Establishment of a linear regression equation for quantification of beta-hemolytic Escherichia coli in different media and survival of hemolytic Escherichia coli after blending with three different media

  • Kim, Jae Cheol (Department of Agriculture and Food Western Australia) ;
  • Pluske, John R. (School of Veterinary and Life Sciences, Murdoch University) ;
  • Yoo, Jaehong (Department of Animal Science and Biotechnology, Chungnam National University) ;
  • Heo, Jung Min (Department of Animal Science and Biotechnology, Chungnam National University)
  • Received : 2014.03.25
  • Accepted : 2014.06.16
  • Published : 2014.06.30


Pathogenic E. coli associated post-weaning diarrhea (PWD) and edema disease are common diseases in commercially-housed weanling pigs. An enterotoxigenic E. coli (ETEC) oral challenge model has been used to mimic the physiological responses observed in commercial conditions. However, an oral challenge procedure has two major limitations: (1) the ETEC cell density is unknown at the point of oral inoculation, and (2) blending ETEC with traditional TSB (trypticase soy broth) is not palatable and hence decreases acceptability by piglets. Therefore, the purposes of this study were to (1) establish a regression equation that can be used for estimation of ETEC concentration in dilution media using the spectrophotometric measurement of cell density; and (2) examine survival of ETEC after blending either with TSB, sweetener or dextrose. A strain of ETEC (serogroup beta-hemolytic E. coli O149; K91; F4; toxins LT, STa, STb) was grown in TSB for 3.5 hours, centrifuged, the supernatant was discarded, and the ETEC pellet was then blended either with TSB (100 mL), sweetener (60 mL TSB + 40 mL fruit flavored concentrate), or dextrose (50 mL TSB + 50 mL dextrose; 0.5g/mL dextrose). Cell density was measured using the colorimetric method and also plated on a 5% sheep blood agar for counting of ETEC colony forming units at 0, 5, 35, 65 and 125 min after blending. The optical density at 600 nm explained 83% of ETEC colony forming units, indicating that the established linear equation (y= 6E+08x - 4E+07, P<0.004) can be used for robust quantification of ETEC cell density in TSB, sweetener and dextrose media. When ETEC was blended with sweetener and dextrose, survival of ETEC was decreased by 45% and 72% within 5 min post-blending. Therefore, further research is required to find out the suitable medium that has potential to improve palatability without compromising survival of ETEC.


Supported by : Rural Development Administration


  1. Bianch AA, Baneyx F. 1999. Hyperosmotic shock induces the ${\sigma}^{32}$ and ${\sigma}^{E}$ stress regulons of Escherichia coli. Molecular Microbiology 34:1029-1038.
  2. Hampson DJ. 1994. Postweaning Escherichia coli diarrhoea in pigs. In: Gyles, C.L. (Ed.), Escherichia coli in domestic animals and humans, CAB International, pp. 171-192, Guildford, UK.
  3. Heo JM, Kim JC, Hansen CF, Mullan BP, Hampson DJ, Pluske JR. 2010. Effects of dietary protein level and zinc oxide supplementation on performance responses and gastrointestinal tract characteristics in weaner pigs challenged with an enterotoxigenic strain of Escherichia coli. Animal Production Science 50:827-836.
  4. Heo JM, Opapeju FO, Pluske JR, Kim JC, Hampson DJ, Nyachoti CM, 2013. Gastrointestinal health and function in weaned pigs: a review of feeding strategies to control post-weaning diarrhoea without using in-feed antimicrobial compounds. Journal of Animal Physiology and Animal Nutrition 97: 207-237.
  5. Kim JC, Hansen CF, Mullan BP, Pluske JR. 2012. Nutrition and pathology of weaner pigs: Nutritional strategies to support barrier function in the gastrointestinal tract. Animal Feed Science and Technology 173:3-16.
  6. McDonald DE, Pethick DW, Mullan BP, Hampson DJ, 2001. Increasing viscosity of the intestinal contents alters small intestinal structure and intestinal growth, and stimulates proliferation of enterotoxigenic Escherichia coli in newlyweaned pigs. The British Journal of Nutrition 86:487-498.
  7. Pluske JR, Pethick DW, Hopwood D, Hampson DJ. 2002 Nutritional influences on some major enteric bacterial diseases of pig. Nutrition Research Reviews 15:333-371.
  8. Small P, Blankenhorn D, Welty D, Zinser E, Slonczewski JL. 1994. Acid and base resistance in Escherichia coli and Shigella flexneri: role of rpoS and growth pH. Journal of Bacteriology 176:1729-1737.