the purpose of this study was to determine, through molecular epidemiology, the most probable source of infection for cattle found with TB lesions in the US. Ninety M. bovis isolates, 50 from Mexico obtained from cattle in 8 different states, and 40 from the US from cattle, deer, elk and feral pigs from 7 different states were included in the study. All samples were analyzed in both laboratories, Mexico and the US, following the same protocol for molecular analysis by spoligotyping. Twenty-seven clusters, ranging from 1 to 18 genetically similar strains were found. Some clustering by country was observed, strains from cattle and deer in Michigan in the US fell into the same cluster, suggesting transmission between species. These results, combined with epidemiological information suggest that despite of the possibility that some animals with lesions in the US come from Mexico as false negatives, the US has its own source of infection, must probably in dairy cattle and wildlife. Genetic diversity of isolates from Mexico was larger than that in the US, which could be a consequence of the endemic status of the disease and the indiscriminate movement of animals between regions.

Polymorphism of the PE domain of PE/PE_PGRS sequences in clinical isolates of Mycobacterium bovis in Mexico.

Polymorphism of the PE domain of PE/PE_PGRS sequences was studied in Mycobacterium bovis isolates from different Mexican states. Samples were analyzed by spolygotyping and RFLP using IS6110 and a 235-bp fragment of the PE domain of PE/PE_PGRS as probes. With the PE probe, three different genotypes were observed, one being predominant in all states. These results confirm the high conservation of the PE domain and suggests a potential role for PE sequence as a stable genetic marker for bovine tuberculosis.

Genotyping of Mycobacterium bovis by geographic location within Mexico.

The spacer oligonucleotide typing (spoligotyping) method was used to differentiate 62 Mycobacterium bovis isolates obtained from tissues with macroscopic lesions typical of tuberculosis in dairy cattle from different regions of Mexico. Our purpose was to see if a strain from one region was genetically different from those of other regions (with the long-term aim of doing molecular trace back of isolates obtained in the laboratory). Results from the genetic analysis indicate that M. bovis isolates cannot be grouped by geographic location due to a wide range of genetic types involved in dairy cattle infections. Isolates even from the same herd showed different spoligotypes but some isolates from different region had similar genetic patterns. Genetic typing without epidemiologic information does not seem to be a plausible method to trace back animals to source of origin to detect and eliminate sources of infection.

Diagnóstico de ileítis porcina por medio de la reacción en cadena de la polimerasa / Polymerase chain reaction for diagnosis of porcine ileitis

La ileítis porcina es una enfermedad que ocasiona importantes pérdidas económicas a la industria porcina. El agente causal de esta enfermedad es Lawsonia intracellularis, un microorganismo de dificl cultivo. Por esta razón, el diagnóstico, generalmente se realiza sólo al sacrificio. Debido a la dificultad del diagnóstico en animales vivos, se han desarrollado técnicas para detectar el ADN de la L. intracellularis por medio de la reacción en cadena de la polimerasa (PCR). El objetivo de este trabajo fue evaluar la técnica de PCR para el diagnóstico de la ileítis porcina en muestras de mucosa intestinal y heces de cerdos sospechosos de la enfermedad. El ADN fue extraído usando tierras diatomeas y tiocainato de guanidina. Para la amplificación del ADN se utilizaron oligonucleótidos específicos que amplifican un fragmento de ADN de L. intracellularis de 319 pb. La estandarización de la técnica se realizó a partir de mucosa intestinal de un cerdo infectado experimentalmente. La mínima cantidad de ADN de mucosa infectada que se detectó por PCR fue de 3.72 ng. Cuando la mucosa infectada fue adicionada a muestras de heces normales se necesitaron 12.4 ng del ADN extraído, para obtener un producto de amplificación visible. El producto de amplificación esperado de 319 pb fue también obtenido de mucosa intestinal o muestras de heces de cerdos infectados con signos clínicos característicos de ileítis porcina. Se concluyó que la técnica de PCR puede ser muy útil para el diagnóstico de esta enfermedad y para la determinación de la prevalencia de la ileítis porcina en diferentes áreas

Actualidades sobre la triquinelosis equina / Current facts about horse trichinellosis

La triquinelosis es una zoonosis causada por parásitos del género Trichinella que es trasmitida principalmente por la ingestión de carne de animales como el cerdo, oso y zorro entre otros. Sin embargo, en Europa se han reportado varios brotes debido al comsumo de carne de caballo. La presencia del parásito no ha sido demostrada de manera directa en la carne de estos animales, sin embargo, la identificación de las especies de Trichinella (T. spiralis T. britovi y T. nativa) involucradas en estos brotes ha sido posible a partir de biopsias tomadas de individuos que consumieron carne de caballo. Recientemente, se identificaron por primera vez larvas de T. spiralis en caballos sacrificados en un rastro del Estado de México, presentando así evidencia directa de la infección de estos animales con el parásito. Por otro lado empleando extractos totales o antígenos TSL-1 de T. spiralis se han detectado anticuerpos en contra de Trichinella en caballos sacrificados en rastros de diferentes paises de Europa así como en México. Asimismo, la infección con varias especies de Trichinella se ha logrado reproducir experimentalmente en caballos y los resultados obtenidos son importantes en el desarrollo de métodos de diagnóstico que permitan estimas la prevalencia de esta infección en caballos cuya carne se destina para el consumo tanto animal como humano y eventualmente instrumentar medias para el control de la trasmisión de la triquinelosis por carne de caballos

Alteraciones de la concentración de leucocitos y subpoblaciones de linfocitos de cerdos que sobrevivieron al desafío con el virus de la fiebre porcina clásica / Changes in leucocyte and lymphocyte subpopulations of pigs that survived classical swine fever virus challenge

La infección con el virus de la fiebre porcina clásica (FPC) provoca la disminución de todas las subpoblaciones de leucocitos circulantes y una inmunosupresión marcada en los cerdos que se enferman y mueren. Sin embargo, no se ha informado del efecto del virus sobre los leucocitos circulantes en cerdos que sobreviven a la infección. Con objeto de conocer este efecto se estudiaron 6 cerdos que, después de ser desafiados experimentalmente con el virus de la FPC, desarrollaron la enfermedad, pero posteriormente se recuperaron. Los parámetros evaluados en estos cerdos fueron los signos clínicos, la temperatura rectal, la biometría hemática y las subpoblaciones del linfocitos determinadas por rosetas de eritrocitos. Los resultados mostraron que hubo un incremento en la temperatura rectal desde el día siete haste el día doce posinoculación, asociada a anorexia, decaimiento y diarrea ligera. En los primeros siete días hubo una disminución marcada de la concentración de leucocitos totales, linfocitos, leucocitos polimorfonucleares segmentados y en banda, monocitos, linfocitos T totales (TE), T de alta afinidad (Taa). B con receptor fc (Bfc) y Null. Al mismo tiempo se observó un incremento de T autólogos (Taut), que son células inmaduras, y no hubo alteración de los linfocitos B con receptor de complemento (Bc). Sin embargo, a partir del séptimo día se incrementaron los valores de la mayoría de las células y disminuyeron los Taut. Estos resultados indican que la recuperación de los animales estuvo asociada al incremento de los leucocitos circulantes