Zoonotic Study of Escherichia coli O157 H7 from Animals to Human through Arbitrarily Primed Polymerase Chain Reaction (AP-PCR) Methods.

ZOONOTIC STUDY OF ESCHERICHIA COLI O157:H7 FROM ANIMALS TO HUMAN THROUGH
ARBITRARILY PRIMED POLYMERASE CHAIN REACTION (AP-PCR) METHOD

A.

Introduction

Escherichia coli O157:H7 is a zoonotic agent of the type of Shiga toxin producing Escherichia coli that can cause disease in human, and cattle is known as
the main reservoir of these bacteria (Karmali et al., 2010). The infection by these bacteria in animals usually asymtomatic, whereas these bacterial infection
in humans usually show clinical symptoms i.e. diarrhea, colitis hemorrhagic and hemolytic uremic syndrome (HUS) (Acheson, 2010; Wani et al., 2004) . This
study report the application of AP-PCR method in order to study the zoonotic potency of E. coli O157:H7 from animals as a main reservoir of these
bacteria to human.

B.

Methods

Cultivation of 14 isolates of E. coli O57 i.e.
ATCC 43894 (positive control), KL52(7),
KL87(7), KL30(4), KL45(1), KL(48(2), KL85(1),
KL83(5), KL24(5), KL68(1), KL-106(3), KL55(6), SM-25(1),SM-7(1)

C.

Extraction of bacterial DNA using QIAamp DNA Mini
Kits

AP-PCR using primers M13F and
M13R with PCR program: I. 94OC, 5
min; II. 39 cycles (94OC, 5 min, 35OC,
1 min, 72OC, 1 min), III. 72OC, 5 min.

Results

Evolutionary distance of each isolate was
measured with algorithm unweighted pair
group method using arithmetic averages
(UPGMA)

A

Fragments and total bands of 14 isolates of E. coli
O157:H7 generated by AP-PCR using primers M13F and
M13R.

B

Phenogram of E. coli O157:H7, which was constructed
using UPGMA, which placed both human and animal
isolates in one clade

D.
AP-PCR profile of genomic DNA of E. coli O157:H7 by using
primers M13F (A) and M13R (B) on 1.5%. agarose gel. Line
1: ATCC 43894 (positive control); Line 2: KL52(7); 3:
KL87(7); 4: KL30(4); 5: KL45(1); 6: KL(48(2); 7: KL85(1); 8:
KL83(5); 9: KL24(5); 10: KL68(1); 11: KL-106(3); 12: KL55(6); 13: SM-25(1); 14: SM-7(1); M: Marker 100 bp DNA
Ladder.

Conclusion

Arbitrarily primed polymerase chain reaction (AP-PCR)
method provides for simples and rapid for tracing of
zoonotic agent E. coli O157:H7, which were supported
by its highly sensitivity
Similarity coefficient among isolates of E. coli O157:H7,
which showed closely similarity between human and
animal isolates

References
1. Acheson, D.W.K., 2000. J. Food Protect. (6): 819-821.
2. Karmali MA, Gannon V, Sargeant JM. 2010. Vet. Microbiol. 140: 360-370.
3. Wani, S.A., Samanta, I., Munshi, Z.H., Bhat, M.A., and Nishikawa, Y., 2004. J. Appl. Microbiol. 100:108–113

Acknowledgments
The research was funded by the Directorate of Research
and Community Services, Directorate General of Higher
Education, Republic of Indonesia through Udayana
Research Grants with contract No.21.34 / UN 14 / SBRC /
2012, January 19th, 2012.

Presented at”Global Health Conference of Researcher in Emerging Disease at Convergence of Animal, Human and Environment Health” on 9-13 February 2015
Chiang Mai University, Thailand.