The fresh-cut produce industry has been the fastest-growing portion of the food retail market during the past 10 years, providing consumers with convenient and nutritious food. However, fresh-cut fruits and vegetables raise food safety concerns, because exposed tissue may be colonized more easily by pathogenic bacteria than intact produce. This is due to the higher availability of nutrients on cut surfaces and the greater potential for contamination because of the increased amount of handling. We found that applied Listeria monocytogenes populations survived and increased only slightly on fresh-cut Red Delicious apples stored at 10°C but increased significantly on fresh-cut honeydew melons stored at 10°C over 7 days. In addition, we examined the effect of lytic, L. monocytogenes-specific phages via two phage application methods, spraying and pipetting, on L. monocytogenes populations in artificially contaminated fresh-cut melons and apples. The phage mixture reduced L. monocytogenes populations by 2.0 to 4.6 log units over the control on honeydew melons. On apples, the reduction was below 0.4 log units. In combination with nisin (a bacteriocin), the phage mixture reduced L. monocytogenes populations by up to 5.7 log units on honeydew melon slices and by up to 2.3 log units on apple slices compared to the control. Nisin alone reduced L. monocytogenes populations by up to 3.2 log units on honeydew melon slices and by up to 2.0 log units on apple slices compared to the control. The phage titer was stable on melon slices, but declined rapidly on apple slices. The spray application of the phage and phage plus nisin reduced the bacterial numbers at least as much as the pipette application. The effectiveness of the phage treatment also depended on the initial concentration of L. monocytogenes.

The preparation and distribution of fresh-cut produce is a rapidly developing industry that provides the consumer with convenient and nutritious food. However, fresh-cut fruits and vegetables may represent an increased food safety concern because of the absence or damage of peel and rind, which normally help reduce colonization of uncut produce with pathogenic bacteria. In this study, we found that Salmonella Enteritidis populations can (i) survive on fresh-cut melons and apples stored at 5 degrees C, (ii) increase up to 2 log units on fresh-cut fruits stored at 10 degrees C, and (iii) increase up to 5 log units at 20 degrees C during a storage period of 168 h. In addition, we examined the effect of lytic, Salmonella-specific phages on reducing Salmonella numbers in experimentally contaminated fresh-cut melons and apples stored at various temperatures. We found that the phage mixture reduced Salmonella populations by approximately 3.5 logs on honeydew melon slices stored at 5 and 10 degrees C and by approximately 2.5 logs on slices stored at 20 degrees C, which is greater than the maximal amount achieved using chemical sanitizers. However, the phages did not significantly reduce Salmonella populations on the apple slices at any of the three temperatures. The titer of the phage preparation remained relatively stable on melon slices, whereas on apple slices the titer decreased to nondetectable levels in 48 h at all temperatures tested. Inactivation of phages, possibly by the acidic pH of apple slices (pH 4.2 versus pH 5.8 for melon slices), may have contributed to their inability to reduce Salmonella contamination in the apple slices. Higher phage concentrations and/or the use of low-pH-tolerant phage mutants may be required to increase the efficacy of the phage treatment in reducing Salmonella contamination of fresh-cut produce with a low pH.

The effect of cultivation practices on fruit quality and antioxidant capacity in highbush blueberries var. Bluecrop (Vaccinium corymbosum L.) was evaluated from random samples of commercial late harvest fields in New Jersey. Results from this study showed that blueberry fruit grown from organic culture yielded significantly higher sugars (fructose and glucose), malic acid, total phenolics, total anthocyanins, and antioxidant activity (ORAC) than fruit from the conventional culture. In organically cultured fruit, the average values for the ORAC, total anthocyanins, and total phenolic content were 46.14 micromol of Trolox (TE)/g of fresh weight (fwt), 131.2 mg/100 g of fwt, and 319.3 mg/100 g of fwt, respectively. In conventionally cultured fruit, the average values for the ORAC, total anthocyanin, and total phenol content were 30.8 micromol of TE/g of fwt, 82.4 mg/100 g of fwt, and 190.3 mg/100 g of fwt, respectively. The organic culture also produced fruit with higher contents of myricetin 3-arabinoside, quercetin 3-glucoside, delphinidin 3-galactoside, delphinidin 3-glucoside, delphinidin 3-arabinoside, petunidin 3-galactoside, petunidin 3-glucoside, and malvidin 3-arabinoside than conventional culture. There was a significant correlation between the ORAC values and total phenolics and total anthocyanins. These results indicate that even though there were variations in phytonutrient content among individual farms within each cultural system, significant differences between two cultivation practices were evident.

No abstract

ABSTRACTStaphylococci cause bovine mastitis, withStaphylococcus aureusbeing responsible for the majority of the mastitis-based losses to the dairy industry (up to $2 billion/annum). Treatment is primarily with antibiotics, which are often ineffective and potentially contribute to resistance development. Bacteriophage endolysins (peptidoglycan hydrolases) present a promising source of alternative antimicrobials. Here we evaluated two fusion proteins consisting of the streptococcal λSA2 endolysin endopeptidase domain fused to staphylococcal cell wall binding domains from either lysostaphin (λSA2-E-Lyso-SH3b) or the staphylococcal phage K endolysin, LysK (λSA2-E-LysK-SH3b). We demonstrate killing of 16 differentS. aureusmastitis isolates, including penicillin-resistant strains, by both constructs. At 100 μg/ml in processed cow milk, λSA2-E-Lyso-SH3b and λSA2-E-LysK-SH3b reduced theS. aureusbacterial load by 3 and 1 log units within 3 h, respectively, compared to a buffer control. In contrast to λSA2-E-Lyso-SH3b, however, λSA2-E-LysK-SH3b permitted regrowth of the pathogen after 1 h. In a mouse model of mastitis, infusion of 25 μg of λSA2-E-Lyso-SH3b or λSA2-E-LysK-SH3b into mammary glands reducedS. aureusCFU by 0.63 or 0.81 log units, compared to >2 log for lysostaphin. Both chimeras were synergistic with lysostaphin againstS. aureusin plate lysis checkerboard assays. When tested in combination in mice, λSA2-E-LysK-SH3b and lysostaphin (12.5 μg each/gland) caused a 3.36-log decrease in CFU. Furthermore, most protein treatments reduced gland wet weights and intramammary tumor necrosis factor alpha (TNF-α) concentrations, which serve as indicators of inflammation. Overall, our animal model results demonstrate the potential of fusion peptidoglycan hydrolases as antimicrobials for the treatment ofS. aureus-induced mastitis.

A phage cocktail was applied to honeydew melon pieces 1, 0.5, and 0 h before contamination with Listeria monocytogenes strain LCDC 81-861 and 0.5, 1, 2, and 4 h after contamination. The phage application was most effective when applied 1, 0.5, or 0 h before contamination with L. monocytogenes, reducing pathogen populations by up to 6.8 log units after 7 days of storage. This indicates that under commercial conditions, if contamination occurs at the time of cutting, phage would have to be applied as soon as possible after cutting the produce. However, all phage applications from 1 h before to 4 h after contamination and all phage concentrations ranging from 10(4) to 10(8) PFU/ml reduced bacterial populations on honeydew melon pieces. Higher phage concentrations were more effective in reducing pathogen populations. A phage concentration of approximately 10(8) PFU/ml was necessary to reduce the pathogen populations to nondetectable levels immediately after treatment, and pathogen growth was suppressed by phage concentrations of 10(6) through 10(8) throughout the storage period of 7 days at 10 degrees C. In an attempt to enhance the effectiveness of the phage cocktail on low pH fruit, such as apples, the phage was applied in combination with MnCl2. This combination, however, did not enhance the effectiveness of the phage on apple tissue. The results from this study indicate that the effectiveness of the phage application on honeydew melon pieces can be optimized by using a phage concentration of at least 10(8) PFU/ml applied up to 1 h after processing of the honeydew melons.

Multi-drug resistant bacteria are a persistent problem in modern health care, food safety and animal health. There is a need for new antimicrobials to replace over used conventional antibiotics. Here we describe engineered triple-acting staphylolytic peptidoglycan hydrolases wherein three unique antimicrobial activities from two parental proteins are combined into a single fusion protein. This effectively reduces the incidence of resistant strain development. The fusion protein reduced colonization by Staphylococcus aureus in a rat nasal colonization model, surpassing the efficacy of either parental protein. Modification of a triple-acting lytic construct with a protein transduction domain significantly enhanced both biofilm eradication and the ability to kill intracellular S. aureus as demonstrated in cultured mammary epithelial cells and in a mouse model of staphylococcal mastitis. Interestingly, the protein transduction domain was not necessary for reducing the intracellular pathogens in cultured osteoblasts or in two mouse models of osteomyelitis, highlighting the vagaries of exactly how protein transduction domains facilitate protein uptake. Bacterial cell wall degrading enzyme antimicrobials can be engineered to enhance their value as potent therapeutics.

No abstract