Mickey E. Parish

Standardization of a Method To Determine the Efficacy of Sanitizers in Inactivating Human Pathogenic Microorganisms on Raw Fruits and Vegetables

The efficacy of sanitizers in killing human pathogenic microorganisms on a wide range of whole and fresh-cut fruits and vegetables has been studied extensively. Numerous challenge studies to determine the effects of storage conditions on survival and growth of pathogens on raw produce have also been reported. Results of these studies are often difficult to assess because of the lack of sufficient reporting of methods or, comparatively, because of variations in procedures for preparing and applying inocula to produce, conditions for treatment and storage, and procedures for enumerating pathogens. There is a need for a standard method to accurately determine the presence and populations of pathogenic microorganisms on produce. The adoption of standard, well-characterized reference strains would benefit a comparative assessment of a basic method among laboratories. A single protocol will not be suitable for all fruits and vegetables. Modifications of a basic method will be necessary to achieve maximum recovery of pathogens on various types of produce subjected to different sanitizer or storage treatments. This article discusses parameters that must be considered in the course of developing a basic standard method against which these modifications could be made.

Survival of Listeria monocytogenes in Low pH Model Broth Systems1

Four strains of Listeria monocytogenes were studied for pH tolerance in pH-adjusted tryptic soy broth supplemented with yeast extract. After incubation at 30°C, all strains grew at pH 4.5 and above. Survival of strain F5069(4b) was further investigated in sterile orange serum in which pH was adjusted from 3.6 to 5.0 in 0.2 unit increments. At 4°C, viable cells were reduced from 106 cfu/ml to less than 25 cfu/ml in 25 d at pH 3.6, 43 d at pH 4.0, and 81 d at pH 4.6. Viable cell populations remained at levels about 102 after 90 d in orange serum at pH 4.8 and 5.0. At 30°C, the same reductions were observed at 5 d at pH 3.6 and 4.0 and 8 d at pH 5.0. Growth in orange serum was observed prior to the reduction in viable cell numbers at pH 4.8 and 5.0 for both storage temperatures.

Coliforms, Escherichia coli and Salmonella Serovars Associated with a Citrus-Processing Facility Implicated in a Salmonellosis Outbreak†

A salmonellosis outbreak occurred during the summer of 1995 among individuals who consumed nonpasteurized orange juice from a Florida citrus-processing facility. Clinical isolates were identified by the Centers for Disease Control as Salmonella serovars Hartford, Gaminara, and Rubislaw. At the processing facility, 70 samples (equipment swabs, fruit surface swabs, juice, and miscellaneous environmental samples) were collected before, during, and after processing runs on two different dates. Bottled juice samples from eight previous extraction dates were also collected. Total plate counts, fecal coliforms, and Escherichia coli were enumerated from each sample. Analysis for Salmonella cells were conducted on all juice samples, fruit surface swabs, environmental samples, and selected equipment swabs using direct enrichment and pre-enrichment techniques. Salmonella serovars Hartford, Rubislaw, Saintpaul, and Newport were detected from either juice, unwashed fruit surfaces, or amphibians (Hyla cinerea and Bufo terrestris) captured outside the processing building. Salmonella cells in juice were associated with population levels of fecal coliforms and E. coli above the upper most probable number (MPN) limits of detection (> 110/ml).

Foodborne illness outbreaks from microbial contaminants in spices, 1973-2010.

This review identified fourteen reported illness outbreaks attributed to consumption of pathogen-contaminated spice during the period 1973-2010. Countries reporting outbreaks included Canada, Denmark, England and Wales, France, Germany, New Zealand, Norway, Serbia, and the United States. Together, these outbreaks resulted in 1946 reported human illnesses, 128 hospitalizations and two deaths. Infants/children were the primary population segments impacted by 36% (5/14) of spice-attributed outbreaks. Four outbreaks were associated with multiple organisms. Salmonella enterica subspecies enterica was identified as the causative agent in 71% (10/14) of outbreaks, accounting for 87% of reported illnesses. Bacillus spp. was identified as the causative agent in 29% (4/10) of outbreaks, accounting for 13% of illnesses. 71% (10/14) of outbreaks were associated with spices classified as fruits or seeds of the source plant. Consumption of ready-to-eat foods prepared with spices applied after the final food manufacturing pathogen reduction step accounted for 70% of illnesses. Pathogen growth in spiced food is suspected to have played a role in some outbreaks, but it was not likely a contributing factor in three of the larger Salmonella outbreaks, which involved low-moisture foods. Root causes of spice contamination included contributions from both early and late stages of the farm-to-table continuum.

Yeasts and molds isolated from spoiling citrus products and by-products

A variety of citrus products, including dried peel used as livestock fodder, unpasteurized orange juice and pasteurized, and chilled orange juice were selectively surveyed for their fungal microflora. Fungi isolated were 1) pasteurized orange juice: Aureobasidium pullulans , Cladosporium sp., and Penicillium sp., 2) unpasteurized orange juice: Candida maltosa , C. sake , Fusarium sp., Geotrichum sp., Hanseniaspora sp., H. guilliermondii , Penicillium sp., Pichia membranaefaciens , Saccharomyces cerevisiae , Schwanniomyces occidentalis , and Torulaspora delbrueckii and 3) dried peel: Aspergillus niger , Aspergillus sp., Byssochlamys sp., Fusarium sp., Penicillium sp., Rhizopus sp., Rhodotorula sp., Sc. occidentalis , and Trichoderma sp.

Microscopic Observation and Processing Validation of Fruit Sanitizing Treatments for the Enhanced Microbiological Safety of Fresh Orange Juice

Studies were conducted to evaluate the infiltration of dye and bacteria into the interior of orange fruit and the impact of possible infiltration on achieving a 5-log microbial reduction during fresh juice processing. Fresh orange fruit were treated at the stem end area with dye and either Salmonella Rubislaw or Escherichia coli strains expressing green fluorescent protein. Microscopic images showed that bacterial contaminants localized at the surface or near surface areas that may be sanitized by surface treatments. Dye infiltration was not a reliable indicator of bacterial penetration in citrus fruit. To quantify the reduction of bacterial contamination, orange fruit were inoculated with E. coli and processed with and without hot water treatments. Greater than 5-log reductions were achieved in juice extracted from fruit immersed in hot water for 1 or 2 min at 80 degrees C, in comparison to the E. coli level detected in the control juice obtained by homogenization of inoculated fruit.

Singleton Sequence Type 382, an Emerging Clonal Group of Listeria monocytogenes Associated with Three Multistate Outbreaks Linked to Contaminated Stone Fruit, Caramel Apples, and Leafy Green Salad

ABSTRACT Three multistate outbreaks between 2014 and 2016, involving case patients in and outside the United States, were linked to stone fruit, caramel apples, and packaged leafy green salad contaminated with Listeria monocytogenes singleton sequence type 382 (ST382), a serotype IVb-v1 clone with limited genomic divergence. Isolates from these outbreaks and other ST382 isolates not associated with these outbreaks were analyzed by whole-genome sequencing (WGS) analysis. The primary differences among ST382 strains were single nucleotide polymorphisms (SNPs). WGS analysis differentiated ST382 from a clonal complex 1 outbreak strain co-contaminating the caramel apples. WGS clustered food, environmental, and clinical isolates within each outbreak, and also differentiated among the three outbreak strains and epidemiologically unrelated ST382 isolates, which were indistinguishable by pulsed-field gel electrophoresis. ST382 appeared to be an emerging clone that began to diverge from its ancestor approximately 32 years before 2016. We estimated that there was 1.29 nucleotide substitution per genome (2.94 Mbp) per year for this clone.

Survey of Foodborne Pathogens, Aerobic Plate Counts, Total Coliform Counts, and Escherichia coli Counts in Leafy Greens, Sprouts, and Melons Marketed in the United States

The objective of this research was to assess the microbiological status of leafy greens, sprouts, and melons from U.S. markets. A total of 14,183 samples of leafy greens, 2,652 samples of sprouts, and 3,411 samples of melons were collected throughout the United States from 2009 to 2014. The samples were analyzed for aerobic plate counts, total coliform counts, Escherichia coli counts, and the presence and levels of Salmonella, Shigella, Listeria monocytogenes, and Shiga toxin-producing E. coli (STEC), depending on the year and type of produce. Among the leafy greens, no E. coli O157:H7 or non-O157 STEC were detected from iceberg lettuce samples. The overall prevalences of Salmonella, E. coli O157:H7, non-O157 STEC, and L. monocytogenes in the 14,183 samples of leafy greens were 0.05, 0.01, 0.07, and 0.11%, respectively. Among sprout samples, no Salmonella or E. coli O157:H7 was detected, and the overall prevalences of non-O157 STEC and L. monocytogenes were 0.04 and 0.11%, respectively. Among melon samples, no Salmonella was detected from cucumbers, no L. monocytogenes was detected from cantaloupes, and the overall prevalences of Salmonella and L. monocytogenes were 0.12 and 0.23%, respectively. L. monocytogenes levels were 0.4 to 1,470 most probable number (MPN)/g in leafy greens, 0.36 to 1,100 MPN/g in sprouts, and <0.03 to 150 MPN/g in melons, and most positive samples had low levels of these pathogens. The isolates from these foods were very diverse genetically. Foodborne pathogens, including Salmonella, STEC, and L. monocytogenes, had relatively low prevalences in the produce surveyed. Because these foods are usually consumed raw, measures should be taken to significantly minimize the presence and levels of human pathogens.