Detection of Mycobacterium avium subsp. paratuberculosis in bovine milk from the state of Pernambuco, Brazil

Abstract

The aim of this study was to detect the IS900 region of Mycobacterium avium subsp. paratuberculosis (MAP) in bovine milk samples using real-time polymerase chain reaction (qPCR) and conventional PCR, and to study the agreement between these tests. A total of 121 bovine milk samples were collected from herds considered positive for MAP, from the State of Pernambuco, Brazil. MAP DNA was detected in 20 samples (16.5%) using conventional PCR and in 34 samples (28.1%) using qPCR. MAP DNA was detected in all of the 6 animal farms studied. Moderate agreement was found between qPCR and conventional PCR results, where the sensitivity and specificity of conventional PCR in relation to qPCR were 50% and 96.6%, respectively. Thus, the IS900 region of MAP was found in bovine milk samples from the State of Pernambuco. To the best of our knowledge, this is the first report of MAP DNA found in bovine milk in Northeast Brazil. We also demonstrated the qPCR technique is more sensitive than conventional PCR with respect to detection of MAP in milk samples.

Occurrence of Mycobacterium avium subsp. paratuberculosis in coalho cheese in the State of Pernambuco, Brazil

“…In our study, a higher occurrence of MAP DNA was observed in coalho type cheese samples, perhaps due to the influence of the area where these samples were collected. In the state of Pernambuco, paratuberculosis has been reported with a certain frequency in cattle (Mota et al, 2007;Sá et al, 2013), and its DNA was detected in milk samples in the Garanhuns microregion (Albuquerque et al, 2017). It is also important to highlight that the qPCR technique used in our study has a greater sensitivity and specificity in the detection of this microorganism when compared to conventional PCR, and also has the capacity to quantify the bacteria (Mackay, 2004).…”

“…An earlier study by Albuquerque et al (2017) in dairy herds from the Garanhuns microregion where some outbreaks of paratuberculosis were detected, an average of 160CFU/ml MAP was quantified in milk samples from positive cows. In this study, the number of CFU/g was higher (mean of 195.9 CFU/g), and may be justified by the fact that the transformation of milk in to cheese basically consists of a process that concentrates a great amount of milk, in which part of the solid components, mainly protein and fat are concentrated in the curd, while the whey proteins, lactose and soluble solids are removed in the whey (Paula et al, 2009).…”

“…After DNA extractions, the Real-Time Polymerase Chain Reaction (qPCR) was performed as described by Albuquerque et al (2017). It consisted of a reaction final volume of 25.0μL containing: 5μL of genomic DNA; 1μL of the specific primers for IS900 at 10μM (DF: 5'-GACGACTCGACCGCTAATTG-3 'and DR-1: 5'-CCGTAACCGTCATTGTCCAG-3') (Taddei et al, 2008); 5.5μL of ultra-pure mili-Q Water and 12.5μL of QuantiFast SYBR Green PCR Kit (QIAGEN® PCR mix) according to manufacturer instructions.…”

“…In the State of Pernambuco, paratuberculosis has been reported in some studies involving cattle (Mota et al, 2007;Sá et al, 2013) and buffaloes (Mota et al, 2010) in dairy farms. The presence of DNA has also been detected in milk samples of cows from several properties in the same State (Albuquerque et al, 2017). Considering the risk of consumption of milk derivatives as a risk for human infection by MAP, the objective of this study was to investigate the occurrence of MAP DNA in coalho cheese samples from Pernambuco, Brazil.…”

Detection of Mycobacterium paratuberculosis in raw cow’s milk using polymerase chain reaction (PCR) technique

“…In this study, M. paratuberculosis was detected in (6%) 3/50 of tested cow's milk, which indicates direct shedding of the pathogen in the raw milk. Since paratuberculosis is a chronic disease and clinical signs may only appear after years of infection, the secretion of M. paratuberculosis in milk is considered as main risk factor for transmission and spreading of the disease to calves (6,11,20,21). In addition, the zoonotic potential of M. paratuberculosis via consumption of contaminated milk and milk products is of public health importance due to frequent isolation of M. paratuberculosis from Crohn's disease patients (22)(23)(24)(25).…”