Isolation And Characterization Of Escherichia Coli From Piglets With Neonatal Diarrhea In Part Of Jiangsu Province And Challenge Of Commercial Pigs With O24ETEC Isolates

Escherichia coli is an important pathogen of neonatal piglets diarrhoea. ETEC infections result in severe economic losses due to high mortality and reduced growth rates among pig farms. From2013to2014,397E. coli strains collect from rectal swabs of piglets with yellow and white sour, duodenum jejunum and ileum of dead piglets from part of Jiangsu province and its surrounding area. Their serogroups, toxin types and adhesin types were characterized.1Isolation and characterization of Escherichia coli from piglets with neonatal diarrhea disease in part of Jiangsu Province397E. coli strains from piglets with neonatal diarrhea were characterized. For O-serogroups,328strains were determined,64strains were unserotypeable, and5strains were self-agglutinated.328E. coli isolates were distributed in thirty-eight serotypes. O24、O101、0141、055、O20and0149were the major serotypes of all the identified isolates, these serotypeable isolates harbor252isolate (76.83%,252/328), they were the predominant serogroups among Escherichia coli strains isolated from newborn piglets. In this study,024、055and O105were serotypes which had seldom reported in the past. So O24、055and0105trend to be the predominant serotypes from Escherichia coli strains isolated from newborn piglets in Jiangsu Province.Multiple PCR was used to characterize the following toxin genes of397E. coli isolates, sta, stb, It, stx2e and eaeA. The result shows that among397isolates,171isolates with toxin genes were detected (43.07%,171/397). According to the result,13toxin-gene types can be grouped. sta、lt、stb and stx2e are the predominant toxin-gene types which represent in93,22,14and8isolates, respectively.The rabbit intestinal-loop assay was used for detection of LT toxin, the neonatal mouse intragastric administration assay was used to detect STA toxin, and the cytopathic effect of Vero cells was used to detect Vero toxin. The isolates with024serotype were examined mentioned above. The result showed that toxin genes of these024isolates expressed and exhibited their virulence.Monoclonal antibody agglutination test and MRHA assay were used to test fimbria of F4, F5, F6and F41of the397E. coli isolates,37isolates which expressed adhesion were detected (9.32%,37/397). These isolates can express one or more adhesions. In this study,11strains belonged to F4,10strains belonged to F5+F41+,7strains belonged to F4+F5+,6strains belonged to F41+and3strains belonged to987P+.Analysis the correlation among O-serogroups, enterotoxin genes and fimbrial-types of the397E. coli isolates was analyzed. The result showed that171toxin gene positive isolates covered17serotypes, O24, O141,0149,0101, O20and0105were the predominant serotypes, including57,22,15,9,9and7isolates, respectively. The predominant toxin-gene types were sta, It, stx2e+eaeA, which occupied59.65%(34/57),10.53%(6/57),8.78%(5/57) of the024toxin-gene positive isolates. The major toxin-gene types were stb and stb+stx2e, which harbored12isolates (75%,12/15)and3isolates (25%,3/15) of the0149toxin-gene positive isolates. The common O101often harbored sta and stx2e. The O20harbored sta which occupied88.89%(8/9). The major toxin-gene was sta, which occupied71.43%(5/7) of the0105toxin-gene positive isolates.37adhesin positive isolates harbored34isolates which owned both toxin-genes and adhesins. The14O149isolates exhibited stb+/F4+F5+、stb+/F41+and stb+stx2e+/F5+F41+. composed of7,5and2isolates, respectively. Five virulence factors types of024, among them2isolates belonged to sta+/F4+,2isolates belonged to stb+/F4+,1isolate belonged to lt/+/F5+F41+,1isolate belonged to sta+stb+/F4+and1isolate belonged to sta+lt+stx2e+/F5+F41+.2Challenge of commercial piglets with O24ETEC isolatesIn this study, new-born, no milk nursing commercial piglets were assigned to three groups,024serotype isolate LYG129(stb+/F5+F41+) challenge group,024serotype isolate LYG141(It+/F41+) challenge group and024serotype isolate YC267(stb+/F4+F5+) challenge group. Mental conditions, food consumption, diarrhea of challenged piglets were monitored. Rectal swab samples were collected per3hour post-challenge, the study was terminated at72hours after inoculation. The results indicated that three challenged pigs all exhibited diarrhea in different degree15hours post-challenge, including yellow loose stool, watery diarrhea, and then visible dehydration, weight lose and so on. Among three groups,024strain YC267(stb+/F4+F5+) challenged pig experienced more sever diarrhea than those challenged with the rest two strains, the piglet was died after24hours post-challenge, the challenged strain could be isolated from the heart blood of dead piglets. Since12hours post-challenge, challenged strains have been isolated from the rectal swab samples of challenged piglets, the O-serotype, toxin-gene types and adhesion types of these isolates were the same with challenged strains.Therefore, ETEC with024serotype can cause diarrhea of neonatal piglets with typical clinical manifestations.