Detection of food hazards in foods: comparison of real time polymerase chain reaction and cultural methods

Abstract

Foodstuffs should not contain microorganisms or their toxins or metabolites in quantities suggesting an unacceptable risk for human health. The detection of food hazards in foods is performed by several tests that produce results dependent on the analytical method used: an analytical reference method, defined as standard, is associated with each microbiological criterion laid down in Regulation 2073/2005/EC, but, analytical methods other than the reference ones, in particular more rapid methods, could be used. Combined screening methods performed by real time-polymerase chain reaction (RT-PCR) are currently validated as alternative methods according to the ISO 16140:2003 and certified by the Association Française de Normalisation. However, the positive results obtained with these alternative methods, the investigated molecular relations that resulted positive have to be confirmed with cultural methods using the same enrichment media in which the molecular screening was performed. Since it is necessary to assess if these testing schemes provide equivalent guarantees of food safety, the aim of this retrospective study is to analyse the data collected, from 2012 to 2014 by Emilia Romagna Region in the field of Piano Regionale Alimenti (Food Regional Plan) during official controls monitoring food samples of animal and other than animal origin. Records performed by combined methods of molecular screening of Salmonella spp., Listeria monocytogenes and thermophilic Campylobacter and cultural confirmation results were gathered together and the results were compared in order to assess the sensitivity of the methods. A total of 10,604 food samples were considered in this study: the comparison of the data revealed that the RT-PCR method detected Salmonella, L. monocytogenes, and thermophilic Campylobacter in 2.18, 3.85 and 3.73% of the samples, respectively, whereas by using cultural method these pathogens were isolated in 0.43, 1.57 and 1.57% of samples, respectively. In spite of the use of the same enrichment broth, the RT-PCR method disclosed a percentage of positive samples that was negative to cultural examination ranging between 20 and 43%, with a PCR/culture ratio between 2.37 to 5.00. In conclusion, the results of this study pose a doubt about the sensitivity of the official cultural methods regarding the isolation of the three investigated foodborne pathogens. Moreover this study may be a useful tool for veterinary authorities to assess appropriate sampling plans to control the risk relating to the consumption of contaminated foods.

Campylobacter Infections in Children

“…Reverse transcriptase polymerase chain reaction identifies Campylobacter from stool 20% to 40% more frequently than culture-based methods. (47) However, because these tests identify the presence of nucleic acid rather than viable organisms, the clinical significance is not always clear. The identification of multiple pathogens is not uncommon and can be difficult to interpret.…”

Point-of-Need Diagnostics for Foodborne Pathogen Screening

“…These culturing methods consist of sample homogenization and multistep selective enrichment, followed by various plating steps for colony isolation. 7,8 Detection is traditionally performed with phenotypic analysis and biochemical confirmation via Gram staining, metabolic assays, and ribotyping. [6][7][8] Despite the reliability of culture-based techniques, the laborious and time-consuming nature and requirement of trained lab personnel, lab equipment, and sterile environments prevent the universal utilization of such methods.…”

“…7,8 Detection is traditionally performed with phenotypic analysis and biochemical confirmation via Gram staining, metabolic assays, and ribotyping. [6][7][8] Despite the reliability of culture-based techniques, the laborious and time-consuming nature and requirement of trained lab personnel, lab equipment, and sterile environments prevent the universal utilization of such methods. 6,9,10 Notably, the 2-3 days required for initial results and 7 days for species confirmation impede timely pathogen detection prior to food consumption.…”

“…Furthermore, culturing techniques reveal challenges in detecting fastidious and viable but not culturable species, especially among samples with high levels of competing microorganisms. 7,11 Recently developed optical sensors based on the analysis of characteristic light diffraction patterns (BARDOT, 12 Scatterometer, 13 converging spherical wave illumination, 14 etc.) have been useful in rapidly differentiating plated bacterial species within a genus and serovars within a species, such as Shiga toxin-producing E. coli (STEC) serogroups.…”

Microfluidic droplet application for bacterial surveillance in fresh-cut produce wash waters

“…During the 1998 to 2008 reporting period, nearly half (46%) of all foodborne illnesses in U.S. were attributable to produce, with 22% attributable to leafy greens alone [9]. Given the global threat of foodborne contamination, the food industry must continually evaluate critical control points (CCPs) for its most vulnerable crops, improve upon antiquated detection methods, and maintain a collaborative relationship with national and international surveillance networks [10][11][12].…”