Search

Salsa-associated outbreaks, including the large multistate outbreak in the United States in 2008 caused by jalapeño and serrano peppers contaminated with SalmonellaSaintpaul, have raised concerns about salsa as a potential vehicle for transmission. Despite these events, there has been relatively limited research on the potential growth of pathogenic bacteria in salsa. The aim of this study was to characterize the survival and growth of Salmonella,including the outbreak strain of SalmonellaSaintpaul (E2003001236), in freshly made salsa and its main ingredients. Chopped tomatoes, jalapeño peppers, cilantro, and onions were tested individually or mixed according to different salsa recipes. Samples were inoculated with five Salmonellaserotypes at 3 log CFU/g: Saintpaul (various strains), Typhimurium, Montevideo, Newport, or Enteritidis. Samples were then stored at room temperature (23°C) for up to 12 h or 3 days. The SalmonellaSaintpaul levels reached approximately 9 log CFU/g after 2 days in tomato, jalapeño pepper, and cilantro. Growth was slower in onions, reaching 6 log CFU/g by day 3. Salsa recipes, with or without lime juice, supported the growth of SalmonellaSaintpaul, and final levels were approximately 7 log CFU/g after 3 days at 23°C. In contrast, the counts of SalmonellaTyphimurium, SalmonellaMontevideo, SalmonellaNewport, and SalmonellaEnteritidis increased only 2 log CFU/g after 3 days in any of the salsas. Other SalmonellaSaintpaul strains were able to grow in salsas containing 10% lime juice, but their final levels were less than 5 log CFU/g. These findings indicate the enhanced ability of the SalmonellaSaintpaul outbreak strain to grow in salsa compared with other Salmonellastrains. Recipe modifications including but not limited to adding lime juice (at least 10%) and keeping fresh salsa at room temperature for less than 12 h before consumption are strategies that can help mitigate the growth of Salmonellain salsa.

Community-associated Clostridium difficileinfection (CA-CDI) now accounts for approximately 50% of CDI cases in central Minnesota; animals and meat products are potential sources. From November 2011 to July 2013, we cultured retail meat products and fecal samples from food-producing and companion animals in central Minnesota for C. difficileby using standard methods. The resulting 51 C. difficileisolates, plus 30 archived local veterinary C. difficileisolates and 208 human CA-CDI case isolates from central Minnesota (from 2012) from the Minnesota Department of Health, were characterized molecularly, and source groups were compared using discriminant analysis. C. difficilewas recovered from 0 (0%) of 342 retail meat samples and 51 (9%) of 559 animal fecal samples. Overall, the 81 animal source isolates and 208 human source isolates were highly diverse genetically. Molecular traits segregated extensively in relation to animal versus human origin. Discriminant analysis classified 95% of isolates correctly by source group; only five (2.5%) human source isolates were classified as animal source. These data do not support meat products or food-producing and companion animals as important sources of CA-CDI in the central Minnesota study region.

Foods have been identified as a potential target for bioterrorism due to their essential nature and global distribution. Foods produced in bulk have the potential to have large batches of product intentionally contaminated, which could affect hundreds or thousands of individuals. Bacillus anthracisspores are one potential bioterrorism agent that may survive pasteurization and remain viable throughout the shelf life of fruit juices and cause disease if consumed. This project examined B. anthracisspore survival in orange, apple, and grape juices, as well as wine. Samples of beverages were inoculated with spores of two nonpathogenic B. anthracisstrains at approximately 106CFU/ml, and the spore count was determined periodically during storage for 30 days at 4°C. After this time, the counts of survival spores never declined more than 1 log CFU/ml in any of the beverage types. These results indicate that spores can survive, with little to no loss in viability, for at least a month in fruit juices and wine.

The growth of Listeria monocytogenesinoculated on frankfurters at four inoculum levels (0.1, 0.04, 0.01, and 0.007 CFU/g) was examined at 4, 8, and 12°C until the time L. monocytogenespopulations reached a detectable limit of at least 2 CFU/g. A scaled-down assumption was made to simulate a 25-g sample from a 100-lb batch size in a factory setting by using a 0.55-g sample from a 1,000-g batch size in a laboratory. Samples of 0.55g were enriched in PDX-LIB selective medium, and presumptive results were confirmed on modified Oxford agar. Based on the time to detect (TTD) from each inoculum level and at each temperature, a shelf life model was constructed to predict the detection or risk levels reached by L. monocytogeneson frankfurters. The TTD increased with reductions in inoculum size and storage temperature. At 4°C the TTDs (±standard error) observed were 42.0 ± 1.0, 43.5 ± 0.5, 50.7 ± 1.5, and 55.0±3.0 days when the inoculum sizes were 0.1, 0.04, 0.01, and 0.007 CFU/g, respectively. From the same corresponding inoculum sizes, the TTDs at 8°C were 4.5 ± 0.5, 6.5 ± 0.5, 7.0 ± 1.0, and 8.5±0.5 days. Significant differences (P< 0.05) between TTDs were observed only when the inoculum sizes differed by at least 2 log. On a shelf life plot of ln(TTD) versus temperature, the Q10(increase in TTD for a 10°C increase in temperature) values ranged from 24.5 to 44.7 and with no significant influence from the inoculum densities. When the observed TTDs were compared with the expected detection times based on the data obtained from a study with an inoculum size of 10 to 20 CFU/g, significant deviations were noted at lower inoculum levels. These results can be valuable in designing a safety-based shelf life model for frankfurters and in performing quantitative risk assessment of listeriosis at low and practical contamination levels.

This research was conducted to study the growth of Listeria monocytogenesinoculated on frankfurters stored at different conditions as a basis for a safety-based consume by shelf life date label. Three L. monocytogenesstrains were separately inoculated at 10 to 20 CFU/cm2onto frankfurters that were previously formulated with or without high pressure and with or without added 2% potassium lactate (PL) and 0.2% sodium diacetate (SD). Inoculated frankfurters were air or vacuum packaged; stored at 4, 8, or 12°C; and L. monocytogenesand psychrotrophic plate counts were determined for 90, 60, and 45 days, respectively, or until the stationary phase was reached. The data (log CFU per square centimeter versus time) were fitted using the Baranyi-Roberts model to determine maximum growth rates and lag-phase time. The maximum growth rates and the lag time under each growth condition were used to calculate the time to reach 100-fold the initial Listeriapopulation. In frankfurters lacking PL and SD, the count of all strains increased by 2 log after 18 to 50 days at 4°C and 4 to 13 days at 8°C. The growth was inhibited at 4 and 8°C in frankfurters containing PL and SD, but one ribotype was capable of growing, with the time to reach 100-fold the initial Listeriapopulation ranging from 19 to 35 days at 12°C. In most cases, the time to reach 100-fold the initial Listeriapopulation of L. monocytogeneswas significantly longer in vacuum-packaged frankfurters as compared with air-packaged samples. Inclusion of PL and SD also inhibited the growth of psychrotrophs, but at all temperatures the psychrotrophic plate counts were greater than 4 log CFU/cm2at the end of the experiments. These results indicated that despite the use of antimicrobials, certain L. monocytogenesstrains could be capable of growing under storage-abuse conditions. Growth kinetics data could be useful for establishing a shelf life date label protocol under different handling scenarios.

The combined effect of heat and hydrogen peroxide (HP) on the inactivation of avirulent Bacillus anthracisspores (Sterne strain 7702; strain ANR-1, an avirulent Ames derivative lacking the pXO2 plasmid; and strain 9131, a plasmid-less Sterne strain) was evaluated in milk. The study temperature ranged from 90 to 95°C, and the concentration of added HP varied from 0.05 to 0.5%. Decimal reduction times (D-values) were determined using a sealed capillary tube technique. The mean D- and z-values of hydrated freeze-dried spores of all three strains in milk ranged from 550 s at 90C to180s at 94°C and from 8.6 to 9.0°C, respectively. When 0.05% HP was added to the milk, the D-values were decreased at least threefold, and at 0.5% HP the D-values ranged from 1 to 10 s. At 90°C, all three strains had similar D-values when 0.05% HP was added. Increasing the concentration of HP to 0.5% had a greater reducing effect on the D-value for strain 7702 than on the values for strains ANR-1 and 9131. The rate of inactivation of each strain followed first-order reaction kinetics at each temperature-peroxide combination. Equations in the form of D =Constant × (HP concentration)nhad R2values greater than 0.97 for strains ANR-1 and 7702 and of at least 0.7 for strain 9131. This study suggests that a combination of high temperature (from 90 to 95°C) and HP could be used for inactivation of B. anthracisspores in the event of deliberate contamination of milk such that the contaminated milk could be disposed of safely.

Samples were collected from 26 organic and conventional farms and 12 county fairs in Minnesota during 2001 and 2002 to identify the presence of Escherichia coliO157. Immunomagnetic separation was used for isolation of E. coliO157. Isolates were further characterized by the presence of virulence marker genes (stx1, stx2, eaeA, E-hly, katP, etpD,and espP), antimicrobial susceptibility profiles, and genotypes. During 2001, E. coliO157 was isolated from 16 (5.2%) of 305 fecal samples and from 7 (36.8%) of 19 farms. During 2002, E. coliO157 was isolated from 6 (4.5%) of 132 fecal samples from weaned calves at 4 (23.5%) of 17 farms. During 2001 and 2002, cattle manure samples were collected from 12 county fairs, and E. coliO157 was isolated from 19 (11%) of 178 samples and 9 (75%) of 12 county fairs. Among 40 E. coliO157 isolates, 17 isolates (43%) had both the stx1and stx2genes, and 21 strains (53%) had the stx2gene only. Thirteen percent of O157 isolates were resistant to tetracycline, and 25% were resistant to sulfadimethoxine. Heterogeneity of E. coliO157 strains was demonstrated by the presence of 22 different pulsed-field gel electrophoresis (PFGE) patterns. Four PFGE patterns matched those of isolates previously found in humans. The presence of E. coliO157 at county fairs suggests the potential for transmission to the public, who may have contact with cattle or their environment.

Microbiological analyses of fresh fruits and vegetables produced by organic and conventional farmers in Minnesota were conducted to determine the coliform count and the prevalence of Escherichia coli, Salmonella, and E. coliO157:H7. A total of 476 and 129 produce samples were collected from 32 organic and 8 conventional farms, respectively. The samples included tomatoes, leafy greens, lettuce, green peppers, cabbage, cucumbers, broccoli, strawberries, apples, and seven other types of produce. The numbers of fruits and vegetables was influenced by their availability at participating farms and varied from 11 strawberry samples to 108 tomato samples. Among the organic farms, eight were certified by accredited agencies and the rest reported the use of organic practices. All organic farms used aged or composted animal manure as fertilizer. The average coliform counts in both organic and conventional produce were 2.9 log most probable number per g. The percentages of E. coli–positive samples in conventional and organic produce were 1.6 and 9.7%, respectively. However, the E. coliprevalence in certified organic produce was 4.3%, a level not statistically different from that in conventional samples. Organic lettuce had the largest prevalence of E. coli(22.4%) compared with other produce types. Organic samples from farms that used manure or compost aged less than 12 months had a prevalence of E. coli19 times greater than that of farms that used older materials. Serotype O157:H7 was not detected in any produce samples, but Salmonellawas isolated from one organic lettuce and one organic green pepper. These results provide the first microbiological assessment of organic fruits and vegetables at the farm level.

A previously identified set of anti– Escherichia coliO157:H7 colicinogenic E. coliwere characterized to assess the suitability of these isolates as a preharvest food safety intervention in cattle. This collection of 23 E. colistrains were screened for virulence factors, antibiotic resistance, type of colicin(s) present, and their ability to inhibit other pathogenic E. coli. With the use of PCR, pathogen genes were detected in six of the 23 colicinogenic E. coli. When the nonpathogenic strains were assessed for antibiotic resistance, four strains showed resistance to at least one antibiotic. The remaining set of 14 strains were evaluated for the presence of previously identified colicins. Seven colicins (B, E1, E2/E7, E7, Ia/Ib, K, and M) were detected. One half of the strains possessed multiple types of colicins. The most commonly detected colicins were B, E2/E7, and M, which were found in six strains each. DNA sequencing was also performed in order to classify the E2/E7 colicins separately from E7 colicins. The 14 colicinogenic E. colialso were evaluated for their ability to inhibit 10 different non-O157 pathogenic E. coli. Six of the colicinogenic E. coliwere capable of inhibiting all 10 pathogens, and the remaining eight strains could each inhibit between six to eight of the pathogenic E. coli. This strain collection has great potential for inhibiting E. coliO157:H7 in cattle.

Escherichia colistrains were screened for their ability to inhibit E. coliO157:H7. An initial evaluation of 18 strains carrying previously characterized colicins determined that only colicin E7 inhibited all of the E. coliO157:H7 strains tested. A total of 540 strains that had recently been isolated from humans and nine different animal species (cats, cattle, chickens, deer, dogs, ducks, horses, pigs, and sheep) were tested by a flip-plating technique. Approximately 38% of these strains were found to inhibit noncolicinogenic E. coliK12 strains. The percentage of potentially colicinogenic E. coliper animal species ranged from 14% for horse isolates to 64% for sheep strains. Those isolates that inhibited E. coliK12 were screened against E. coliO157:H7, and 42 strains were found to be capable of inhibiting all 22 pathogenic strains tested. None of these 42 strains produced bacteriophages, and only 24 isolates inhibited serotype O157:H7 in liquid culture. The inhibitory activity of these strains was completely eliminated by treatment with proteinase K. When mixtures of these 24 colicinogenic strains were grown in anaerobic continuous culture, the four-strain E. coliO157:H7 population was reduced at a rate of 0.25 log10cells per ml per h, which was fivefold faster than the washout rate. Two strains originally isolated from cat feces (F16) and human feces (H30) were identified by repetitive sequences polymerase chain reaction as the predominant isolates in continuous cultures. The results of this work indicate that animal species other than cattle can be sources of anti-O157 colicinogenic strains, and these results also lead to the identification of at least two isolates that could potentially be used in preharvest control strategies.