Jeff Farber

Assessment of the microbiological quality of ready-to-use vegetables for health-care food services

The microbiological quality of ready-to-use (RTU) vegetables, including chopped lettuce, salad mix, carrot sticks, cauliflower florets, sliced celery, coleslaw mix, broccoli florets, and sliced green peppers was determined before and after processing. Microbial profiles were obtained 24 h after processing and on days 4, 7, and 11 after storage at 4 and 10°C to simulate temperature abuse. In addition, the microbial profiles of four RTU vegetables, coleslaw mix, salad mix, cauliflower florets, and sliced green peppers were determined 7 days after distribution to a select group of Ontario hospitals. RTU vegetables, with the exception of green peppers, showed up to a 1-log decrease in aerobic colony counts after processing. These counts increased to preprocessing levels after 4 days of storage at both 4 and 10°C. RTU vegetables stored at temperature abuse conditions (10°C)had significantly higher counts (P < 0.001) on days 4 to 11 as compared to those stored at 4°C. Green peppers had the highest bacterial counts while cauliflower and chopped lettuce had the lowest counts at both storage temperatures (P < 0.05). Increased levels of Listeria monocytogenes in RTU vegetables were associated with temperature abuse. Levels of >100 MPN/g for L. monocytogenes were detected in 8 of 120 (6.7%) samples stored at 10°C but not in 175 samples stored at 4°C after 7 days (P < 0.05). Overall, L. monocytogenes was detected in 13 of 120 (10.8%) RTU vegetables stored for up to 11 days at 10°C and 5 of 176 (2.8%) samples stored at 4°C (P < 0.05). E. coli was detected in 2 of the 120 (1.7%) processed RTU vegetables after day 7 of storage at 10°C and 1 of the 65 (1.5%) unprocessed vegetables from the same batches of vegetables used for processing. This indicator organism was not detected in RTU vegetable samples stored at 4°C or in any of the RTU vegetable samples obtained from hospital coolers. Other pathogenic bacteria, such as Salmonella spp., Campylobacter spp., Yersinia enterocolitica (serotype O:3) and verocytotoxigenic E. coli (VTEC) were not detected in any of the RTU vegetables tested, Recommendations regarding processing, distribution, and storage of these products are presented.

Foodborne spread of hepatitis A: Recent studies on virus survival, transfer and inactivation.

Hepatitis A virus (HAV) is responsible for considerable morbidity and economic losses worldwide, and is the only reportable, foodborne viral pathogen in Canada. Outbreaks caused by it occur more frequently in settings such as hospitals, daycare centres, schools, and in association with foods and food service establishments. In recent years, the incidence of hepatitis A has increased in Canada. Many factors, including changing lifestyles and demographics, faster and more frequent travel, and enhanced importation of foods from hepatitis A-endemic regions, may be behind this increase. Despite its increasing significance as a human pathogen, not much was known until recently about the survival and inactivation of HAV, and even less was understood about the effectiveness of measures to prevent and control its foodborne spread. Studies conducted in the past decade have shown that HAV can survive for several hours on human hands and for several days on environmental surfaces indoors. The virus can also retain its infectivity for several days on fruits and vegetables which are often consumed raw, and such imported items have already been incriminated in disease outbreaks. Casual contact between contaminated hands and clean food items can readily lead to a transfer of as much as 10% of the infectious virus. HAV is also relatively resistant to inactivation by heat, gamma irradiation and chemical germicides. In view of these findings, better approaches to prevent the contamination of foods with HAV and more effective methods for its inactivation in foods, on environmental surfaces and on the hands of food handlers are needed.

An overview of microbial food safety programs in beef, pork, and poultry from farm to processing in Canada.

Canadas vision for the agri-food industry in the 21st century is the establishment of a national food safety system employing hazard analysis and critical control point (HACCP) principles and microbiological verification tools, with traceability throughout the gate-to-plate continuum. Voluntary on-farm food safety (OFFS) programs, based in part on HACCP principles, provide producers with guidelines for good production practices focused on general hygiene and biosecurity. OFFS programs in beef cattle, swine, and poultry are currently being evaluated through a national recognition program of the Canadian Food Inspection Agency. Mandatory HACCP programs in federal meat facilities include microbial testing for generic Escherichia coli to verify effectiveness of the processors dressing procedure, specific testing of ground meat for E. coli O157:H7, with zero tolerance for this organism in the tested lot, and Salmonella testing of raw products. Health Canadas policy on Listeria monocytogenes divides ready-to-eat products into three risk categories, with products previously implicated as the source of an outbreak receiving the highest priority for inspection and compliance. A national mandatory identification program to track livestock from the herd of origin to carcass inspection has been established. Can-Trace, a data standard for all food commodities, has been designed to facilitate tracking foods from the point of origin to the consumer. Although much work has already been done, a coherent national food safety strategy and concerted efforts by all stakeholders are needed to realize this vision. Cooperation of many government agencies with shared responsibility for food safety and public health will be essential.

Rotaviruses from Canadian farm samples

Animal rotavirus (RoV) strains detected in Canadian swine and dairy cattle farms were characterized by sequence analysis of viral protein 4 (VP4), VP6, VP7 and non-structural protein 4 segments from 15 RoV strains. Some porcine strains were found to contain a mixture of segments typical of human and animal viruses. One strain represented a novel VP6 genotype “I14”, G2-P[27]-I14. Other strains detected in porcine samples represented multiple different segment types. These results illustrate the active evolution of animal RoV strains and underline the need for surveillance of both animal and human strains in public health-monitoring programs.

Microfluidic filtration and extraction of pathogens from food samples by hydrodynamic focusing and inertial lateral migration.

Detecting pathogenic bacteria in food or other biological samples with lab-on-a-chip (LOC) devices requires several sample preparation steps prior to analysis which commonly involves cleaning complex sample matrices of large debris. This often underestimated step is important to prevent these larger particles from clogging devices and to preserve initial concentrations when LOC techniques are used to concentrate or isolate smaller target microorganisms for downstream analysis. In this context, we developed a novel microfluidic system for membrane-free cleaning of biological samples from debris particles by combining hydrodynamic focusing and inertial lateral migration effects. The microfluidic device is fabricated using thermoplastic elastomers being compatible with thermoforming fabrication techniques leading to low-cost single-use devices. Microfluidic chip design and pumping protocols are optimized by investigating diffusive losses numerically with coupled Navier–Stokes and convective-diffusion theoretical models. Stability of inertial lateral migration and separation of debris is assessed through fluorescence microscopy measurements with labelled particles serving as a model system. Efficiency of debris cleaning is experimentally investigated by monitoring microchip outlets with in situ optical turbidity sensors, while retention of targeted pathogens (i.e., Listeria monocytogenes) within the sample stream is assessed through bacterial culture techniques. Optimized pumping protocols can remove up to 50xa0% of debris from ground beef samples while percentage for preserved microorganisms can account for 95xa0% in relatively clean samples. However, comparison between inoculated turbid and clean samples (i.e., with and without ground beef debris) indicate some degree of interference between debris inertial lateral migration and hydrodynamic focusing of small microorganisms. Although this interference can lead to significant decrease in chip performance through loss of target bacteria, it remains possible to reach 70xa0% for sample recovery and more than 50xa0% for debris removal even in the most turbid samples tested. Due to the relatively simple design, the robustness of the inertial migration effect itself, the high operational flow rates and fabrication methods that leverage low-cost materials, the proposed device can have an impact on a wide range of applications where high-throughput separation of particles and biological species is of interest.

Qualitative Microbiological Risk Assessment of Unpasteurized Fruit Juice and Cider

Until recent decades, unpasteurized fruit juice and cider have been considered non-hazardous with respect to microbiological pathogens due to their acidic nature. However, in light of the many global foodborne illness outbreaks associated with these products, it is apparent that certain bacterial, viral and parasitic pathogens can survive these acidic conditions and remain infectious. Specifically, outbreaks of human illness have been attributed to infection with Escherichia coli O157:H7, Salmonella spp., Shigella spp., Cryptosporidium spp., Trypanosoma cruzi, and hepatitis A, and have been associated with the consumption of apple juice or cider, orange juice and various other types of unpasteurized juices. The most likely mechanisms by which juice, and the fruit it is processed from, becomes contaminated with pathogenic microorganisms are through direct contact with animal or human faeces, or indirect contact with contaminated water, soil, processing equipment, or infected food handlers. This risk assessment reviews foodborne outbreaks linked to unpasteurized fruit juice and cider, and evaluates the evidence for effectiveness of measures to control pathogens in these products.

Assessment of Sources of Exposure for Mycobacterium avium subsp. paratuberculosis in Food and Water

Mycobacterium avium subsp. paratuberculosis (MAP) is believed to be the causative agent of paratuberculosis (Johnes disease) in ruminants. MAP infection has also been proposed as the cause of Crohns disease (CD) in humans. An increasing number of recent studies suggest some association between MAP and CD, however a cause-effect relationship has yet to be proved or disproved. Infected cattle appear to be the most important source of human exposure to MAP, and the associated suspected vehicles of transmission are milk, dairy products and beef.

Rapid and Specific Enzyme Immunoassay on Hydrophobic Grid Membrane Filter for Detection and Enumeration of Thermophilic Campylobacter spp. from Milk and Chicken Rinses

Six commercially available anti-Campylobacter antibodies were examined for their applicability in an enzyme immunoassay on hydrophobic grid membrane filters, both for the detection and enumeration of Campylobacter spp. When a panel of nine Campylobacter (seven Campylobacter jejuni and two Campylobacter coli) and eight non-Campylobacter strains were used in a dot-blot format enzyme immunoassay to test the specificity of these antibodies, only one polyclonal antibody (Biodesign) detected all Campylobacter strains. Escherichia coli O157:H7 produced weak nonspecific signals due to endogenous peroxidase activity. The specificity of this Biodesign antibody was further tested against 30 more Campylobacter strains and more than 600 non-Campylobacter strains on hydrophobic grid membrane filters grown on modified Campylobacter agar with charcoal and deoxycholate, a Campylobacter selective medium. All the Campylobacter strains were detected, whereas only two (Acinetobacter calcoaceticus, Salmonella Minnesota) of the approximately 130 non-Campylobacter strains, which grew on modified Campylobacter agar with charcoal and deoxycholate, gave false-positive signals. This simple, rapid, and specific enzyme immunoassay also detected Campylobacter spp. from inoculated milk and chicken rinses and naturally contaminated chicken rinses.

Neptune: a bioinformatics tool for rapid discovery of genomic variation in bacterial populations

Abstract The ready availability of vast amounts of genomic sequence data has created the need to rethink comparative genomics algorithms using ‘big data’ approaches. Neptune is an efficient system for rapidly locating differentially abundant genomic content in bacterial populations using an exact k-mer matching strategy, while accommodating k-mer mismatches. Neptune’s loci discovery process identifies sequences that are sufficiently common to a group of target sequences and sufficiently absent from non-targets using probabilistic models. Neptune uses parallel computing to efficiently identify and extract these loci from draft genome assemblies without requiring multiple sequence alignments or other computationally expensive comparative sequence analyses. Tests on simulated and real datasets showed that Neptune rapidly identifies regions that are both sensitive and specific. We demonstrate that this system can identify trait-specific loci from different bacterial lineages. Neptune is broadly applicable for comparative bacterial analyses, yet will particularly benefit pathogenomic applications, owing to efficient and sensitive discovery of differentially abundant genomic loci. The software is available for download at: http://github.com/phac-nml/neptune.

For the Safety of Fresh Produce: Regulatory Considerations for Canada on the Use of Whole Genome Sequencing to Subtype Salmonella

Salmonella is one of the oldest bacteria known to man, yet it is also one of the most prevalent when it comes to foodborne-related diseases and outbreaks. Naturally present in the environment and difficult to treat on fresh produce, Salmonella represents an important food safety challenge. Emerging technologies such as whole genome sequencing (WGS) and next generation sequencing (NGS) now offer promising applications within the realm of food safety that can significantly change the way routine testing, inspections and disease surveillance are done. They offer potential avenues that may foster more sustainable agricultural and environmental practices to detect and reduce the presence of Salmonella. Strategies are being developed to better cluster, integrate and share genomic data to facilitate the development of diagnostic tests and control methods, as well as generate robust evidence to better inform future policy and regulatory decision-making. Using the approaches developed by the Salmonella Syst-OMICS consortium, a large-scale Canadian-based genomic project, this paper discusses the policy and regulatory considerations for the applications of WGS and NGS technologies in the development of testing and biocontrol tools for food safety. The paper presents an overview of the current regulatory framework for the approval of testing methodologies for Salmonella. It discusses considerations related to (1) the development of a new test for Salmonella, (2) the potential establishment of a Salmonella risk virulence classification scheme, and (3) the development of a biocontrol method to reduce the presence of Salmonella on fresh produce.