Isolation Identification and Characterization of Bacterial pathogens causing Calf Diarrhea with special reference to Escherichia coli.
N Hemashenpagam, B Kiruthiga, T Selvaraj, A Panneerselvam
antibiotic sensitivity, calf diarrhea, dna extraction, escherichia coli, pcrtetagene
N Hemashenpagam, B Kiruthiga, T Selvaraj, A Panneerselvam. Isolation Identification and Characterization of Bacterial pathogens causing Calf Diarrhea with special reference to Escherichia coli.. The Internet Journal of Microbiology. 2008 Volume 7 Number 2.
Though India is an agricultural country, livestock plays an important role. In bovine, calf mortality is due to many diseases, in that diarrhea or white scour plays a major role. Enteriocolibacillosis is the disease caused by
The livestock population of India is huge and animals as a whole play an importance role in the agricultural economy even though they often receive inadequate nourishment. Diarrhea in calves can be caused by a variety of pathogens including bacteria, viruses, protozoa and intestinal parasites. Among bacteria, enterotoxigenic
The disease characteristically affects calves within the first 10-14 days of age, usually within the first week when there is sudden onset of profuse yellow or white diarrhea causing rapid and severe dehydration. The calf quickly becomes recumbent. Accumulation of fluid in the abomasums and intestines gives the abdomen a bloated appearance. These enterotoxigenic
Materials And Methods
Collection of sample
Sterilized new polypropylene storage vial of 5ml capacity was used for collection of fecal sample from the diarrheic calves.
MacConkey agar, Eosin Methylene Blue agar, Brillant Green agar, Xylose-Lysin Deoxycholate, Nutrient agar, Peptone water, MR-VP broth, Nitrate broth, Phenol Red Base broth, Simmon’s citrate agar, Triple Sugar Iron agar, Christensen’s urea agar base, Motility test medium and Muller Hinton agar. It was autoclaved at 121oC for 15 minutes.
Antibiogram of the
Isolation of DNA from the
Result and Discussion
Out of 16 fecal samples from the diarrheic calves 12 were positive for bacterial isolation on to media like MacConkey agar, EMB, BGA, XLD. All the 12 isolates produced positive reaction to lactose fermentation on MacConkey agar plate, metallic green sheen colonies on EMB plates, yellowish green colonies on BGA, yellow colonies on XLD. These findings were in correlation with the presentation of Cowan (1993) and John Barnes
Gram staining were performed for all the isolates and revealed a Gram negative, non-acid fast, uniform staining non-spore forming bacilli. These findings were in correlation with the presentation of John Barnes
All the 12 isolates evinced positive reaction for indole test by converting the aminoacid tryptophane to indole with help of tryptophanes enzyme by the organism, Methyl Red test by utilization of glucose in the MRVP medium, triple sugar iron agar test, the isolates utilized all three sugars glucose, lactose and sucrose which results in acid slant or acid butt with gas production, carbohydrate fermentation test, utilized all three sugars by change of red colour (indicator phenol red) to yellow with gas production in Durham’s tube in Phenol red base medium, nitrate reduction test by converting the nitrate to nitrite, catalase test by the development of air bubbles on adding 3%H2O2 to the isolate in the glass slide.
All the 12 isolates were negative for Voges proskauer test where no acetone production was seen, citrate utilization test where no colour change in Simmon’s citrate medium and also negative for oxidase test.These findings were in correlation with Bettelheim, (1994) and Cowan, (1993).
Antibiotic susceptibility test were done for all the isolates. All the isolates were sensitive to ciprofloxacin(100%) followed by pefloxacin(100%), cephotaxime(100%), chloramphenicol(83.33%), co-trimoxazole(75%) and gentamicin(75%). In the present study the isolates were sensitive to gentamicin was similar to the findings of Jones
PCR amplification were done for two isolates. In the present study the PCR amplified fragments of approximately 417 bp was observed with use of tet(A) specific primers F- CGA GCC ATT CGC GAG AGC and R- GCC TCC TGC GCG ATC TGG in the 1.5% agarose with ethidium bromide stain electrophorsed and viewed under UV illumination. This findings were in agreement with that of Furushita