IAFP 2009 Program Book and Abstract Book - AB176

Purpose: This study aimed to isolate and characterize bacteriophages specific for B. cereus and able to cause complete lysis of the target organism. Methods: Sewage and sludge samples were collected and mixed with a cocktail of B. cereus strains to test for the presence of bacteriophages. Isolation of bacteriophages was based on the plaque size and shape. Transmission electron microscopy was carried out to examine phage morphology. Different strains of B. cereus as well as other Bacillus species were used to determine the host specificity of the isolated phages. The sizes and restriction patterns of the phage DNA was determined. The infectivity rate was tested under the stress of some chemical factors such as chloroform, and physical factors such as ambient and high temperatures. Results: Four bacteriophages were isolated and purified. TEM images showed three phages with isometric heads and long non-contractile tails, believed to belong to the family Siphoviridae. A fourth phage with an isometric head and contractile tail was provisionally identified as belonging to the Myoviridae. The isolated phages showed different degrees of infectivity and specificity against tested organisms. Different restriction enzymes showed distinct restriction DNA patterns for the isolated phages. Also the phages showed different responses depending on the level of the stress of chemical and physical factors. Significance: This study recommends using the isolated phages to control B. cereus and suggests that the effect will be maximized if combined with other preservation methods. P3-75 Isolation and Characterization of Lytic Bacteriophages against Enterohemorrhagic Escherichia coli DSC STELIOS VIAZIS, Mastura Akhtar, Joellen Feirtag and Francisco Diez-Gonzalez University of Minnesota, Food Science and Nutrition, St. Paul, MN, USA Introduction: Since the 1980s, Enterohemorrhagic Escherichia coli (EHEC) strains have been recognized as foodborne pathogens and serotype O157:H7 has caused frequent gastroenteritis outbreaks. Bacteriophages can be used as a natural non-antibiotic method with the potential to reduce bacterial pathogens, such as EHEC, from the food supply. Purpose: The objective of this study was to isolate, identify and characterize a diverse collection of lytic bacteriophages capable of infecting EHEC serotypes O26, O111 and O157. Methods: Phages previously isolated were provided by Dr. Andrew Brabban and Dr. Lawrence Goodridge. Additional phages were isolated from dairy and feedlot manure using EHEC O157, O26 and O111 strains as hosts. Manure was enriched using Tryptic Soy (TS) Broth and exponentially growing cultures of specific bacterial hosts, and was incubated overnight at 37°C. The enriched extracts were centrifuged, filtered, combined with the host strain in tryptone top agar, and plated on TS agar. Plaques were purified and screened against additional strains (14, O157; 10, O26; 10, O111) using the efficiency of plaquing method (EOP). Purified phages were observed using transmission electron microscopy (TEM) after staining with 2% ammonium molybdate. Results: Phage CEV2 and five other phages previously isolated were able to lyse all 14 O157 strains with EOP values consistently above 0.001. Four out of six phages isolated from a cattle feedlot were effective against all O157 strains and one O26 strain with EOP values greater than 0.001. After spot testing, they were found to be effective against seven O26 strains. Five phages isolated from dairy manure were highly specific against O26 strains but were not effective against other serotypes. Based on TEM, some of the phages were classified as Group 1 Caudovirales. Significance: These results indicated that the isolated bacteriophages were highly effective against multiple strains of two EHEC serotypes. This collection of phages can be grouped and potentially used as an antimicrobial cocktail to inactivate O157 and O26 serotypes and reduce their incidence in the food chain. P3-76 Analysis of Antimicrobial Resistance in Enterococcus spp. Recovered from a Commercial Beef Processing Plant MUEEN ASLAM, Cara Service, Heidi Rempel and Moussa Diarra Lacombe Research Centre, Agriculture and Agri-Food Canada, Lacombe, AB, Canada Introduction: Antimicrobial resistant enterococci are considered to be a human health concern, as they can provide a pool of resistance genes transferable to pathogens. Purpose: The goal of this study was to analyze antimicrobial- resistant (AMR) phenotypes and resistance genes in enterococci isolates from a commercial beef processing plant. Methods: Samples were obtained from conveyers used for moving carcasses, before the start of operation (CC), two hours after operation has started (DC), and from ground beef (GB). Samples were screened for presumptive enterococci on KF agar, and three isolates from each positive sample (26 from CC; 27 from DC; 26 from GB) were randomly selected and confirmed to species level using PCR and the API® 20 Strep kit. A total of 201 isolates comprising 48, 81 and 72 from CC, DC, and GB, respectively, were analyzed for AMR and resistance genes and were genetically characterized. The AMR was determined by use of a sensititre system®, and results were interpreted according to the criteria of the Clinical Laboratory Standard Institute. PCR was used for the detection of resistance genes. Results: Enterococcus faecalis (80%) was the most common species found, followed by E. faecium (6%) and E. durans (3%). Complete resistance to tetracycline and lincomycin was found in 48% and 10% of isolates, respectively, and 83%, 59% and 37% of isolates were intermediate resistant to lincomycin, erythromycin and ciprofloxacin, respectively. Resistance to quinupristin/dalfopristin was found in 36% of E. faecium isolates. Tetracycline resistant enterococci were positive for tetM gene, whereas ermB and vatE genes were found in 18% and 49% of enterococci. The analysis showed that the majority of enterococci from individual sample sources were genetically related. Data on the prevalence of antimicrobial resistant enterococci and resistant genes among various sample sources will be discussed. Significance: These findings suggest that antimicrobial resistant Enterococcus spp. are prevalent during commercial beef processing. This study underscores the need for improving hygienic conditions in meat processing plants to improve the safety of meat. 176 — Abstract Book Poster

IAFP 2009 Program Book and Abstract Book

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