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Consumption of Salmonella-contaminated jalapeño peppers has been implicated in one of the largest foodborne illness outbreaks in the summer of 2008. The objective of this study was to investigate representative groups of native microflora and the distribution, growth, and inactivation of experimentally-inoculated Salmonella Saintpaul on jalapeño peppers. Two genetically modified strains of Salm. Saintpaul producing either green- or red-fluorescent protein were constructed and used in the study. Microbiological analyses showed that jalapeño peppers contained an average of 5.6 log units of total aerobic count and 3.5, 1.8, and 1.9 log units, respectively, of enterobacteriaceae, lactic acid bacteria, and yeast/mold per gram of tissue. Strains typical of Pseudomonas accounted for 8.3% of total aerobic count, and 0.2% of which exhibited pectolytic activity. On inoculated peppers, a vast majority (>90%) of Salm. Saintpaul was recovered from stem/calyx and only a small proportion recovered from fleshy pods. Growth of Salm. Saintpaul on peppers was indicated by an increase in the population of 3 log units after incubation of samples at 20 degrees C for 48 h. Fluorescent Salm. Saintpaul aggregates could be readily detected on stem/calyx using stereofluorescence imaging microscopy and scanning electron microscopy. Data presented showed that Salm. Saintpaul could survive for at least 8 wk on peppers stored at 4 degrees C. Immersion of inoculated peppers in 200 ppm of sodium hypochlorite, acidified sodium chlorite, or peroxy acetic acid for 10 min could reduce the number of Salm. Saintpaul on stem/calyx by 1.5 to 1.7 and that on flesh by 2.1 to 2.4 log units. Practical Application: Consumption of Salmonella-contaminated jalapeño peppers has been implicated in foodborne illness outbreaks. The vast majority of Salmonella Saintpaul recovered from inoculated jalapeño peppers (>90%) was from stem/calyx. Salmonella increased by 3 log units during storage at 68 degrees F (20 degrees C) for 48 h. Salmonella could survive for at least 8 wk on peppers stored at 4 degrees C. Immersion of inoculated peppers in 200 ppm of sodium hypochlorite, acidified sodium chlorite, or peroxyacetic acid for 10 min reduced Salmonella on stem/calyx by 1.5 to 1.7 log units, compared with reductions of 2.1 to 2.4 log units on flesh. These results highlight the need to consider the stem/calyx as the most likely area for contamination of jalapeño peppers, and to process this commodity accordingly to minimize exposure and cross-contaminations.

The health and environmental hazard associated with the use of chlorine for food processing has been documented previously. This study was conducted to determine if acidified sodium chlorite (ASC) could be used to replace calcium hypochlorite (Ca[OCl](2)) for disinfection of alfalfa seeds. Contaminated seeds containing approximately 1.5 x 10(7) CFU/g of Salmonella were treated with ASC or Ca(OCl)(2) at different concentrations and for different periods of time. Results showed that the efficacy of ASC and Ca(OCl)(2) for elimination of Salmonella on contaminated seeds could be improved greatly by extending the treatment time from the traditional 15 to 45 min. Treatment of seeds with 800 ppm of ASC for 45 min reduced the number of Salmonella by 3.9 log units, approximately 1.2 log units higher than that treated with 20000 ppm of Ca(OCl)(2). Treatment of seeds with a lower concentration (100 to 400 ppm) of ASC for 45 min reduced the number of Salmonella by 1.3 to 2.2 log units. Soaking alfalfa seeds in 800 ppm of ASC for 45 min did not affect seed germination. However, soaking seeds in 20000 ppm of Ca(OCl)(2) for 45 min reduced seed germination by 20%. Unlike Ca(OCl)(2), antimicrobial efficiency of ASC was not affected by pre-exposure to alfalfa seeds. Data presented also showed that Salmonella on newly inoculated seeds that had been stored at 4 degrees C for less than 7 d were more sensitive to sanitizer treatment than those on seeds that had been stored for 4 wk or longer.

Aims: To investigate the growth of salmonellae on sprouting alfalfa seeds as affected by the inoculum size, microbial load and Pseudomonas fluorescens 2-79., Methods and Results: Alfalfa seeds pre-inoculated with < or =10(1)-10(3) CFU g(-1) of salmonellae and with or without Ps. fluorescens 2-79 were sprouted in glass jars and the population of salmonellae were determined daily for up to 6 days. The population of salmonellae on germinating seeds reached the maximum 2-3 days after sprouting when total bacterial count reached the maximum (10(9) CFU g(-1)). The population of salmonellae on sprouting seeds not treated with Ps. fluorescens 2-79 showed a net increase of 3-4 log units. However, the population of salmonellae on alfalfa seeds treated with Ps. fluorescens 2-79 showed a net increase of only 1-2 log units. Disinfection of seeds with calcium hypochlorite enhanced the growth of salmonellae., Conclusions: Treatment of seeds with Ps. fluorescens 2-79 reduced the growth of salmonellae by 2-3 log units., Significance and Impact of the Study: The potential of Ps. fluorescens 2-79 as a biological agent for use in control of salmonellae on sprouting seeds was demonstrated and warrants further investigation.

Aims: To investigate the resuscitation of acid-injured Salmonella enterica in selected enrichment broths, in apple juice and on cut surfaces of apple and cucumber slices., Methods and Results: Following exposure to 2.4% acetic acid for 7 min, S. enterica (serovars Mbandaka, Chester and Newport) cells were used to inoculate enrichment broths, phosphate-buffered saline (PBS), apple juice and fruit slices. Injured Salmonella cells resuscitated and regained the ability to form colonies on selective agar (Xylose-Lysine-Tergitol 4) if they were incubated in lactose broth (LB), universal pre-enrichment broth (UPB) or buffered peptone water (BPW), but not in tetrathionate broth, PBS or apple juice. The resuscitation occurred at a significantly (P > 0.05) faster rate in UPB than in LB or BPW. The resuscitation also occurred on the surfaces of fresh-cut cucumber at 20 degrees C, but not at 4 degrees C., Conclusions: Acid-injured Salmonella cells resuscitated in nonselective enrichment broths at different rates, but not in selective enrichment broth, apple juice, PBS or on fresh-cut apple., Significance and Impact of the Study: Pre-enrichment of food samples in UPB prior to selective enrichment is recommended. Injured Salmonella cells have the ability to resuscitate on fresh-cut surfaces of cucumber when stored at abusive temperatures.

Aims: To investigate and prevent the undesirable effect of native bacteria and alfalfa seed homogenates on detection of Salmonella in alfalfa seeds by indicator agar media and polymerase chain reaction (PCR)., Methods and Results: The relative sensitivity of five indicator agar media, including modified semisolid RV (MSRV), xylose-lysine-Tergitol 4 (XLT4), Hektoen enteric agar (HEA), brilliant green agar (BGA) and bismuth sulphite agar (BSA), for detection of Salmonella in the presence of a large number of native bacteria from alfalfa seeds was examined. The detection limit as measured by the ratio between the numbers of native bacteria and Salmonella was estimated to be 10(6) to 1 for MSRV and 10(3) to 1 for XLT4, HEA, BGA or BSA. Presence of alfalfa seed homogenates markedly reduced the sensitivity of Salmonella detection by PCR. The minimal number of Salmonella detectable by PCR was determined to be 1-10 and 100-1000 CFU in the absence and presence of seed homogenate, respectively. Application of anti-Salmonella immunomagnetic beads permitted detection of 2-5 CFU of heat-injured cells in 25 g of seeds within 24 h by PCR., Conclusions: The MSRV medium is more sensitive than other indicator agars for detecting a small number of motile Salmonella in samples containing a large number of native bacteria. Application of immunomagnetic beads eliminates the PCR-inhibitory activity of seed homogenates and improves the detection of Salmonella in inoculated seeds., Significance and Impact: The results generated from this study will aid the seed distributors, sprout growers and public health officials to identify and recall the Salmonella-contaminated seed lots to be used for sprout production.

The native microflora of three types of produce (green bell peppers, Romaine lettuce, and prepeeled baby carrots) and two types of sprouting seeds (alfalfa and clover) were investigated. Aerobic plate count (APC) for each produce or seed type as determined on Pseudomonas agar F (PAF) with incubation at 28 degrees C was in the range of 4 to 7 log CFU per g of tissue or seed. There was no significant difference (P > or = 0.05) in APC when the determinations were made with three agar media including PAF, brain heart infusion agar, and plate count agar. However, the APC as determined from plates that were incubated at 28 degrees C was significantly (P < or = 0.05) higher than with incubation at 37 degrees C. Fluorescent pseudomonads accounted for 23 to 73% of APC and 6 to 18% of APC recovered from carrots, pepper, and lettuce were pectolytic. Forty-eight strains of pectolytic bacteria were randomly isolated and identified, respectively, as members of the genera of Pseudomonas, Erwinia, Bacillus, Xanthomonas, or Flavobacterium. Lactic acid bacteria and/or yeast were consistently isolated from baby carrots, lettuce, and sprouting seeds (alfalfa or clover) but not from green bell peppers. Approximately 120 strains of indigenous microflora were tested for their ability to inhibit the growth of Salmonella Chester, Listeria monocytogenes, Escherichia coli, or Erwinia carotovora subsp. carotovora on PAF. Six isolates capable of inhibiting the growth of at least one pathogen were isolated and identified, respectively, as Bacillus spp. (three strains), Pseudomonas aeruginosa (one strain), Pseudomonas fluorescens (strain A3), and yeast (strain D1). When green pepper disks were inoculated with strains A3 and D1, the growth of Salmonella Chester and L. monocytogenes on the disks was reduced by 1 and 2 logs, respectively, over a period of 3 days. Application of strains A3 and D1 as potential biopreservatives for enhancing the quality and safety of fresh produce is discussed.

A laboratory model using green pepper disks was developed to investigate the attachment of Salmonella Chester on plant tissue and to evaluate the effectiveness of sanitizer agents in inactivating attached bacteria on fruits. Pepper disks (14 mm in diam, and 3-4 mm in thickness) were immersed in a bacterial suspension containing 1.5 x 107 cfu x mL(-1) of S. Chester for 30 s and subsequently air-dried at room temperature for 10 min. Approximately 30% of the bacteria retained on the disk after immersion were firmly attached and could not be removed by two washes and agitation. A positive correlation was observed between the number of bacteria attached and the concentration of bacteria in the suspension. Population studies and scanning electron microscopic examinations revealed that attachment of S. Chester on pepper disks occurred mainly on the surfaces of injured (cut) tissue but rarely on the unbroken skin. When inoculated disks were treated with 3% to 12% (w/v) of trisodium phosphate (TSP) at pH 12.3 for 5 min, the population of bacteria on the disk was reduced by 10- to 100-fold. A small portion (0.7% to 7.1%) of bacteria attached to the disk were either resistant to or protected from the TSP treatment. When the pH of TSP solution was reduced from 12.3 to 4.5, the effectiveness of TSP in inactivating S. Chester on pepper disks was reduced by 26%. This study shows that surfaces of injured fruit tissue are the principal sites for bacterial attachment, and a small portion of the bacteria attached to the tissue are resistant to the sanitizer treatment. Avoiding mechanical injuries to fresh fruits during and after harvest would reduce the chance of pathogen attachment and contamination on green pepper and fruits of similar nature.

Attachment and growth of Salmonella Chester on fresh-cut apple disks and in vivo response of attached bacteria to sanitizer treatments were investigated. Apple disks (14 mm in diameter and 3 to 4 mm in thickness) were immersed in a bacterial suspension that contained 8.17 log CFU/ml of Salmonella Chester and air dried at room temperature for 10 min. After two rinses, the population of Salmonella Chester retained on apple disks that contained no skin was 13 to 19% higher than that retained on disks that contained skin, indicating that Salmonella Chester attached more firmly to the surfaces of injured tissue than to the unbroken skin. The number of bacteria attached to the disk was not affected by the immersion time but was directly proportional to the concentration of bacteria in the suspension. The distribution of artificially inoculated Salmonella Chester on the surfaces of three different parts of whole fruit was determined; 94% of attached bacteria was found on the stem and calyx cavity areas and 6% on the skin of the remaining area of the fruit. Despite their acidic pH (4.1), apple disks supported the growth of Salmonella Chester at 20 degrees C but not at 8 degrees C. All four sanitizers tested in the study, including 6% hydrogen peroxide, 2% trisodium phosphate, 0.36% calcium hypochlorite, and 1.76% sodium hypochlorite, were effective in reducing the population of Salmonella Chester on apple disks by 1 to 2 logs. However, 5 to 13% of bacteria survived the sanitizer treatments. Hydrogen peroxide, which reduced the population of Salmonella Chester on skin by 3 to 4 logs and the population of bacteria on stem or calyx by 1 to 2 logs, was the most effective among the four sanitizers tested. Firm attachment of bacteria on calyx, stem, and injured tissue and partial resistance of attached bacteria to sanitizer treatments are two major obstacles to be considered when developing methods for cleaning and decontaminating apple fruits destined for juice production and fresh consumption.

The article presents a study of antimicrobial action of acetic acid (AA) on Salmonella mbandaka in vitro and on cut surfaces of apple slices based on acid concentration, exposure time and culture age. Findings reveal that the most effective sanitizer treatments tested was a mixture of AA and hydrogen peroxide (H2O2). Salmonella strains that are more sensitive to AA action in vitro are also found to be more susceptible to AA action when present on apple slices.