A new platform for Real-Time PCR detection of Salmonella spp., Listeria monocytogenes and Escherichia coli O157 in milk

Intoxications and infections caused by food-borne pathogens represent an increasing public health problem, and diagnostic tests in multiplex format are needed for the rapid identification of food contaminations caused by more than one microbial species. We have developed a multiple PCR-based platform for the simultaneous detection of the widespread milk-associated pathogens Salmonella spp., Listeria monocytogenes and Escherichia coli O157. The assay combines an enrichment step in a medium properly formulated for the simultaneous growth of target pathogens, a DNA isolation method, and a multiplex Real-Time PCR detection system based either on dual-labelled probes (mRT-PCR), or on melting curve analysis (mHRM). The second, producing a distinct peak for each amplification product, allows the qualitative assessment of pathogen presence. Moreover, the internal amplification control (IAC) included in the reaction, ensuring the reliability of results, complies with quality management programmes. Inclusivity and exclusivity were 100% each, with a detection limit of 1 CFU for each pathogen in a total of five 25 ml-aliquots of raw milk, and a duration of two working days.The assay represents an alternative approach for the qualitative detection of the cited bacterial species, suitable for a relatively inexpensive screening of several milk samples, reducing the turnaround time and the workload.     

肾脏内科住院患者感染病原菌分布及耐药性分析

目的:调查肾脏内科住院患者感染病原菌分布和耐药性,为临床治疗细菌感染的抗菌药物选择提供依据。方法:收集2016年1月至2019年6月期间入住皖南医学院弋矶山医院肾脏内科患者送检标本培养阳性病原菌数据。结果:共培养出286株细菌,以呼吸道感染和泌尿道感染为主。革兰阴性菌占比89.51%,革兰阳性菌占比10.49%。革兰阴性菌中,肠杆菌科细菌检出前三位细菌分别是大肠埃希菌、肺炎克雷伯菌和阴沟肠杆菌,非发酵阴性菌检出前二位细菌分别是铜绿假单胞菌、鲍曼不动杆菌;其中产超广谱β内酰胺酶细菌（ESBLs）检出率为32.87%,以大肠埃希菌和肺炎克雷伯菌为主,对氨曲南和头孢曲松耐药率分别为83.8%和100%;碳青霉烯类耐药菌检出率为6.29%。在革兰阳性菌中金黄色葡萄球菌最常见,其中耐甲氧西林金黄色葡萄球菌（MRSA）检出率为4.89%。结论:在本院肾脏内科住院患者细菌感染中,产ESBLs大肠埃希菌和肺炎克雷伯菌检出率高。     

软腐猕猴桃果实真菌类病原菌的生物多样性分析

本文对湖南地区主要猕猴桃产区猕猴桃软腐果实真菌类病害的多样性分布规律进行系统分析,为猕猴桃软腐病原真菌多样性的深入研究提供基础数据和理论依据。采用IlluminaHi Seq高通量测序技术对5处软腐猕猴桃(A、B、C、D、E),15组果实样品中微生物ITS扩增子二代测序数据采集,进行生物信息学分析。5处软腐猕猴桃果实中真菌种类较为丰富,真菌丰富度指数和多样性指数较高,尤其是A、B、C处,相对于D、E处存在极显著差异(p<0.01)。在不同猕猴桃果实中得到了2个门的优势微生物。子囊菌门(Ascomycota)为最优势菌门,主要分布在D和E处,群类相对丰度为19.20%,且远远大于次优势菌门-担子菌门(Basidiomycota,6.80%,主要分布在A、B、C处),表现出非常明显的优势;间座壳属Diaporthe相对丰度最高,达到了14.00%,相对于其它属病原菌均具有极显著性差异(p<0.01),此菌在D、E组含量相对较高,在本组内达到了30%以上,其余3组几乎没有。布勒弹孢酵母属Bullera相对丰度为4.90%,此菌为各组共有的优势菌属,A、B、C组略高,在本组内达到了5.50%左右,D、E组则为3.50%左右。葡萄座腔菌属Neofusicoccum相对丰度为3.20%,此菌在E组含量相对较高,在本组内达到了15.00%以上,其余4组几乎没有。红菇属Russula相对丰度为0.60%,及其它较少的担子菌门菌属Tomentella、Phialocephala、oidiodendron、cenococcum等,此外,具有软腐猕猴桃果实潜力的菌属有茎点酶菌属(homa)、青霉属(penicillium)及拟青霉属(simplicillium)等。实验为猕猴桃软腐病的防治提供了一定理论基础,今后对猕猴桃果实软腐病防治时可针对性的杀灭间座壳属和葡萄座腔菌属微生物,以提高果实的耐贮性。     

Evaluation of Antibacterial Activity of Whey Protein Isolate Coating Incorporated with Nisin,Grape Seed Extract,Malic Acid,and EDTA on a Turkey Frankfurter System

ABSTRACT: The effectiveness of whey protein isolate (WPI) coatings incorporated with grape seed extract (GSE), nisin (N), malic acid (MA), and ethylenediamine tetraacetic acid (EDTA) and their combinations to inhibit the growth of Listeria monocytogenes, E. coli O157:H7, and Salmonella typhimurium were evaluated in a turkey frankfurter system through surface inoculation (approximately 10⁶ CFU/g) of pathogens. The inoculated frankfurters were dipped into WPI film forming solutions both with and without the addition of antimicrobial agents (GSE, MA, or N and EDTA, or combinations). Samples were stored at 4 °C for 28 d. The L. monocytogenes population (5.5 log/g) decreased to 2.3 log/g after 28 d at 4 °C in the samples containing nisin (6000 IU/g) combined with GSE (0.5%) and MA (1.0%). The S. typhimurium population (6.0 log/g) was decreased to approximately 1 log cycles after 28 d at 4 °C in the samples coated with WPI containing a combination of N, MA, GSE, and EDTA. The E. coli O157:H7 population (6.15 log/g) was decreased by 4.6 log cycles after 28 d in samples containing WPI coating incorporated with N, MA, and EDTA. These findings demonstrated that the use of an edible film coating containing nisin, organic acids, and natural extracts is a promising means of controlling the growth and recontamination of L. monocytogenes, S. typhimurium, and E. coli O157:H7 in ready‐to‐eat poultry products.     

Effect of UV-C irradiation on the inactivation of inoculated pathogens and quality of chicken breasts during storage

In this study, we evaluated the inactivation of foodborne pathogens inoculated on chicken breasts by UV-C treatment. Chicken breasts were inoculated with Campylobacter jejuni, Listeria monocytogenes, and Salmonella enterica serovar Typhimurium at 6–7 log CFU/g. The inoculated chicken breasts were then irradiated with UV-C light of dose 0, 0.5, 1, 3, and 5 kJ/m². Microbiological data indicated that the populations of the foodborne pathogens decreased significantly (p < 0.05) with increasing UV-C irradiation. In particular, UV-C irradiation at 5 kJ/m² reduced the initial populations of C. jejuni, L. monocytogenes, and S. typhimurium by 1.26, 1.29, and 1.19 log CFU/g, respectively. After UV-C irradiation, the samples were individually packed using polyethylene terephthalate containers and stored at 4 ± 1 °C for 6 d. The pH of the control increased more than the samples irradiated at 0.5, 1, 3, and 5 kJ/m², while TBARS values increased during storage regardless of UV-C irradiation. UV-C irradiation caused negligible changes in Hunter L, a, and b values. These results suggest that UV-C irradiation can be useful in improving the microbial safety of chicken breasts during storage, without impairing quality.     

Effects of acid adaptation on the survival of Listeria monocytogenes on modified atmosphere packaged vegetables

The survival and growth of Listeria monocytogenes and L. innocua on ready-to-use (RTU) packaged vegetables (lettuce, swedes, dry coleslaw mix, bean-sprouts) were studied. The effects of acid adaptation of Listeria spp. on their survival during subsequent storage were also investigated. Listeria innocua behaviour was similar to that of L. monocytogenes on all vegetables examined. The survival and growth patterns of Listeria varied with the packaged product. Populations on packaged lettuce and swedes significantly increased ( P  < 0.05, by 1–1.5 log cycles) during a 14-day storage period. During the same period, Listeria counts gradually decreased (by 1–1.5 log cycles) on coleslaw mix. Acid adaptation enhanced survival of Listeria spp. during storage in packages of vegetables which had relatively high in-pack CO₂ levels (i.e. 25% in packaged coleslaw, bean-sprouts). It is concluded that adapting listerial cells to mildly acidic conditions rendered cultures more resistant to relatively high (25–30%) CO₂ atmospheres.     

Assessment of fermentation as a household technology for improving food safety: a joint FAO/WHO workshop

Food safety and nutritional aspects of lactic acid fermentation processes for the purpose of complementary food preparation at household scale in tropical countries were assessed during an FAO/WHO workshop held in Pretoria, South Africa in December 1995.

Lactic acid fermentation was evaluated in particular, taking into consideration that microbial fermentation is only a part of the food preparation processes involving other operations such as soaking, cooking and the use of germinated cereals. The latter are of special interest as they enable the preparation of semiliquid porridges of high-nutrient density.

The present state of knowledge concerning the antimicrobial effects imparted by the acidity in lactic-fermented foods was reviewed, as well as the nutritional benefits of fermentation and use of germinated cereals. Areas requiring further research were identified and prioritized.

It was concluded that high priority should be given to research on: the effect of lactic acid fermentation on viruses, parasites, some bacteria and mycotoxins; risk assessment using the HACCP approach, health education of food handlers and consumer perception of fermented foods; characterization and optimization of fermentation processes and development of appropriate starters; and some physiological and nutritional effects of consumption of fermented foods.     

Relationships between Bacteroides 16S rRNA genetic markers and presence of bacterial enteric pathogens and conventional fecal indicators

Occurrence and prevalence of different bacterial enteric pathogens as well as their relationships with conventional (total and fecal coliforms) and alternative fecal indicators (host-specific Bacteroides 16S rRNA genetic markers) were investigated for various water samples taken from different sites with different degrees of fecal contamination. The results showed that a wide range of bacterial pathogens could be detected in both municipal wastewater treatment plant samples and in surface water samples. Logistic regression analysis revealed that total and human-specific Bacteroides 16S rRNA genetic markers showed significant predictive values for the presence of Escheriachia coli O-157, Salmonella, heat-labile enterotoxin (LT) of enterotoxigenic E. coli (ETEC), and heat-stable enterotoxin for human (STh) of ETEC. The probability of occurrence of these pathogenic bacteria became significantly high when the concentrations of human-specific and total Bacteroides 16S rRNA genetic markers exceeded 10(3) and 10(4) copies/100 mL. In contrast, Clostridium perfringens was detected at high frequency regardless of sampling sites and levels of Bacteroides 16S rRNA genetic markers. No genes related to Shigella spp., Staphylococcus aureus and Vibrio cholerae were detected in any samples analyzed in this study. Conventional indicator microorganisms had low levels of correlation with the presence of pathogens as compared with the alternative fecal indicators. These results suggested that real-time PCR-based measurement of alternative Bacteroides 16S rRNA genetic markers was a rapid and sensitive tool to identify host-specific fecal pollution and probably associated bacterial pathogens. However, since one fecal indicator might not represent the relative abundance of all pathogenic bacteria, viruses and protozoa, combined application of alternative indicators with conventional ones could provide more comprehensive pictures of fecal contamination, its source and association with pathogenic microorganisms.     

肠出血性大肠杆菌O157∶H7的检测方法进展

肠出血性大肠杆菌(EHEC)O157:H7为新近报道的食源性强致病菌,对病原菌快速、简便、准确、灵敏的检测方法是预防控制的关键,本文对国内外常用的鉴别培养、免疫检测以及PCR等检测方法进行了系统的综述。