Abstract
BackgroundThe nematode Caenorhabditis elegans has become increasingly used for screening antimicrobials and probiotics for pathogen control. It also provides a useful tool for studying microbe-host interactions. This study has established a C. elegans life-span assay to preselect probiotic bacteria for controlling K88+ enterotoxigenic Escherichia coli (ETEC), a pathogen causing pig diarrhea, and has determined a potential mechanism underlying the protection provided by Lactobacillus.Methodology/Principal FindingsLife-span of C. elegans was used to measure the response of worms to ETEC infection and protection provided by lactic acid-producing bacteria (LAB). Among 13 LAB isolates that varied in their ability to protect C. elegans from death induced by ETEC strain JG280, Lactobacillus zeae LB1 offered the highest level of protection (86%). The treatment with Lactobacillus did not reduce ETEC JG280 colonization in the nematode intestine. Feeding E. coli strain JFF4 (K88+ but lacking enterotoxin genes of estA, estB, and elt) did not cause death of worms. There was a significant increase in gene expression of estA, estB, and elt during ETEC JG280 infection, which was remarkably inhibited by isolate LB1. The clone with either estA or estB expressed in E. coli DH5α was as effective as ETEC JG280 in killing the nematode. However, the elt clone killed only approximately 40% of worms. The killing by the clones could also be prevented by isolate LB1. The same isolate only partially inhibited the gene expression of enterotoxins in both ETEC JG280 and E. coli DH5α in-vitro.Conclusions/SignificanceThe established life-span assay can be used for studies of probiotics to control ETEC (for effective selection and mechanistic studies). Heat-stable enterotoxins appeared to be the main factors responsible for the death of C. elegans. Inhibition of ETEC enterotoxin production, rather than interference of its intestinal colonization, appears to be the mechanism of protection offered by Lactobacillus.
Highlights
Probiotics are live microorganisms that can confer a healthy benefit on their host [1]
Establishment of a Life-span Assay of C. elegans Infected with enterotoxigenic Escherichia coli (ETEC) To establish a C. elegans life-span assay capable of measuring the response of worms to ETEC infection, K88+ ETEC strain JG280 at different concentrations, ranging from 107 to 109 colony forming units (CFU) per ml, was used in the assay instead of E. coli OP50 that is normally used as food to maintain the nematode when C. elegans reaches the L4 stage
The assay with C. elegans infected by JG280 was used to assess selected lactic acidproducing bacterial (LAB) isolates for their protection against the death of worms caused by JG280 infection
Summary
Probiotics are live microorganisms that can confer a healthy benefit on their host [1]. They have long been used in food animal production and widely considered to be a potential alternative to dietary antibiotics. The nematode has become increasingly used for screening antimicrobials, including preselecting probiotic bacteria for Salmonella control [7,11,12]. The nematode Caenorhabditis elegans has become increasingly used for screening antimicrobials and probiotics for pathogen control. This study has established a C. elegans life-span assay to preselect probiotic bacteria for controlling K88+ enterotoxigenic Escherichia coli (ETEC), a pathogen causing pig diarrhea, and has determined a potential mechanism underlying the protection provided by Lactobacillus
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