Salmonellosis in calves without intestinal lesions / Salmonelose em bezerros sem manifestações intestinais

Salmonellosis is a known cause of enteric disorders in calves. However, cases in the septicemic form may not present enteric lesions, which may lead the veterinary practitioner to not suspect salmonellosis, compromising the diagnosis. The current study describes the epidemiological, clinical, pathological and immunohistochemical aspects of septicemic salmonellosis in calves without enteric lesions. The protocols involving bovine material submitted to the Pathology Laboratory (LAP) of the "Faculdade de Medicina Veterinária e Zootecnia" (FAMEZ) of the "Universidade Federal do Mato Grosso do Sul" (UFMS) from January 1995 to July 2018 were studied. Cases confirmed or suggestive of septicemic salmonellosis in calves without enteric manifestations were selected. Fragments of the liver, lung, and spleen embedded in paraffin were submitted to immunohistochemistry (IHC). Only cases in which there was positive marking on the IHC or culture isolation of Salmonella were included in this study. Of a total of 5,550 cattle examined in the period, ten presented septicemic salmonellosis without enteric lesions. Clinical signs included mucosal pallor, apathy, hyperthermia, and dyspnea. Only three calves presented diarrhea, and two were found dead before clinical changes were observed. The most common necropsy findings were hepatosplenomegaly; yellow, orange or brown discolored livers; pale mucous membranes; inflated and sometimes red lungs; fibrin or fluid within body cavities; and gallbladder filled with inspissated bile. Jaundice was observed in three calves that had a concomitant infection with Anaplasma sp. Microscopically, paratyphoid hepatic nodules and interstitial pneumonia were the most frequent manifestations, followed by thrombosis and bacterial colonies in the spleen, lung, liver, and brain. A strong positive marking was observed in IHC, predominantly in the lung and to a lesser extent in the liver. Polymerase chain reaction (PCR) indicated the Dublin serotype as the causative agent in the samples of the four calves submitted to this procedure. In calves, the septicemic form was the major cause of death due to salmonellosis. Septicemic salmonellosis was usually not accompanied by diarrhea. The clinical signs of septicemia are nonspecific and of little assistance in the diagnosis. IHC has been shown to be efficient in the detection of the agent, mainly in the lung and especially in situations where it is not possible to perform bacterial culture.(AU)

A salmonelose é uma causa conhecida de distúrbios entéricos em bezerros. Porém, casos na forma septicêmica podem não apresentar manifestação entérica, o que leva o médico veterinário a não suspeitar de salmonelose, comprometendo o diagnóstico. Este estudo descreve os aspectos epidemiológicos, clínicos, patológicos e imuno-histoquímicos da salmonelose septicêmica em bezerros sem lesões entéricas. O estudo foi realizado a partir dos protocolos referentes a materiais de bovinos enviados para diagnóstico ao Laboratório de Anatomia Patológica (LAP) da Faculdade de Medicina Veterinária e Zootecnia (FAMEZ) da Universidade Federal de Mato Grosso do Sul (UFMS) de janeiro de 1995 a julho de 2018. Foram selecionados os casos de bezerros confirmados ou sugestivos de salmonelose septicêmica sem lesões entéricas. Fragmentos de fígado, pulmão e baço embebidos em parafina foram submetidos ao exame de imuno-histoquímica (IHQ). Somente foram incluídos neste estudo casos em que houve marcação positiva na IHQ ou isolamento da bactéria em cultura. De um total de 5.550 bovinos examinados no período, dez apresentaram salmonelose septicêmica sem lesão entérica. Os sinais clínicos incluíram palidez de mucosas, apatia, hipertermia e dispneia. Apenas três bezerros apresentaram diarreia e dois foram encontrados mortos sem terem sido observadas alterações clínicas. Os achados mais frequentes de necropsia foram hepatoesplenomegalia, fígado amarelado, alaranjado ou acastanhado, palidez de mucosas, pulmões inflados e, por vezes, vermelhos, fibrina ou líquido nas cavidades do organismo e vesícula biliar repleta de bile grumosa. Icterícia foi observada em três bezerros que apresentavam infecção concomitante por Anaplasma sp. Microscopicamente, os nódulos paratifoides hepáticos e pneumonia intersticial foram as manifestações mais encontradas, seguidas por trombose e colônias bacterianas no baço, pulmão, fígado e encéfalo. Na IHQ, marcação fortemente positiva foi observada, predominantemente, no pulmão e, em menor intensidade, no fígado. A técnica de reação em cadeia de polimerase (PCR) tipificou o sorotipo Dublin como agente etiológico nas amostras dos quatro bezerros submetidos a este procedimento. Em bezerros, a forma septicêmica foi a principal responsável pelas mortes por salmonelose. Na maioria das vezes essa forma não estava acompanhada por diarreia. Os sinais clínicos da forma septicêmica são inespecíficos e de pouco auxílio no direcionamento do diagnóstico. A IHQ mostrou-se eficiente na detecção do agente principalmente no pulmão e especialmente nas situações em que não é possível a realização da cultura bacteriana.(AU)

Pathogen translocation and histopathological lesions in an experimental model of Salmonella Dublin infection in calves receiving lactic acid bacteria and lactose supplements

The purpose of this study was to evaluate the capacity of a lactic acid bacteria (LAB) inoculum to protect calves with or without lactose supplements against Salmonella Dublin infection by evaluating histopathological lesions and pathogen translocation. Fifteen calves were divided into three groups [control group (C-G), a group inoculated with LAB (LAB-G), and a group inoculated with LAB and given lactose supplements (L-LAB-G)] with five, six, and four animals, respectively. The inoculum, composed of Lactobacillus (L.) casei DSPV 318T, L. salivarius DSPV 315T, and Pediococcus acidilactici DSPV 006T, was administered with milk replacer. The LAB-G and L-LAB-G received a daily dose of 109 CFU/kg body weight of each strain throughout the experiment. Lactose was provided to the L-LAB-G in doses of 100 g/day. Salmonella Dublin (2 x 1010 CFU) was orally administered to all animals on day 11 of the experiment. The microscopic lesion index values in target organs were 83%, 70%, and 64.3% (p < 0.05) for the C-G, LAB-G, and L-LAB-G, respectively. Administration of the probiotic inoculum was not fully effective against infection caused by Salmonella. Although probiotic treatment was unable to delay the arrival of pathogen to target organs, it was evident that the inoculum altered the response of animals against pathogen infection.

Serum protein concentrations, including acute phase proteins, in calves experimentally infected with Salmonella Dublin / Teores séricos de proteínas, incluindo proteínas de fase aguda, em bezerros infectados experimentalmente com Salmonella Dublin

The aim of this study was to evaluate serum protein concentrations in calves experimentally inoculated with Salmonella Dublin. Twelve healthy 10 to 15-day-old Holstein calves were randomly allotted into two groups, control and infected with 10(8) CFU of Salmonella Dublin orally. The calves were subjected to physical evaluation and blood samples were collected shortly before administration of the bacteria and also 24, 48, 72, 96, 120 and 168 hours post-infection. The concentration of serum proteins was determined through sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE). Thirty serum proteins ranging from molecular weight of 24,000 Da to molecular weight of 236,000 Da were detected. Serum concentrations of ceruloplasmin (125,000 Da), haptoglobin (45,000 Da), acid glycoprotein (40,000 Da) and a 34,000 Da protein were significantly increased in the experimentally infected calves, when compared with their concentrations in the control animals. Therefore, this study showed that S. Dublin infection could lead to the increase of certain serum proteins in calves.

O objetivo do estudo foi avaliar as concentrações séricas de proteínas em bezerros inoculados experimentalmente com Salmonella Dublin. Foram examinados 12 bezerros sadios da raça Holandesa com 10 a 15 dias de idade, distribuídos aleatoriamente em dois grupos, controle e infectado experimentalmente com 10(8) UFC de Salmonella Dublin. As amostras de sangue foram colhidas minutos antes e 12, 24, 48, 72, 96, 120 e 168 horas após a inoculação. As concentrações séricas das proteínas ceruloplasmina (125.000 Da), haptoglobina (45.000 Da), glicoproteína ácida (40.000 Da) e proteína de peso molecular de 34.000 Da foram significativamente maiores nos bezerros infectados do que nos do grupo controle. Portanto, o estudo mostrou que a infecção por S. Dublin pode elevar a concentração de certas proteínas séricas em bezerros.

Efficacy of florfenicol and intravenous fluid therapy for treatment of experimental salmonellosis in newborn calves / Eficácia do florfenicol e da fluidoterapia parenteral no tratamento da salmonelose experimental em bezerros neonatos

The efficacy of florfenicol associated or not to intravenous fluid therapy for treatment of Salmonella Dublin-infected calves was determined. Twenty-four healthy 10 to 15-day-old Holstein calves were randomly allotted into four groups, with six animals each: control (group 1); infected with 10(8)CFU Salmonella Dublin and not treated (group 2); infected with 10(8)CFU Salmonella Dublin and treated with florfenicol (group 3); and infected with 10(8)CFU Salmonella Dublin and treated with florfenicol associated to fluid therapy (group 4). All animals were submitted to physical examination just before inoculation and every 24 hours, during seven days after experimental infection. Rectal swabs and blood samples were collected for Salmonella Dublin isolation and pH and blood electrolytes determination. The experimental infection with Salmonella Dublin induced clinical signs of salmonellosis, such as diarrhea and fever, and caused reduction in blood concentrations of pH, sodium, potassium and chlorides. The treated calves showed good clinical recovery, and the group treated with antibiotic in combination to fluid therapy presented a faster and more efficient correction of the hydro-electrolyte balance.

Avaliou-se a eficácia terapêutica do florfenicol associado ou não à fluidoterapia intravenosa no tratamento de bezerros infectados experimentalmente com Salmonella Dublin. Foram utilizados 24 bezerros sadios da raça Holandesa com 10 a 15 dias de idade, distribuídos aleatoriamente em quatro grupos experimentais, constituídos por seis animais cada: controle (grupo 1); infectado com 10(8)UFC de Salmonella Dublin e não tratado (grupo 2); infectado com 10(8)UFC de Salmonella Dublin e tratado com florfenicol (grupo 3); e infectado com 10(8)UFC de Salmonella Dublin (grupo 4) e tratado com florfenicol associado à fluidoterapia. Todos os animais foram submetidos ao exame físico logo antes da inoculação e a cada 24 horas, durante sete dias após a infecção experimental. Foram colhidas amostras de suabes retais para o isolamento de Salmonella Dublin e amostras de sangue para determinação dos valores de pH e dosagem de eletrólitos sanguíneos. A infecção experimental com Salmonella Dublin induziu sinais clínicos de salmonelose, como diarreia e febre, e provocou redução do valor do pH e das concentrações sanguíneas de sódio, potássio e cloreto. Os bezerros submetidos aos tratamentos mostraram boa recuperação clínica, sendo que o grupo tratado com antibiótico combinado à fluidoterapia apresentou correção mais rápida e eficiente do equilíbrio hidroeletrolítico.

Avaliação da reação em cadeia da polimerase e do isolamento bacteriológico convencional na detecção de Salmonella Dublin em amostras de fezes de bezerros infectados experimentalmente / Evaluation of polymerase chain reaction and standard microbiological techniques for detection of Salmonella Dublin in fecal samples of infected calves

The efficiency of the Polymerase Chain Reaction (PCR) combined with selective enrichment broth was compared with the standard microbiological techniques for detection of Salmonella Dublin in fecal samples of 10 to 15-days-old Holstein calves, experimentally infected with 10(8) CFU of Salmonella Dublin. Seventy-six fecal samples were analyzed using PCR associated with selenite cystine (SC) and Muller-Kauffmann tetrathionate (TMK) broths and standard microbiological techniques. Regardless of the selective enrichment broth used, the standard microbiological techniques were significantly better than PCR in detection of positive samples of Salmonella Dublin. However, the simultaneous use of both techniques provided detection of a larger number of positive samples. The SC broth was the best option as selective enrichment in both techniques.

Hemograma e perfil bioquímico sérico, inclusive hemogasométrico, de bezerros infectados experimentalmente com Salmonella Dublin / Blood cell counts and serum biochemical profile, including blood gas levels, in Salmonella Dublin-infected calves

O objetivo do estudo foi avaliar o hemograma e o perfil bioquímico sérico, inclusive hemogasométrico, de bezerros infectados experimentalmente com Salmonella Dublin. Foram utilizados 12 bezerros sadios da raça Holandesa com 10 a 15 dias de idade, distribuídos aleatoriamente em dois grupos experimentais: grupo-controle (n= 6) e grupo infectado com 10(8)UFC de Salmonella Dublin (n=6). Os bezerros foram submetidos ao exame físico diário, e as amostras de sangue foram coletadas minutos antes da inoculação (0h) e 24, 48, 72, 96, 120, 144 e 168h após a inoculação. Além do hemograma e das análises hemogasométricas, foram mensuradas as atividades séricas das enzimas aspartato aminotransferase (AST), fosfatase alcalina (ALP), creatina cinase (CK), gamaglutamiltransferase (GGT) e lactato desidrogenase (LDH), e os teores de albumina, bilirrubinas, cálcio total, cálcio ionizado, sódio, potássio, cloretos, creatinina, ferro, fibrinogênio, fósforo, glicose, magnésio, proteína totais e ureia. As principais alterações foram: redução das concentrações de albumina, ferro, glicose, magnésio e proteína total, aumento do teor de fibrinogênio, leucocitose e acidose metabólica e hiponatremia.

The blood cell counts and biochemical profile, including blood gas levels, were evaluated, in Salmonella Dublin-infected calves. Twelve healthy 10 to 15-day old Holstein calves were randomly allotted into two groups: control (n=6) and group orally infected with 10(8) CFU Salmonella Dublin (n=6). The calves were submitted to physical examination and the blood samples were taken just before the inoculation (0h) and at 24, 48, 72, 96, 120, 144, and 168h later. Besides, blood cell counts, blood gas levels, and the serum concentrations of aspartate aminotransferase (AST), alkaline phosphatase (ALP), creatine kinase (CK), gammaglutamyltransferase (GGT), lactate desidrogenase (LDH), albumin, bilirubin, total calcium, ionic calcium, sodium, potassium, chlorides, creatinine, iron, fibrinogen, phosphorus, glucose, magnesium, total protein, and urea were measured. The main findings included decrease in albumin, iron, glucose, magnesium, and total protein concentrations, and increase in fibrinogen level, leukocytosis, metabolic acidosis, and hyponatremia.

Comparação da eficiência dos caldos de enriquecimento seletivo no isolamento de Salmonella Dublin / Comparison of the efficiency of selective enrichment broths for Salmonella Dublin isolation

The aim of this study was to compare three different selective enrichment broths: Rappaport-Vassiliadis (RV), selenite cystine (SC) and Muller-Kauffmann tetrathionate (MKT) for Salmonella Dublin isolation from faecal samples of calf experimentally infected. The bacteriological procedure involved pre-enrichment stages in Hajna-GN broth (only for the samples inoculated in RV broth), selective enrichment, culture in modified brilliant green agar (BGA), presumptive biochemistry tests (using triple-sugar-iron agar and lysine-agar) and slide agglutination test with poli-O and poli-H Salmonella antiserum. The effects of enrichment temperatures using RV broth were also evaluated (37ºC and 42ºC). SC broth was significantly more efficient in the isolation of Salmonella Dublin (P<0,05), whereas RV broth incubated at 42ºC had a lower efficiency in the microbiological isolation.

Avaliação clínica da infecção experimental de bezerros com Salmonella Dublin / Clinical evaluation of experimental Salmonella Dublin infection in calves

The clinical conditions of healthy calves infected with experimental 10(8)CFU of Salmonella Dublin were evaluated and the viability of the experimental model in disease induction in calves was verified. Twelve 10 to 15-day-old male Holstein calves were examined. They were allocated into two groups, control and experimentally infected with 10(8)CFU of Salmonella Dublin. Animals were submitted to clinical examination after inoculation and at every 12 hours, during seven days after the experimental infection. Samples of rectal swabs were collected for Salmonella Dublin isolation. All animals had severe diarrhea, with mucus and bleeding, 12 to 84 hours after the experimental infection with Salmonella Dublin, accompanied by fever, dehydration and respiratory signs. The isolation of Salmonella Dublin from rectal swabs occurred 12 hours after the infection. Two out of the six animals inoculated with Salmonella Dublin died with symptoms of enteritis, fibrinous pneumonia, centrilobular hepatic steatosis, hepatocyte necrosis, spleen congestion, interstitial nephritis, and tubular degeneration. Thus, the oral administration of 10(8)CFU of Salmonella Dublin induced clinical signs of salmonellosis in 10 to 15-day-old calves.