Mangieri Nicola, Picozzi Claudia, Cocuzzi Riccardo, Foschino Roberto
Department of Food, Environmental and Nutritional Sciences, Università degli Studi di Milano, Milan, Italy.
Front Microbiol. 2020 Jul 24;11:1801. doi: 10.3389/fmicb.2020.01801. eCollection 2020.
Shiga-toxin producing (STEC) are important foodborne pathogens involved in gastrointestinal diseases. Furthermore, the recurrent use of antibiotics to treat different bacterial infections in animals has increased the spread of antibiotic-resistant bacteria, including , in foods of animal origin. The use of bacteriophages for the control of these microorganisms is therefore regarded as a valid alternative, especially considering the numerous advantages (high specificity, self-replicating, self-limiting, harmless to humans, animals, and plants). This study aimed to isolate bacteriophages active on STEC strains and to set up a suspension of viral particles that can be potentially used to control STEC food contamination. Thirty-one STEC of different serogroups (O26; O157; O111; O113; O145; O23, O76, O86, O91, O103, O104, O121, O128, and O139) were investigated for their antibiotic resistance profile and sensitivity to phage attack. Ten percent of strains exhibited a high multi-resistance profile, whereas ampicillin was the most effective antibiotic by inhibiting 65% of tested bacteria. On the other side, a total of 20 phages were isolated from feces, sewage, and bedding material of cattle. The viral particles proved not to carry genes codifying Shiga-toxins and intimin. No STEC was resistant to all phages, although some strains revealed weak sensitivity by forming turbid plaques. Three different bacteriophages (forming the "cocktail") were selected considering their different RAPD (Random Amplification of Polymorphic DNA) profiles and the absence of virulence-encoding genes and antibiotic-resistance genes. The lytic ability against STEC strains was investigated at different multiplicity of infection (MOI = 0.1, 1, and 10). Significant differences ( < 0.05) among mean values of optical density were observed by comparing results of experiments at different MOI and controls. An effective reduction of bacterial population was obtained in 81% of cases, with top performance when the highest MOI was applied. The efficacy of the phage cocktail was tested on fresh cucumbers. Results showed a reduction in pathogenic by 1.97-2.01 log CFU/g at 25°C and by 1.16-2.01 log CFU/g at 4°C during 24 h, suggesting that the formulated cocktail could have the potential to be used in bio controlling STEC different serogroups.
产志贺毒素大肠杆菌（STEC）是引起胃肠道疾病的重要食源性病原体。此外，在动物中反复使用抗生素治疗不同的细菌感染增加了包括动物源食品中耐药菌的传播。因此，使用噬菌体来控制这些微生物被认为是一种有效的替代方法，特别是考虑到其众多优点（高特异性、自我复制、自我限制、对人类、动物和植物无害）。本研究旨在分离对STEC菌株有活性的噬菌体，并建立一种病毒颗粒悬浮液，其可潜在用于控制STEC食品污染。对31株不同血清型（O26；O157；O111；O113；O145；O23、O76、O86、O91、O103、O104、O121、O128和O139）的STEC菌株进行了抗生素耐药性分析和噬菌体攻击敏感性研究。10%的菌株表现出高度多重耐药性，而氨苄青霉素是最有效的抗生素，可抑制65%的受试细菌。另一方面，从牛的粪便、污水和垫料中总共分离出20种噬菌体。这些病毒颗粒被证明不携带编码志贺毒素和紧密黏附素的基因。没有STEC菌株对所有噬菌体都耐药，尽管一些菌株通过形成浑浊噬菌斑显示出较弱的敏感性。考虑到它们不同的随机扩增多态性DNA（RAPD）图谱以及不存在毒力编码基因和抗生素耐药基因，选择了三种不同的噬菌体（组成“鸡尾酒”）。在不同感染复数（MOI = 0.1、1和10）下研究了对STEC菌株的裂解能力。通过比较不同MOI实验结果和对照，观察到光密度平均值之间存在显著差异（P < 0.05）。在81%的情况下获得了细菌数量的有效减少，当应用最高MOI时效果最佳。在新鲜黄瓜上测试了噬菌体鸡尾酒的功效。结果表明，在25℃下24小时内，致病菌减少了1.97 - 2.01 log CFU/g，在4℃下减少了1.16 - 2.01 log CFU/g，这表明配制的鸡尾酒有可能用于生物控制不同血清型的STEC。
