Development of a dipstick assay format for detection of Salmonellae in food and feeds
Date
2015
Abstract
Salmonellosis remains a public health concern and a problem in the livestock industry. Early detection of the causal organism is a major strategy for control and management of the spread of the disease. Contaminated water, food, feed and infected animals/animal products are the vectors of Salmonellae. Current methods for Salmonella detection include antibody or DNA-based formats or a combination of both. DNA-based detection shows more reliability, particularly when applied through exponential amplification of the target sequences e.g. via the polymerase chain reaction. A dipstick assay was established for the detection of Salmonella via DNA hybridization. This utilized DIG-labeled and unlabeled DNA fragments generated in a PCR reaction using specifically designed primers. The established assay involved the sequential hybridization of the capture probe fixed on the dipstick with the target DNA followed by the detection of the labeled probe. Consistent detection results at 55 deg C hybridization temperature, a 50ng of unlabeled fragment fixed on the nylon membrane can capture a minimum of 10ng Salmonella target DNA were observed. This was detected upon the addition of DIG-labeled detector fragment. The appearance of a definite blue colored spot was observed for positive result on the prepared dipsticks. Initial storage studies of prepared dipsticks at room temperature and 10 deg C for seven weeks in air-tight container exhibited similar positive results. Selective enrichment procedures to complete the dipstick assay protocol were tested on selected food samples. A two-step enrichment was employed to provide a next-day result of the analysis and convenience to the analyst. Other medium for the second stage enrichment which will provide better selectivity in the presence of other pathogens will be evaluated. Some modifications on the hybridization conditions will be conducted to establish the dipstick assay protocol.
Language
English
Document Type
Article
Recommended citation
Sedano, S. A.; Tavanlar, M.A. T.; Perez, M.A.T. M.; Araguirang, G. E.; de Padua, Z. P.; and Alip, T. G., "Development of a dipstick assay format for detection of Salmonellae in food and feeds" (2015). Report. 620.
https://www.ukdr.uplb.edu.ph/reports/620
Pages /Collation
44 leaves
En – AGROVOC descriptors
SALMONELLA; FOODS; FEEDS; DNA HYBRIDIZATION; BACTERIA; DISEASE TRANSMISSION; PCR; ANALYTICAL METHODS