Mycoplasma hyopneumoniae infection dynamics in naïve replacement gilts introduced to positive farms.

This study was designed to characterize the dynamics of infection of Mycoplasma hyopneumoniae in naïve replacement gilts after introduction to positive systems. Ninety-eight naïve gilts were monitored in three positive commercial farms (A, B, and C). The näive gilts were housed for 21 days in pens adjacently located to older gilt cohorts (named seeders), which have been naturally exposed to the positive farms. The infection dynamics was evaluated by PCR and ELISA, from laryngeal swabs and serum samples, respectively. Samples were collected at 150 (arrival), 165, 180, 210, 240, 270, 300 days of age (doa), and pre-farrowing. Infection occurred rapidly on farms A and B, taking 25.2 and 23.9 days for 95% of gilts to be PCR positive, respectively. There was no influence on the number of seeders at the time of exposure, but their absence (farm C) could explain the extended period it took for gilts to get infected (69.4 days). On average, it took 162.2 days after the first PCR detection for 85% of gilts to stop shedding the bacterium. The serology results were consistent with the herd infection curve. At pre-farrowing, 100% of gilts seroconverted and 36.7% remained PCR positive. A total of 1.33% of piglets were positive at weaning. Fifteen variants were detected among the three farms by MLVA. The acclimation protocol was efficient and easy to perform, and the presence of seeders was likely critical for early acclimation for M. hyopneumoniae.

The effect of gilt flow management during acclimation on Mycoplasma hyopneumoniae detection.

The objective of this study was to characterize the Mycoplasma hyopneumoniae (M. hyopneumoniae) detection and seroconversion patterns in recently acclimated gilts to be introduced to endemically infected farms using different types of replacement management. Three gilt developing units (GDUs) belonging to sow farms were included in this investigation: two farms managed gilts in continuous flow, and one farm managed gilts all-in/all-out. Two replicates of 35 gilts each were selected per GDU and sampled approximately every 60 days for a total of four or five samplings, per replicate and per GDU. Detection of M. hyopneumoniae was evaluated by PCR, while antibodies were measured using a commercial ELISA assay. Also, M. hyopneumoniae genetic variability was evaluated using Multiple-Locus Variable number tandem repeat Analysis. Detection of M. hyopneumoniae was similar across GDUs. Although a significant proportion of gilts was detected positive for M. hyopneumoniae after acclimation, an average of 30.3 % of gilts was negative at any point during the study. Detection of M. hyopneumoniae antibodies was similar among GDUs regardless of flow type or vaccination protocol. The genetic variability analysis revealed a limited number of M. hyopneumoniae types within each GDU. Results of this study showed a similar pattern of M. hyopneumoniae detection by PCR and seroconversion by ELISA among GDUs, regardless of the type of flow management strategies applied to gilts.

Agents of pneumonia in slaughtered pigs in southern Brazil

ABSTRACT: A high prevalence of pneumonic lesions has been reported to affect slaughtered pigs in southern Brazil. In order to identify which microorganisms have been causing those lesions, 30 pig lungs presenting pneumonic gross lesions were collected from five different slaughterhouses, totaling 150 lungs. Samples for bacterial isolation, molecular, histopathologic and immunohistochemistry (IHC) evaluation were taken from each lung. The pneumonic lesion scoring ranged from 1.53 to 2.83. The most frequent histopathological lesions found was the concomitant Influenza A virus (IAV) and Mycoplasma hyopneumoniae infection, corresponding to 55.3% (83/150), and Pasteurella multocida type A was isolated in 54.2% (45/83) of these cases. In 102 samples (68%), there was histopathologic suggestion of involvement of more than one infectious agent. M. hyopneumoniae was the most frequent agent associated with pneumonic lesions, being present in 92.1% (94/102) of the lungs with coinfections, followed by IAV in 89.2% (91/102). Besides the coinfections, IAV lesions were observed also in six samples without another pathogenic microorganism detected. A total of 46 samples with acute and subacute IAV suspected lesions in histopathological examination were assessed for IHC and real time RT-PCR for IAV. A total of 35% (16/46) of them were positive by IHC and 13% (6/46) by real time RT-PCR. Regarding M. hyopneumoniae, 79.3% (119/150) of samples were positive by qPCR and 84.9% (101/119) of them also presented M. hyopneumoniae suspected lesions in the histopathological examination. The results of this study suggest the importance of IAV in respiratory diseases in finishing pigs, even though this virus is more frequently reported in the nursery phase. In addition, our results emphasize the importance of lung coinfections in finishing pigs.

RESUMO: Lesões sugestivas de pneumonia são frequentemente encontradas em altas prevalências em suínos abatidos no sul do Brasil. Para identificar quais microrganismos causam essas lesões, foram coletados 30 pulmões de suínos com lesão macroscópica sugestiva de pneumonia em cinco frigoríficos diferentes, totalizando 150 pulmões. Amostras para isolamento bacteriano, avaliação molecular, histopatológica e imuno-histoquímica (IHC) foram coletadas de cada pulmão. O escore de lesão pulmonar variou entre 1,53 a 2,83. O achado histopatológico mais observado foi a lesão sugestiva de infecção concomitante pelo vírus Influenza A (IAV) e Mycoplasma (M.) hyopneumoniae, correspondendo a 55,3% (83/150), e em 54,2% (45/83) desses casos Pasteurella (P.) multocida tipo A foi isolado. Em 102 amostras (68%), houve lesão histopatológica sugestiva do envolvimento de mais de um agente infeccioso. M. hyopneumoniae foi o microrganismo mais frequente associado a lesões de pneumonia, estando presente em 92,1% (94/102) dos pulmões com coinfecções, seguido de IAV, que foi encontrado em 89,2% (91/102). Além das coinfecções, lesões de IAV foram observadas em mais seis amostras que não aparentavam envolvimento de outro agente infeccioso. Um total de 46 amostras com suspeita de lesão aguda e subaguda de IAV no exame histopatológico foram avaliadas para IHC e RT-PCR em tempo real para IAV e 35% (16/46) delas foram positivas por IHC e 13% (6/46) foram positivas por RT-PCR em tempo real. Com relação a M. hyopneumoniae, 79,3% (119/150) das amostras foram positivas por qPCR e 84,9% (101/119) delas também apresentaram lesões suspeitas de M. hyopneumoniae no exame histopatológico. Os resultados deste trabalho sugerem a importância do IAV como agente causador de pneumonias em suínos de terminação, embora esse vírus seja mais frequentemente relatado na fase de creche. Além disso, os achados deste trabalho demonstram a presença frequente de coinfecções pulmonares em suínos de terminação.

Agents of pneumonia in slaughtered pigs in southern Brazil / Agentes patogênicos de pneumonia em suínos abatidos no sul do Brasil

A high prevalence of pneumonic lesions has been reported to affect slaughtered pigs in southern Brazil. In order to identify which microorganisms have been causing those lesions, 30 pig lungs presenting pneumonic gross lesions were collected from five different slaughterhouses, totaling 150 lungs. Samples for bacterial isolation, molecular, histopathologic and immunohistochemistry (IHC) evaluation were taken from each lung. The pneumonic lesion scoring ranged from 1.53 to 2.83. The most frequent histopathological lesions found was the concomitant Influenza A virus (IAV) and Mycoplasma hyopneumoniae infection, corresponding to 55.3% (83/150), and Pasteurella multocida type A was isolated in 54.2% (45/83) of these cases. In 102 samples (68%), there was histopathologic suggestion of involvement of more than one infectious agent. M. hyopneumoniae was the most frequent agent associated with pneumonic lesions, being present in 92.1% (94/102) of the lungs with coinfections, followed by IAV in 89.2% (91/102). Besides the coinfections, IAV lesions were observed also in six samples without another pathogenic microorganism detected. A total of 46 samples with acute and subacute IAV suspected lesions in histopathological examination were assessed for IHC and real time RT-PCR for IAV. A total of 35% (16/46) of them were positive by IHC and 13% (6/46) by real time RT-PCR. Regarding M. hyopneumoniae, 79.3% (119/150) of samples were positive by qPCR and 84.9% (101/119) of them also presented M. hyopneumoniae suspected lesions in the histopathological examination. The results of this study suggest the importance of IAV in respiratory diseases in finishing pigs, even though this virus is more frequently reported in the nursery phase. In addition, our results emphasize the importance of lung coinfections in finishing pigs.(AU)

Lesões sugestivas de pneumonia são frequentemente encontradas em altas prevalências em suínos abatidos no sul do Brasil. Para identificar quais microrganismos causam essas lesões, foram coletados 30 pulmões de suínos com lesão macroscópica sugestiva de pneumonia em cinco frigoríficos diferentes, totalizando 150 pulmões. Amostras para isolamento bacteriano, avaliação molecular, histopatológica e imuno-histoquímica (IHC) foram coletadas de cada pulmão. O escore de lesão pulmonar variou entre 1,53 a 2,83. O achado histopatológico mais observado foi a lesão sugestiva de infecção concomitante pelo vírus Influenza A (IAV) e Mycoplasma (M.) hyopneumoniae, correspondendo a 55,3% (83/150), e em 54,2% (45/83) desses casos Pasteurella (P.) multocida tipo A foi isolado. Em 102 amostras (68%), houve lesão histopatológica sugestiva do envolvimento de mais de um agente infeccioso. M. hyopneumoniae foi o microrganismo mais frequente associado a lesões de pneumonia, estando presente em 92,1% (94/102) dos pulmões com coinfecções, seguido de IAV, que foi encontrado em 89,2% (91/102). Além das coinfecções, lesões de IAV foram observadas em mais seis amostras que não aparentavam envolvimento de outro agente infeccioso. Um total de 46 amostras com suspeita de lesão aguda e subaguda de IAV no exame histopatológico foram avaliadas para IHC e RT-PCR em tempo real para IAV e 35% (16/46) delas foram positivas por IHC e 13% (6/46) foram positivas por RT-PCR em tempo real. Com relação a M. hyopneumoniae, 79,3% (119/150) das amostras foram positivas por qPCR e 84,9% (101/119) delas também apresentaram lesões suspeitas de M. hyopneumoniae no exame histopatológico. Os resultados deste trabalho sugerem a importância do IAV como agente causador de pneumonias em suínos de terminação, embora esse vírus seja mais frequentemente relatado na fase de creche. Além disso, os achados deste trabalho demonstram a presença frequente de coinfecções pulmonares em suínos de terminação.(AU)

Infection dynamics and genetic variability of Mycoplasma hyopneumoniae in self-replacement gilts.

The aim of this study was to assess the longitudinal pattern of M. hyopneumoniae detection in self-replacement gilts at various farms and to characterize the genetic diversity among samples. A total of 298 gilts from three M. hyopneumoniae positive farms were selected at 150days of age (doa). Gilts were tested for M. hyopneumoniae antibodies by ELISA, once in serum at 150 doa and for M. hyopneumoniae detection in laryngeal swabs by real time PCR two or three times. Also, 425 piglets were tested for M. hyopneumoniae detection in laryngeal swabs. A total of 103 samples were characterized by Multiple Locus Variable-number tandem repeats Analysis. Multiple comparison tests were performed and adjusted using Bonferroni correction to compare prevalences of positive gilts by ELISA and real time PCR. Moderate to high prevalence of M. hyopneumoniae in gilts was detected at 150 doa, which decreased over time, and different detection patterns were observed among farms. Dam-to-piglet transmission of M. hyopneumoniae was not detected. The characterization of M. hyopneumoniae showed 17 different variants in all farms, with two identical variants detected in two of the farms. ELISA testing showed high prevalence of seropositive gilts at 150 doa in all farms. Results of this study showed that circulation of M. hyopneumoniae in self-replacement gilts varied among farms, even under similar production and management conditions. In addition, the molecular variability of M. hyopneumoniae detected within farms suggests that in cases of minimal replacement gilt introduction bacterial diversity maybe farm specific.

Mycoplasma hyopneumoniae detection in nylon-flocked and rayon-bud swabs.

The objective of this study was to compare the absorption and M. hyopneumoniae detection by real-time PCR using nylon-flocked and rayon-bud swabs. Results of this study showed that absorption and detection of M. hyopneumoniae were significantly higher in nylon-flocked swabs with Ct differences only ranging from 0.5 to 1.7.

Detection of Mycoplasma hyopneumoniae in naturally infected gilts over time.

Mycoplasma hyopneumoniae causes a chronic respiratory infection in pigs and its transmission occurs mainly by direct contact and by vertical transmission (sow-to-piglet). The objective of this study was to assess the detection dynamics and persistence of M. hyopneumoniae natural infection in replacement gilts. Forty-four twenty-day-old gilts were selected from a M. hyopneumoniae positive farm and followed up to one day prior to their first weaning. Laryngeal swabs were collected every 30days, starting at day 20, for M. hyopneumoniae detection by real-time PCR, resulting in 12 samplings. Piglets born to selected females were sampled via laryngeal swabs one day prior to weaning to evaluate sow-to-piglet transmission. The M. hyopneumoniae prevalence was estimated at each one of the 12 samplings in gilts and a multiple comparison test and Bonferroni correction were performed. Bacterial detection in gilts started at 110days of age (doa) and a significant increase (p<0.05) occurred at 140 doa. The M. hyopneumoniae prevalence remained above 20% from 140 to 230 doa, decreasing thereafter. However, it did not reach 0% at any sampling after 110 doa. In this study, M. hyopneumoniae was not detected in piglets sampled prior to weaning. The M. hyopneumoniae detection pattern showed that in natural infections, gilts were positive for M. hyopneumoniae for one to three months, but occasionally long-term detection may occur. Moreover, the lack of M. hyopneumoniae detection throughout the study in 18.2% of gilts indicated the existence of negative subpopulations in positive herds.

Virulence profiles of enterotoxigenic Escherichia coli isolated from piglets with post-weaning diarrhea and classification according to fecal consistency / Perfis de virulência de Escherichia coli enterotoxigênica isoladas de leitões desmamados com diarreia do Sul do Brasil e classificação das amostras de acordo com a consistência fecal

The aim of this study was to assess the frequency and association of virulence factors of Escherichia (E.) coli isolated from weaned piglets with diarrhea and to correlate it with fecal consistency. A total of 152 rectal swabs were collected from 25-40 day-old piglets with diarrhea, in farms of Southern Brazil. Phenotypical and molecular techniques were used for bacterial isolation, characterization and classification of enterotoxigenic E. coli (ETEC) pathotypes. Statistical analysis was carried out to determine the frequency of virulence factors and virotypes, of fimbriae F4, F5, F6, F18, F41 and toxins LT, STa, STb and STx2e. Out of 456 E. coli isolates, 287 (62.9%) samples showed significant growth of E. coli. Among them, 194 (67.6%) samples showed at least one virulence factor, indicating that ETEC is an important etiological agent of diarrhea in weaned piglets. Higher frequencies were found of fimbria F4 and F18 and enterotoxins LT, STa and STb. Significant association was found to F4, LT, STa and STb; between F18 and STa and STx2e; between F5 and LT, STa and STb. The most frequent virotypes were F18-STa, F4-LT-STa-STb, F4-STa, F4-LT-STb and F18-STa-STx2e. Beta-hemolysis was observed in 47.4% of samples and there was significant association between hemolytic samples and virulence factors F4, F18, STa and STx2e. Regarding fecal consistency, there was significant association of liquid feces and F4 fimbria, STa toxin and virotypes F4-STa and F4-F5-LT-STa-STb. Since there was significant association of ETEC and liquid feces in nursery piglets, it is important to prioritize the sampling of liquid feces for the diagnosis etiologic cause of diarrhea.

O objetivo deste estudo foi avaliar a frequência e associação de fatores de virulência de Escherichia (E.) coli isoladas de leitões desmamados com diarreia e correlacioná-la com consistência fecal. Suabes retais foram coletados em leitões com 25-40 dias de idade com sinal clínico de diarreia, em granjas do Sul do Brasil, totalizando 456 amostras. Foram utilizadas técnicas fenotípicas e moleculares para isolamento bacteriano, caracterização e classificação de patotipos de E. coli enterotoxigênica (ETEC). A análise estatística foi realizada para determinar a frequência de fatores de virulência e virotipos, de fímbrias F4, F5, F6, F18, F41 e toxinas LT, STa, STB e STx2e. Duzentas e oitenta e sete (62,9%) amostras apresentaram crescimento significativo de E. coli. Entre os quais, 194 (67,6%) amostras apresentaram pelo menos um fator de virulência, indicando que ETEC é um importante agente etiológico de diarreia em leitões desmamados. As frequências mais elevadas foram encontradas para as fímbrias F4 e F18 e enterotoxinas LT, STa e STb. Associação significativa foi encontrada para F4, LT, STa e STb; entre F18 e STa e STx2e; entre F5 e LT, STa e STb. Os virotipos mais frequentes foram F18-STa, F4-LT-STa-STb, F4-STa, F4-LT-STb e F18-STa-STx2e. Beta-hemólise foi observada em 47,4% das amostras e houve associação significativa entre amostras hemolíticas e fatores de virulência F4, F18, STa e STx2e. Quanto consistência fecal, houve associação significativa de fezes líquidas e fímbria F4, toxina STa e virotipos F4-STa e F4-F5-LT-STa-STb. A associação significativa da ETEC e fezes líquidas em leitões de creche, é importante para priorizar a amostragem de fezes com essa consistência para no diagnóstico etiológico da diarreia.