摘要

       生产过程中会发生长叶莴苣被人类病原体，抗生素抗性细菌（ARB）和抗生素抗性基因（ARG）污染的情况。采后采取干预措施以减轻病原体，但也可以减轻蔬菜上的ARB和ARG。这项研究的目的是确定用消毒剂清洗，伽马射线辐照和冷藏后生菜叶微生物区群，接种的ARB以及电阻组（ARG的概况）的变化。为了模拟收获前污染的潜在来源，将长叶莴苣叶用含有致病性抗生素菌株（大肠杆菌O157：H7）和腐败细菌（铜绿假单胞菌）代表的堆肥浆料接种。比较了用次氯酸钠（50 ppm游离氯）洗涤，在改良的气氛（98％氮气）下包装，辐照（1.0 kGy）并在4°C下保存1天和14天的各种组合。 shot弹枪宏基因组测序分析了收获后处理对抵抗力的影响。进行细菌16S rRNA基因扩增子测序，以确定对叶球菌群的变化。通过选择培养基上的计数评估接种的ARB的存活率和再生长。在含消毒剂的水中清洗生菜与降低ARG类的含量有关，这些ARG类赋予了对糖肽，β-内酰胺类，酚类和磺酰胺类的抗性（Wilcoxon，p <0.05）。洗涤后再进行辐照导致产生不同的抗药性，这主要是由于多药，三氯生，多粘菌素，β-内酰胺和喹诺酮ARG类的减少（Wilcoxon，p <0.05）。辐照，然后在4°C下保存14天，与治疗后1天相比，导致ARGs宿主细菌的β多样性发生明显变化（ANOSIM，R = 0.331； p = 0.003）。将经过清洗和辐射的生菜在4°C下保存14天会增加假单胞菌科和食肉杆菌科的相对丰度（Wilcoxon，p <0.05），这两组与生菜叶球体中10种ARG类的检测相关（p <0.05）。辐照导致大肠杆菌O157：H7和铜绿假单胞菌的接种菌株显着降低（〜3.5 log CFU / g）（ANOVA，p <0.05）。结果表明，在4°C的条件下，将经过包装的生菜的洗涤，辐照和保存是降低耐药性的大肠杆菌O157：H7和铜绿假单胞菌以及各种ARG类型相对丰度的有效策略。

        Contamination of romaine lettuce with human pathogens, antibiotic-resistant bacteria (ARB), and antibiotic resistance genes (ARGs) occurs during production. Post-harvest interventions are emplaced to mitigate pathogens, but could also mitigate ARB and ARGs on vegetables. The objective of this research was to determine changes to lettuce phyllosphere microbiota, inoculated ARB, and the resistome (profile of ARGs) following washing with a sanitizer, gamma irradiation, and cold storage. To simulate potential sources of pre-harvest contamination, romaine lettuce leaves were inoculated with compost slurry containing antibiotic-resistant strains of pathogenic (Escherichia coli O157:H7) and representative of spoilage bacteria (Pseudomonas aeruginosa). Various combinations of washing with sodium hypochlorite (50 ppm free chlorine), packaging under modified atmosphere (98% nitrogen), irradiating (1.0 kGy) and storing at 4°C for 1 day versus 14 days were compared. Effects of post-harvest treatments on the resistome were profiled by shotgun metagenomic sequencing. Bacterial 16S rRNA gene amplicon sequencing was performed to determine changes to the phyllosphere microbiota. Survival and regrowth of inoculated ARB were evaluated by enumeration on selective media. Washing lettuce in water containing sanitizer was associated with reduced abundance of ARG classes that confer resistance to glycopeptides, β-lactams, phenicols, and sulfonamides (Wilcoxon, p < 0.05). Washing followed by irradiation resulted in a different resistome chiefly due to reductions in multidrug, triclosan, polymyxin, β-lactam, and quinolone ARG classes (Wilcoxon, p < 0.05). Irradiation followed by storage at 4°C for 14 days led to distinct changes to the β-diversity of the host bacteria of ARGs compared to 1 day after treatment (ANOSIM, R = 0.331; p = 0.003). Storage of washed and irradiated lettuce at 4°C for 14 days increased the relative abundance of Pseudomonadaceae and Carnobacteriaceae (Wilcoxon, p < 0.05), two groups whose presence correlated with detection of 10 ARG classes on the lettuce phyllosphere (p < 0.05). Irradiation resulted in a significant reduction (∼3.5 log CFU/g) of inoculated strains of E. coli O157:H7 and P. aeruginosa (ANOVA, p < 0.05). Results indicate that washing, irradiation and storage of modified atmosphere packaged lettuce at 4°C are effective strategies to reduce antibiotic-resistant E. coli O157:H7 and P. aeruginosa and relative abundance of various ARG classes.