Development of a monoclonal antibody-based co-agglutination test to detect enterotoxigenic Escherichia coli isolated from diarrheic neonatal calves
نویسندگان
چکیده
Escherichia coli (E. coli) strains were collected from young diarrheic calves in farms and field. Strains that expressed the K99 (F5) antigen were identified by agglutination tests using reference antibodies to K99 antigen and electron microscopy. The K99 antigen from a selected field strain (SAR-14) was heat-extracted and fractionated on a Sepharose CL-4B column. Further purification was carried out by sodium deoxycholate treatment and/or ion-exchange chromatography. Monoclonal antibodies to purified K99 antigen were produced by the hybridoma technique, and a specific clone, NEK99-5.6.12, was selected for propagation in tissue culture. The antibodies, thus obtained, were affinity-purified, characterized and coated onto Giemsastained Cowan-I strain of Staphylococcus aureus (S. aureus). The antibody-coated S. aureus were used in a coagglutination test to detect K99+ E. coli isolated from feces of diarrheic calves. The specificity of the test was validated against reference monoclonal antibodies used in co-agglutination tests, as well as in ELISA. Specificity of the monoclonal antibodies was also tested against various Gram negative bacteria. The developed antibodies specifically detected purified K99 antigen in immunoblots, as well as K99+ E. coli in ELISA and co-agglutination tests. The co-agglutination test was specific and convenient for large-scale screening of K99+ E. coli isolates.
منابع مشابه
Passive protective effect of anti-K99 antibodies against enterotoxigenic E.coli infection in neonatal calves
BACKGROUND: BACKGROUND: Enterotoxigenic Escherichia coli (ETEC) is the main causative agents of neonatal calf diarrhea. Passive transfer of antibodies that is derived from the dams of the calves through their colostrum plays an important role in disease prevention. OBJECTIVES: To determine the presence of natural specific antibodies against K99 antigen in dam’s serum and colostrum as well as in...
متن کاملCorrelation between neonatal calf diarrhea and the level of maternally derived antibodies
Passively acquired antibodies through colostrum will protect calves against etiological agents of neonatal calf diarrhea. Among them enteric diseases due to strains of Enterotoxigenic Escherichia coli (ETEC) are the most commonly occurring form of colibacillosis in newborn calves. Specific antibodies against whole ETEC cells and total immunoglobulin G in dam serum, colostrum and calf serum were...
متن کاملMolecular characterization of enterotoxigenic Escherichia coli isolated from neonatal calves diarrhea
Enterotoxigenic Escherichia coli (ETEC) produces different virulence factors allowing the bacterium to colonize and develop watery diarrhea. Proteomics studies have also introduced new protein belonging to the secretion pathways, antigen 43 (Ag43), which plays important role in E. coli pathogenesis. The objective of this study was to investigate O-types and virulence factors o...
متن کاملProduction, Purification and Characterization of Chicken Egg Yolk Monoclonal Antibody Against Colonization factor antigen -1 of Enterotoxigenic Escherichia coli Causing Diarrhea
Enterotoxigenic Escherichia coli (ETEC) causes diarrhea in both humans and animals. The contaminated food and water are the most common vehicles for ETEC infection. The colonization factor antigen (CFA-1) is a fimbriae protein that promotes adherence of the ETEC strain to the epithelium of the small intestine of the host. In this study IgY proteins were produced against the CFA-1 of ETEC in imm...
متن کاملGenetic variation among Escherichia coli isolates from human and calves by using RAPD PCR
Background: Various strains of Escherichia coli (E. coli) are known as major causes of intestinal and extraintestinal infections in humans and various animal species. Molecular methods are important for the identification of bacterial isolates and nucleotide sequence variations, as well as information on tracking bacterial agents related to the outbreaks, the frequency of the bacterial genetic ...
متن کامل