First Isolation of Bacillus licheniformis from Bovine Mastitis in Iran

Document Type : Original Articles

Authors

1 Department of Microbiology and Immunology , Faculty of Veterinary Medicine, University of Tehran, Tehran, Iran

2 Department of Microbiology and Immunology, Faculty of Veterinary Medicine, University of Tehran, Tehran, Iran

10.32592/ARI.2023.78.6.1690

Abstract

Bacillus licheniformis is a gram-positive, endospore-forming, saprophytic and facultative anaerobe that is resistant to heat and environmental conditions. This study was the first to isolate and confirm B. licheniformis as a cause of bovine mastitis in Iran. In the summer of 2020, 105 samples of mastitic milk were collected from dairy farms around Tehran and sent to the microbiology laboratory of the Faculty of Veterinary Medicine at the University of Tehran. The bacterial pathogens were identified using selective and differential culture media and confirmed by PCR to contain the toxin synthetase genes licA, licB and licC in mastitic isolates of B. licheniformis. Resistance patterns to 19 antibiotics were determined for two isolates of B. licheniformis. Staphylococcus aureus and Escherichia coli were identified as the most important organisms in the samples. B. licheniformis was isolated from the two samples containing all three genes. Both isolates were resistant to streptomycin, trimethoprim-sulfamethoxazole, cefixime, ampicillin, bacitracin, clindamycin, and gentamicin. B. licheniformis was reported for the first time in Iran as a cause of bovine mastitis with clinical symptoms. The first isolation of toxin-producing strains of B. licheniformis from mastitic cows in Iran raises concerns about the safety of dairy products. In principle, selected strains with toxigenic potential should not be used as feed additives and animal feed. However, whole genome sequencing is proposed to search for genes coding for toxins.

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References
1. Yakoubou S, Côté J-C. Assessment of a short
phylogenetic marker based on comparisons of 3'end 16S
rDNA and 5'end 16S-23S ITS nucleotide sequences on
the genus Xanthomonas. NS. 2010;2(12):1369-74.
2. Cutting SM. Bacillus probiotics. Food Microbiol.
2011;28(2):214-20.
3. Motta AS, Lorenzini DM, Brandelli A. Purification and
partial characterization of an antimicrobial peptide
produced by a novel Bacillus sp. isolated from the
Amazon Basin. Curr Microbiol. 2007;54:282-6.
4. Syrjälä P, Anttila M, Dillard K, Fossi M, Collin K,
Nylund M, et al. Causes of bovine abortion, stillbirth
and neonatal death in Finland 1999–2006. Acta Vet
Scand. 2007;49(1):1-2.
5. Sirohi P, Yadav AK, Singh NK. INHIBITORY
MECHANISM OF N-HEXANE AND DICHLOROMETHANE LEAF EXTRACTS OF CLERODENDRUM PHLOMIDIS LINN. ON FOOD BORN
PATHOGEN BACILLUS LICHENIFORMIS. Journal
of Plant Development Sciences Vol. 2019;11(9):501-10.
6. Shaheen R, Andersson MA, Apetroaie C, Schulz A,
Ehling-Schulz M, Ollilainen V-M, et al. Potential of
selected infant food formulas for production of Bacillus
cereus emetic toxin, cereulide. Int J Food Microbiol.
2006;107(3):287-94.
7. Markey B, Leonard F, Archambault M, Cullinane A,
Maguire D. Clinical veterinary microbiology e-book:
Elsevier sci; 2013.
8. DebMandal M, Mandal S, Kumar Pal N. Antibiotic
resistance prevalence and pattern in environmental
bacterial isolates. The Open Antimicrobial Agents
Journal. 2011;3(1).
9. Nieminen T, Rintaluoma N, Andersson M, Taimisto AM, Ali-Vehmas T, Seppälä A, et al. Toxinogenic
Bacillus pumilus and Bacillus licheniformis from
mastitic milk. Vet Microbiol. 2007;124(3-4):329-39.
10. Tapi A, Chollet-Imbert M, Scherens B, Jacques P. New
approach for the detection of non-ribosomal peptide
synthetase genes in Bacillus strains by polymerase chain
reaction. Applied microbiology and biotechnology.
2010;85:1521-31.
11. Xue Z, Marco ML. Milk and dairy products. Food
Microbiol (5th Ed). 2019:101-23.
12. Ostrov I, Sela N, Belausov E, Steinberg D, Shemesh M.
Adaptation of Bacillus species to dairy associated
environment facilitates their biofilm forming ability.
Food Microbiol. 2019;82:316-24.
13. Kent D, Chauhan K, Boor K, Wiedmann M, Martin N.
Spore test parameters matter: Mesophilic and
thermophilic spore counts detected in raw milk and
dairy powders differ significantly by test method. J
Dairy Sci. 2016;99(7):5180-91.
14.Rückert A, Ronimus RS, Morgan HW. A RAPD-based
survey of thermophilic bacilli in milk powders from
different countries. Int J Food Microbiol.
2004;96(3):263-72.
15. Ariznabarreta A, Gonzalo C, San Primitivo F.
Microbiological quality and somatic cell count of ewe
milk with special reference to staphylococci. J Dairy
Sci. 2002;85(6):1370-5.
16. Sadashiv S, Kaliwal B. Isolation, characterization and
antibiotic resistance of Bacillus sps. from bovine
mastitis in the region of north Karnataka, India. Int J
Curr Microbiol Appl Sci. 2014;3:360-73.
17. Adimpong DB, Sørensen KI, Thorsen L, StuerLauridsen B, Abdelgadir WS, Nielsen DS, et al.
Antimicrobial susceptibility of Bacillus strains isolated
from primary starters for African traditional bread
production and characterization of the bacitracin operon
and bacitracin biosynthesis. Appl Environ Microbiol.
2012;78(22):7903-14.
Nayeri Fasaei et al / Archives of Razi Institute, Vol. 78, No. 6 (2023) 1690-1697 1697
18. Allam NAT, Sedky D, Mira EK. The clinical impact of
antimicrobial resistance genomics in competition with
she-camels recurrent mastitis metabolomics due to
heterogeneous Bacillus licheniformis field isolates. Vet
World. 2017;10(11):1353.
19. Suliman R, Salih R. Bacterial Bovine Mastitis and
Formation of Specific antibodies in Milk and Sera,
Sudan. J Vet Med Health. 2019;3:113.
20. SALIH RRM. COMPARISON BETWEEN THE
PERCENTAGE OF INCIDENCE OF MASTITIS
CAUSED BY Bacillus spp. AND Staphylococcus spp.
IN WINTER SEASON IN KHARTOUM STATE,
SUDAN. Online J Anim Feed Res. 2015;5(4):112-6.
21. Scheldeman P, Herman L, Foster S, Heyndrickx M.
Bacillus sporothermodurans and other highly
heatā€resistant spore formers in milk. J Appl Microbiol.
2006;101(3):542-55.
22. Taylor JM, Sutherland AD, Aidoo KE, Logan NA.
Heat-stable toxin production by strains of Bacillus
cereus, Bacillus firmus, Bacillus megaterium, Bacillus
simplex and Bacillus licheniformis. FEMS
microbiology letters. 2005;242(2):313-7.
23. Nonnemann B, Lyhs U, Svennesen L, Kristensen KA,
Klaas IC, Pedersen K. Bovine mastitis bacteria resolved
by MALDI-TOF mass spectrometry. J Dairy Sci.
2019;102(3):2515-24.
24. Veith B, Herzberg C, Steckel S, Feesche J, Maurer KH,
Ehrenreich P, et al. The complete genome sequence of
Bacillus licheniformis DSM13, an organism with great
industrial potential. Microbial Physiology. 2004;
7(4):204-11.
25. Madslien E, Rønning H, Lindbäck T, Hassel B,
Andersson M, Granum P. Lichenysin is produced by
most Bacillus licheniformis strains. J Appl Micrbiol.
2013;115(4):1068-80.