A new integrated approach to analyze bulk tank milk and raw milk filters for the presence of the E. coli serogroups frequently associated with VTEC status

Abstract

A new integrated approach to analyze bulk tank milk and raw milk filters for the presence of the E. coli serogroups frequently associated with VTEC status, Research in Veterinary Science (2017Science ( ), doi: 10.1016Science ( / j.rvsc.2017 This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. AbstractWe optimized a combination of microbiological and molecular methods to quickly identify the presence of the O157 and the six non-O157 serogroups (O26, O45, O103, O111, O121 and O145) most frequently associated with VTEC status, at herd level. The lower detection limit of this methodology is 10 1 CFU/ml for each of the serogroups tested.We tested 67 bulk tank milk (BTM) and raw milk filters (RMF) derived from dairy herds located in Lombardy and Trentino Alto Adige. We identified 3 positive samples and 20 positive samples out of 67 respectively in the BTM and RMF. Interestingly, several samples showed positivity for more than one serogroups at the same time. We also identified the presence of E.coli O45 and O121 for the first time in raw milk and raw milk filters. Once screened the seven serogroups of interest in our ACCEPTED MANUSCRIPT A C C E P T E D M A N U S C R I P T2 samples, we evaluated the real pathogenicity of our positive, non-O157 samples through two parallel molecular biology methods: virulence gene research by PCR, and HRMA and sequencing.The most frequently isolated serogroups in milk were O157 (2.64%), O103 (2.11%), and O145(1.06%), while in RMF the frequencies were, respectively 14.92%, 4.48%, and 2.98%. Moreover, this is the first published report in Italy of positive recovery of O45 and O121 serogroups in milk and milk filters. The new diagnostic approach proposed investigate the presence of the O157 and big six non-O157 serogroups at farm level and not to identify VTEC hazard only once the product is processed and/or is ready to be consumed.

Evaluation of virulence factors profiles and antimicrobials resistance of Escherichia coli isolated from bulk tank milk and raw milk filters

“…This study was designed to contribute to fill this gap by investigating the presence of pathogenic and CoEC in 149 isolates collected from BTM and RMF and their association with AMR pattern. The higher proportion of isolates from RMF, when compared to BTM confirmed the usefulness of this sampling in monitoring the presence of hazards at dairy herd level (Albonico et al, 2017;Murphy et al, 2005). Among all the isolates, 53 (35.6%) were classified as pathogenic while the other ones were classified as CoEC.…”

“…Monitoring the raw milk at herd level showed to be a useful way to detect the presence of enteropathogenic strains and AMR bacteria, before being used to feed calves or to produce food for human consumption such as raw milk or raw milk products (Albonico et al, 2017;Berge et al, 2007). Besides the analysis of BTM, also RMF after milking showed to be a very useful tool to monitor the presence of hazards at milk and herd level (Albonico et al, 2017;Murphy et al, 2005).…”

“…All the samples were stored at 4 °C and delivered under refrigeration to the Department of Veterinary Medicine of the University of Milan within 8 h after collection. Once arrived in the laboratory, the samples were frozen at −20 °C until processed (Albonico et al, 2017).…”

“…The RMF is a component of milking machines, which blocks the entry of debris, large particles of organic material and foreign objects in BTM. The material of the filter is similar to a tissue with a variable weight of 60 to 80 g/m 2 and the analysis of the filter showed to be useful in identifying the presence of foodborne pathogens (Albonico et al, 2017;Murphy et al, 2005).…”

Prevalence and molecular characterization of shiga toxin-producing Escherichia coli in animal source foods and green leafy vegetables

“…Although, E. coli was found in all of the bulk tank milk samples (100%) in this study, none of the isolates had the STEC virulent gene. Albonico et al (2017) found the STEC O45 serotype in only 2 (2.9%) of 67 tank milk samples, as well as O157 and O145 in 1 (1.5%) sample. Another study found the stx gene in 32 (12.5%) of 255 bulk tank milk samples with 4 (12.5%) of these isolates were E. coli O157 and 6 (18.8%) were E. coli O26 serotypes (Trevisani et al, 2014).…”