Genes Encoding the Virulence and the Antimicrobial Resistance in Enterotoxigenic and Shiga-Toxigenic E. coli Isolated from Diarrheic Calves

Abstract

Calf diarrhea is one of the considerable infectious diseases in calves, which results in tremendous economic losses globally. To determine the prevalence of Shiga-toxigenic E. coli (STEC) and Enterotoxigenic E. coli (ETEC) incriminated in calf diarrhea, with special reference to Shiga- toxins genes (stx1 and stx2) and enterotoxins genes (lt and sta) that govern their pathogenesis, as well as the virulence genes; eaeA (intimin) and f41(fimbrial adhesion), and the screening of their antibiogram and antimicrobial resistance genes; aadB, sul1, and bla-TEM, a total of 274 fecal samples were collected (April 2018-Feb 2019) from diarrheic calves at different farms in El-Sharqia Governorate, Egypt. The bacteriological examination revealed that the prevalence of E. coli in diarrheic calves was 28.8%. The serotyping of the isolated E. coli revealed 7 serogroups; O₂₆, O₁₂₈, O₁₁₁, O₁₂₅, O₄₅, O₁₁₉ and O₉₁. Furthermore, the Congo red binding test was carried out, where 89.8% of the examined strains (n = 71) were positive. The antibiogram of the isolated strains was investigated; the majority of E. coli serotypes exhibit multidrug resistance (MDR) to four antimicrobial agents; neomycin, gentamycin, streptomycin, and amikacin. Polymerase chain reaction (PCR) was used to detect the prevalence of the virulence genes; stx1, stx2 lt, sta, f41 and eaeA, as well as the antibiotic resistance genes; aadB, sul1, and bla-TEM. The prevalence of STEC was 20.2% (n = 16), while the prevalence of ETEC was 30.4% (n = 24). Briefly, the Shiga toxins genes; stx1 and stx2, are the most prevalent virulence genes associated with STEC, which are responsible for the pathogenesis of the disease and helped by the intimin gene (eaeA). In addition, the lt gene is the most prevalent enterotoxin gene accompanied by the ETEC strains, either alone or in combination with sta and/or f41 genes. The majority of pathogenic E. coli incriminated in calf diarrhea possesses the aadB resistance gene, followed by the sul1 gene. Enrofloxacin, florfenicol, amoxicillin-clavulanic acid, and ampicillin-sulbactam, are the most effective antimicrobial agents against the isolated STEC and ETEC strains.

Keywords: ETEC; STEC; antimicrobial resistance genes; calves; diarrhea; virulence genes.

Figure 1

(A) Electrophoretic pattern of aadB (319 bp): M: 100–1000 bp DNA ladder; POS: Positive control; Neg: Negative control; Lanes: 1–8: positive E. coli strains. (B) Electrophoretic pattern of sul1 gene (433 bp): M: 100–1000 bp DNA ladder, POS: Positive control; Neg: Negative control; Lanes: 1, 4, 5: positive E. coli strains; Lanes: 2, 3, 6, 7, 8: negative E. coli strains. (C) Electrophoretic pattern of blaTEM gene (516 bp): M: 100–600 bp DNA ladder; POS: Positive control; Neg: Negative control; Lanes: 2–3: positive E. coli strains; Lanes: 1, 4, 5, 6, 7, 8: negative E. coli strains.

Figure 2

(A) Electrophoretic pattern of stx1 gene (614 bp); M: 100–1000 bp DNA ladder; POS: Positive control; Neg: Negative control; Lanes 1–6: positive E. coli strains; Lanes 7, 8: negative E. coli strains. (B) Electrophoretic pattern of stx2 gene (779 bp); M: 100–1000 bp DNA ladder, POS: Positive control; Neg: Negative control; Lanes 1–7: positive E. coli strains; Lane 8: negative E. coli strain. (C) Electrophoretic pattern of eaeA gene (248 bp): M: 100–600 bp DNA ladder; POS: Positive control; NEG: Negative control; 1–7: positive E. coli strains; Lane 8: negative E. coli strain.

Figure 3

(A) Electrophoretic pattern of lt gene (605 bp): M: 100–1000 bp DNA ladder; POS: Positive control; Neg: Negative control; Lanes: 3, 6, 7: positive E. coli strains; Lanes 1, 2, 4, 5, 8: negative E. coli strains. (B) Electrophoretic pattern of sta gene (219 bp): M: 100–1000 bp DNA ladder, POS: Positive control; Neg: Negative control; Lanes: 2, 3, 5: positive E. coli strains; Lanes 1, 4, 6, 7, 8: negative E. coli strains. (C) Electrophoretic pattern of f41 gene (380 bp): M: 100–600 bp DNA ladder, POS: Positive control; Neg: Negative control; Lanes: 1–6: positive E. coli strains; Lanes 7, 8: negative E. coli strains.