Department of Medicine and Infectious Diseases, Faculty of Veterinary Medicine, Cairo University, Cairo, Egypt.
Laboratory of Molecular Epidemiology (LME), Faculty of Veterinary Medicine, Cairo University, Cairo, Egypt.
Trop Anim Health Prod. 2020 Nov;52(6):3173-3185. doi: 10.1007/s11250-020-02343-1. Epub 2020 Jul 10.
Escherichia coli field isolates from calves were characterized and categorized into the most significant diarrheagenic pathotypes using polymerase chain reaction (PCR) assays with different specific primers. The used PCR systems were designed to detect sequences representing the group-specific virulence genes encoding fimbriae f5 (K99), Shiga toxins (stx and stx), heat-stable enterotoxins (st), heat-labile enterotoxins (lt), intimin (eae), hemolysin (hylA), and EAEC heat-stable enterotoxin (astA). In the present work, a total of 150 E. coli field isolates were recovered from 150 fecal swabs collected from 100 diarrheic and 50 apparently healthy in-contact cattle and buffalo calves under 3 months old. Out of these 150 isolated E. coli, 106 isolates from 77 diarrheic and 29 in-contact calves harbored one or more of the investigated virulence genes. The pathotyping of the isolates could classify them into shigatoxigenic E. coli (STEC), enteropathogenic E. coli (EPEC), enterotoxigenic E. coli (ETEC), and enteroaggregative E. coli (EAEC) with a 30.7, 2.7, 12.7, and 7.3% distribution, respectively. Meanwhile, the detection rates of f5, stx, stx, st, lt, eae, hylA, and astA genes were 17.3, 27.3, 6.7, 10, 37.3, 17.7, 9.3, and 20.7%, respectively. These virulence genes were found either single or in different combinations, such as stx/eae, stx/st/f5, eae/st/f5, or st/lt/f5. Four attaching-effacing shigatoxigenic E. coli isolates (AE-STEC) harboring stx/eae were retrieved from diarrheic calves. Although none of the stx-or eae-positive isolates was verified as O157:H7, STEC isolates detected in apparently healthy calves have potential pathogenicity to humans highlighting their zoonotic importance as reservoirs. Atypical combinations of ETEC/STEC and ETEC/EPEC were also detected in percentages of 14.7 and 2.7%, respectively. Most of these atypical combinations were found more in buffalo calves than in cattle calves. While STEC and EPEC isolates were detected more in cattle calves than in buffalo calves, ETEC isolates were the same in the two species. The pathogenic E. coli infection in calves was recorded to be higher in the first weeks of life with the largest numbers of virulence factor-positive isolates detected at the age of 4 weeks. Histopathological examination of five intestinal samples collected from four dead buffalo calves revealed typical attaching and effacing (AE) lesion which was correlated with the presence of intimin encoding virulence gene (eae). Other lesions characterized by hemorrhagic enteritis, shortening and fusion of intestinal villi and desquamation of the lining epithelium of intestinal mucosa had also been detected.
从犊牛分离的大肠杆菌田间分离株，通过使用不同特异性引物的聚合酶链反应（PCR）检测，被鉴定并分类为最重要的腹泻致病性血清型。使用的 PCR 系统旨在检测代表编码菌毛 f5（K99）、志贺毒素（stx 和 stx）、耐热肠毒素（st）、不耐热肠毒素（lt）、侵袭素（eae）、溶血素（hylA）和 EAEC 耐热肠毒素（astA）的特定毒力基因的序列。在本研究中，从 100 头腹泻和 50 头无临床症状的接触牛以及 3 个月以下的水牛犊牛的 150 份粪便拭子中，共分离出 150 株大肠杆菌。在这 150 株分离的大肠杆菌中，106 株来自 77 头腹泻和 29 头无临床症状的接触牛，携带一种或多种研究的毒力基因。对分离株的分型可将其分类为志贺毒素产生型大肠杆菌（STEC）、肠致病性大肠杆菌（EPEC）、肠产毒性大肠杆菌（ETEC）和肠聚集性大肠杆菌（EAEC），分别占 30.7%、2.7%、12.7%和 7.3%。同时，f5、stx、stx、st、lt、eae、hylA 和 astA 基因的检出率分别为 17.3%、27.3%、6.7%、10%、37.3%、17.7%、9.3%和 20.7%。这些毒力基因单独存在或存在于不同的组合中，如 stx/eae、stx/st/f5、eae/st/f5 或 st/lt/f5。从腹泻犊牛中分离出 4 株带有 stx/eae 的附着-破坏型志贺毒素产生大肠杆菌（AE-STEC）。尽管没有一株 stx 或 eae 阳性分离株被证实为 O157：H7，但在无临床症状的犊牛中检测到的 STEC 分离株对人类具有潜在的致病性，强调了它们作为人类疾病来源的人畜共患病重要性。还检测到 14.7%和 2.7%的 ETEC/STEC 和 ETEC/EPEC 非典型组合。这些非典型组合大多在水牛犊牛中比在奶牛犊牛中更常见。虽然 STEC 和 EPEC 分离株在奶牛犊牛中的检出率高于水牛犊牛，但 ETEC 分离株在这两种牛中相同。犊牛中致病性大肠杆菌感染在生命的前几周更高，在 4 周龄时检测到的携带毒力因子的分离株数量最多。对从 4 头死亡水牛犊牛收集的 5 个肠组织样本的组织病理学检查显示出典型的附着和破坏（AE）病变，这与侵袭素编码毒力基因（eae）的存在相关。还检测到其他特征为出血性肠炎、肠绒毛缩短和融合以及肠黏膜衬里上皮脱落的病变。
