Open AccessArticle
Detection of Plasmid-Mediated Colistin
Resistant mcr-1 Gene in Escherichia coli Isolated
from Infected Chicken Livers in Nepal
Sayara Bista 1,†,Upendra Thapa Shrestha 1,†,Binod Dhungel 1,Pragya Koirala 2,Tulsi Ram Gompo 2,Nabaraj Shrestha 2,Nabaraj Adhikari 1,Dev Raj Joshi 1,Megha Raj Banjara 1,Bipin Adhikari 3,Komal Raj Rijal 1,* andPrakash Ghimire 1
1Central Department of Microbiology, Tribhuvan University,
Kirtipur, Kathmandu 44618, Nepal
2Central Veterinary Laboratory Ministry of Agriculture, Land
Management and Cooperatives, Government of Nepal, Tripureshwor, Kathmandu
44618, Nepal
3Centre for Tropical Medicine and Global Health, Nuffield
Department of Medicine, University of Oxford, Oxford OX1 3SY, UK
* Author to whom correspondence should be addressed.
† These authors have equally contributed in this study.
Animals 2020, 10(11),
2060; https://doi.org/10.3390/ani10112060 (registering DOI)
Received: 30 August 2020 / Revised:
22 October 2020 / Accepted: 25 October 2020 / Published: 7 November 2020
(This article belongs to the Special Issue Poultry
Microbiology and Immunology)
Simple Summary
The poultry industry is one of the
top agribusinesses in Nepal. However, despite the government’s restriction on
the use of antibiotics as growth promotors in animals, the overuse and misuse
of antibiotics can be seen all over the country. Such inappropriate use of
antibiotics has led to the rise of antibiotic resistance among treatment
options for both human and animal pathogens. Several findings suggest the
failure of colistin, a polymyxin E antibiotic (once regarded as the last resort
drug), in the treatment of human bacterial infections is due to the emergence
and spread of the plasmid-mediated colistin resistance gene (mcr-1)
among Gram-negative bacterial pathogens. The emergence and rapid transfer of
resistant strains in poultry farms are associated with unwanted loss of
livestock, economic burden and spread of drug-resistance to other animals,
humans and the environment, as well. In this study, we characterized the mcr-1 gene
from infected chicken livers, where prevalence was found to be alarmingly high.
This study identifies the result of regulatory failures. Therefore, this report
provides valuable reference to the policy makers so that a more effective
policy can be formulated and implemented to curb the spread of drug-resistant
pathogens.
Background: Plasmid-mediated resistance to the colistin in poultry
is considered as an emerging problem worldwide. While poultry constitutes the
major industry in Nepal, there is a paucity of evidence on colistin resistance
in Escherichia coli isolates causing natural infections in
poultry. This study aimed to explore the prevalence of plasmid-mediated
colistin resistance gene, mcr-1 in E. coli isolated
from liver samples of dead poultry suspected of E. coli infections.
Methods: A total of two hundred and seventy liver samples (227
broilers and 43 layers) from dead poultry suspected of colibacillosis were
collected from post-mortem in the Central Veterinary Laboratory (CVL),
Kathmandu, between 1 February and 31 July 2019. The specimens were processed to
isolate and identify E. coli; an antimicrobial susceptibility test
(AST) using disk diffusion method was performed with 12 different antibiotics:
Amikacin (30 µg), ampicillin (10 µg), ciprofloxacin (5 µg), chloramphenicol (30
µg), cefoxitin (30 µg), ceftazidime (30 µg), ceftriaxone (30 µg), cotrimoxazole
(25 µg), gentamicin (10 µg), imipenem (10 µg), levofloxacin (5 µg) and
tetracycline (30 µg). Colistin resistance was determined by agar dilution
method and colistin-resistant strains were further screened for
plasmid-mediated mcr-1 gene, using conventional polymerase
chain reaction (PCR).
Results: Out of 270 liver samples, 53.3% (144/270) showed
growth of E. coli. The highest number (54%; 109/202) of E.
coli isolates was obtained in the liver samples from poultry birds (of
both types) aged less than forty days. In AST, 95.1% (137/144) and 82.6%
(119/144) of E. coli isolates were resistant against
tetracycline and ciprofloxacin, respectively, while 13.2% (19/144) and 25.7%
(37/144) isolates were resistant to cefoxitin and imipenem, respectively. In
the same assay, 76.4% (110/144) E. coli isolates were
multi-drug resistant (MDR). The phenotypic prevalence of colistin resistance
was 28.5% (41/144). In the PCR assay, 43.9% (18/41) of colistin-resistant
isolates were screened positive for plasmid-mediated mcr-1.
Conclusion: The high prevalence of mcr-1 in
colistin-resistant E. coli isolates in our study is a cause of
concern for the probable coming emergence of colistin resistance in human
pathogens, due to horizontal transfer of resistant genes from poultry to human
isolates.
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