ABSTRACT
Detection of Salmonella sp. is important for the broiler chicken production chain because it is one microorganisms involved in food-borne diseases. Thus, this study performed the optimization of a technique of Loop-mediated isothermal DNA amplification (LAMP) through the 3MTM Molecular Detection Assay 2: Salmonella (MDS®), in accordance with Ordinance number 126 of the Ministry of Agriculture, for the detection of Salmonella sp. in drag swab. The methodology followed ISO 16140-2: 2016, with the analysis naturally contaminated drag swab samples collected from broiler aviaries and artificially contaminated with salmonella ATCCs. Of the 300 samples processed in protocol A (pre-enrichment tetrathionate broth (TT)), 45 were positive for Salmonella sp., 242 negative, one false-positive, and 12 false-negative, while of the 300 samples analyzed in protocol B (pre-enrichment brain-heart infusion broth (BHI)), 40 were positive, 256 negative, one false-positive, and three false-negative. The result for protocol A was a sensitivity of 79%, specificity of 99.6%, Positive Predictive Value (PPV) of 98%, and Negative Predictive Value (NPV) of 95%; and for protocol B, 93% sensitivity, 99.6% specificity, 98% PPV, and 99% NPV. Both protocols were associated with the reference method (p>0.05), concluding that the MDS® can be used for the qualitative detection of Salmonella sp.
Subject(s)
Chickens , Nucleic Acid Amplification Techniques , Salmonella , Sensitivity and Specificity , Salmonella/isolation & purification , Salmonella/genetics , Animals , Nucleic Acid Amplification Techniques/methods , Chickens/microbiology , Molecular Diagnostic Techniques/methods , Salmonella Infections, Animal/diagnosis , Salmonella Infections, Animal/microbiology , DNA, Bacterial/analysis , Reproducibility of ResultsABSTRACT
The storage of fresh raw milk at low temperature does not prevent proliferation of psychrotrophic bacteria that can produce heat-resistant proteolytic enzymes contributing to the reduced shelf life of dairy products. This study aimed to identify the dominant psychrotrophic proteolytic enzyme-producing population of raw milk from Brazil. Raw milk samples collected in 3 different cooling tanks in Brazil were stored at optimal (45 h at 4 °C followed by 3 h at 7 °C) and suboptimal (45 h at 7 °C followed by 3 h at 10 °C) conditions to simulate farm storage and transportation allowed by Brazilian laws. The highly proteolytic enzyme-producing strains isolated from stored cold raw milk were characterized by repetitive sequence-based Polymerase Chain Reaction (PCR) analysis. This clustering resulted in 8 different clusters and 4 solitary fingerprints. The most proteolytic isolates from each rep-cluster were selected for identification using miniaturized kit, 16S rDNA and rpoB gene sequencing. Serratia liquefaciens (73.9%) and Pseudomonas spp. (26.1%) were identified as the dominant psychrotrophic microorganisms with high spoilage potential. The knowledge of milk spoilage microbiota will contribute to improved quality of milk and dairy products.