Application of F+RNA Coliphages as Source Tracking Enteric Viruses on Parsley and Leek Using RT-PCR

Abstract

The objective of this study was to identify sources of fecal contamination in leek and parsley, by using four different F(+)RNA coliphage genogroups (IV, I indicate animal fecal contamination and II, III indicate human fecal contamination). Three different concentrations (10(2), 10(4), 10(6) pfu/ml) of MS2 coliphage were inoculated on the surface of parsley and leek samples for detection of phage recovery efficiency among two methods of elution concentration (PEG-precipitation and Ultracentrifugation) by performing double agar layer (DAL) assay in three replications. Highest recovery of MS2 was observed in PEG method and in 10(6) inoculation concentration. Accordingly, the PEG method was used for washing and isolation of potentially contaminated phages of 30 collected samples (15 samples from the market and 15 samples from the farm). The final solutions of PEG method were tested for the enumeration of plaques by DAL assay. Total RNA was then extracted from recovered phages, and RT-PCR was performed by using four primer sets I, II, III, and IV. Incidence of F(+)RNA coliphages was observed in 12/15 (80 %) and 10/15 (66/6 %) of samples were obtained from farm and market, respectively, using both DAL and RT-PCR test methods. Different genotypes (I, II, and IV) of F(+)RNA coliphages were found in farm samples, while only genotype I was detected in market samples by using the primer sets. Due to the higher frequency of genotype I and IV, the absence of genotype III, and also the low frequency of genotype II, it is concluded that the contamination of vegetable (parsley and leek) in Neyshabour, Iran is most likely originated from animal sources.

Abundance and Distribution of Enteric Bacteria and Viruses in Coastal and Estuarine Sediments—a Review

“…Advantages of this approach includes, target specificity (each phage is typically specific to one host) and their greater environmental persistence in comparison to FIOs; typically 3-fold longer under controlled conditions (Allwood et al, 2003). In addition, source apportionment can be undertaken using different genogroups of F + RNA coliphages (e.g., I, IV for animal and II and III for human) (Shahrampour et al, 2015). Concentrations of F + RNA coliphages were between 9 and 20 fold higher in sediments than the overlying water column (Alcântara and Almeida, 1995).…”

Application of bacteriophage MS2 in preliminary simulation of inactivation rate of pathogenic viruses in commercial beef and chicken packaging

“…Beef (fillet) and chicken (breasts) meats were purchased from local stores and kept in two forms, minced and chopped to undergo the designed experiments at a freezing temperature of −18 °C (Shahrampour et al ., 2015).…”

“…Male‐specific F‐RNA bacteriophages are now being studied as a surrogate and model for enteric viruses due to their closeness to mammalian viral infections in form, size, morphology, isoelectric point, resistance to environmental stress, and transport properties. These coliphages infect gram‐negative bacterial cells that produce F or sex‐pili (e.g., E. coli K12 and Salmonella typhimurium WG49), and unlike other viral and bacterial indicators, working with them in the laboratory is simple, inexpensive, and poses little risk to human health (Shahrampour et al ., 2015).…”

A bioassay-based protocol for chemical neutralization of human faecal wastes treated by physico-chemical disinfection processes: A case study on benzalkonium chloride

“…Such chemical neutralization of disinfectant chemicals in disinfection experiments is standard practice to allow for reliable culture based assays of faecal indicator bacteria (e.g. Escherichia coli or Enterococcus faecalis (APHA, 2010a; c)) as bacterial pathogen surrogates, bacteriophages such as somatic coliphages (APHA, 2010d;Gall et al, 2016), FRNA phages (Shahrampour et al, 2015), Bacteroides phages (Ebdon et al, 2012), and bacteriophage φ6 (Casanova and Weaver, 2015b) (Casanova and Weaver, 2015a) as virus pathogen surrogates in complex sewage and faecal waste matrices. Failure to achieve adequate chemical neutralization of the disinfectant used in an experiment can result in flawed disinfection kinetics estimates based on CT conditions because inactivation could continue after the experiment is completed due to variable and unknown remaining concentrations of the disinfectant or its biocidal by-products.…”