Application of Multiplex PCR Assay for Campylobacter fetus Sub sp. Detection and Differentiation in Bovine Preputial Wash

https://doi.org/10.21059/buletinpeternak.v42i1.26114

Seruni Agistiana(1*), Surachmi Setiyaningsih(2), Agustin Indrawati(3)

(1) Indonesian Agriculture Quarantine Agency, Indonesian Ministry of Agriculture, Jakarta Selatan, 12550, Indonesia
(2) Department of Animal Diseases and Veterinary Health, Faculty of Veterinary Medicine, Bogor Agricultural University, Bogor, 16680, Indonesia
(3) Department of Animal Diseases and Veterinary Health, Faculty of Veterinary Medicine, Bogor Agricultural University, Bogor, 16680, Indonesia
(*) Corresponding Author

Abstract


The aim of this research was to evaluate the detection and differentiation of C. fetus subspecies through multiplex PCR (mPCR) method from the bovine preputial wash. The evaluation was done by comparing 2 extraction methods, which was by commercial kit extraction and boiling method, and also by measuring the effect of Transport Enrichment Media (TEM) addition as bacterial enrichment. Spiked bovine preputial washes with 2 extraction methods (commercial kit and boiling), with and without TEM addition were used as a sample. The results showed that the highest sensitivity/specificity was by boiling extraction without TEM addition, which reached 83.3%/100% for all or grouped sample. The results also supported by Poisson regression analysis which showed the comparison between sample with and without TEM addition which had 1.3 Incidence Rate Ratio (IRR). The obtained IRR score indicated that sample without TEM addition had 30% better result compared to the sample with TEM addition. Furthermore, a comparison between commercial kit and boiling extraction showed no significant difference (1.029 IRR). The research concluded that boiling extraction without TEM addition is the most accurate, fast and economical extraction method regarding its high sensitivity and specificity value, with less than 24 hours testing time and without any commercial kit requirements. However, a validation and verification with field sample were still needed before applied in the laboratory.


Keywords


Bovine genital campylobacteriosis; Campylobacter fetus; Cff; Cfv; Multiplex PCR

Full Text:

PDF


References

Al-Soud, W. A. and P. Rådström. 2001. Purification and characterization of PCR-inhibitory components in blood cells. J. Clin. Microbiol. 39:  485-493.

BBKP Soekarno Hatta. 2017. Rekapitulasi Sapi Impor. Laporan Tahunan Tahun Anggaran 2016, Tangerang.

BBKP Tanjung Priok. 2017. Rekapitulasi Sapi Impor. Laporan Tahunan Tahun Anggaran 2016, Jakarta.

Beniawan, A. 2017. Alternatif media transport untuk Campylobacter fetus subs. Venerealis. Tesis Sekolah Pascasarjana Institut Pertanian Bogor, Bogor.

Chaban, B., S. Chu, S. Hendrick, C. Waldner and J. E. Hill. 2012. Evaluation of a Campylobacter fetus subspecies venerealis real-time quantitative polymerase chain reaction for direct analysis of bovine preputial samples. Can. J. Vet. Res. 76: 166-173.

Chang, W. and J. E. Ogg. 1971. Transduction and mutation to glycine tolerance in Vibrio fetus. Am.  J.Vet. Res. 23: 649-653.

Corkill, G. and R. Rapley. 2008. The manipulation of ucleic acids: basic tools and techiques. In: Molecular Biomethods Handbook Second Edition. J.M. Walker and N.J. Rapley (Eds.). Humana Press. USA. Pp. 3-16.

Eaglesome, M. D. and M. M Garcia. 1992. Microbial agents associated with bovine genital tract infections and semen. Part I. Brucella abortus, Leptospira, Campylobacter fetus and Tritrichomonas foetus. Vet. Bull. 62: 743-775.

Gilardi, G. L. 1985. Pseudomonas. In:  Manual of Clinical Microbiology (4th edn). E.H. Lenette, A. Balows, W.J. Hausler, H.J. Shadomy (Eds). American Society of Microbiology, Wahsington. pp. 350-372.

Hardjoutomo, S. 1998. Tinjauan Tentang Vibriosis Sapi di Indonesia. Wartazoa 7: 10-14.

Hum, S., K. Quinn, J. Brunner, and S. L. W. On. 1997. Evaluation of a PCR assay for identification and differentiation of Campylobacter fetus subspecies.  Aust. Vet. J. 75: 827-831.

Hum, S., M. Hornitzky and T. Berg. 2009. Bovine Genital Campylobacteriosis. Australia and New Zealand Standard Diagnostic Procedures November 2009. Pp. 1-19.

Iraola, G., M. Hernandez, L. Calleros, F. Paolicchi, S. Silveyra, A. Velilla, L. Carretto, E. Rodriguez and R. Perez. 2012. Application of a multiplex PCR assay for Campylobacter fetus detection and subspecies differentiation in uncultured samples of aborted bovine fetuses. J. Vet. Sci. 13: 371-6.

Kementan RI. 2016. Mentan luncurkan upsus SIWAB (Sapi Indukan Wajib Bunting) http://www.pertanian.go.id/ap_posts/detil/672/2016/10/08/22/36/43/Mentan%20Luncurkan%20UPSUS%20SIWAB%20-Sapi%20Indukan%20Wajib%20Bunting-.  Accessed 5 June 2017.

Lander, K. P. 1990. The application of a transport and enrichment medium to the diagnosis of  Campylobacter fetus  infections in bulls. British. Vet. J. 146: 334-340.

Markoulatos, P., N. Siafakas and M. Moncany. 2002. Multiplex polymerase chain reaction: a practical approach. J. Clin. Lab. Anal. 16: 47-51.  

Modolo, J. R., C. A. M Lopes and T. Genari. 2000. Occurrence of Campylobacter in the genitals of teaser bulls maintained at an embryo transfer center. Arquivo Brasileiro de Medicina Veterinária e Zootecnia, 52: 96-97.

OIE. 2016. Disease Distribution Maps. http://www.oie.int/wahis_2/public/wahid.php/Diseaseinformation/Diseasedistributionmap?disease_type_hidden=&disease_id_hidden=&selected_disease_name_hidden=&disease_type=0&disease_id_terrestrial=31&species_t=0&disease_id_aquatic=-999&species_a=0&sta_method=semesterly&selected_start_year=2015&selected_report_period=1&selected_start_month=1&date_submit=OK.  Accessed 12 July 2016.

OIE. 2017. Chapter 2.4.4: Bovine Genital Campylobacteriosis. http://www.oie.int/fileadmin/Home/eng/Health_standards/tahm/2.04.04_BGC.pdf. Accessed 10 Juli 2017.

Rådström, P., R. Knutsson, P. Wolffs, M. Lövenklev, and C. Löfström. 2004. Pre-PCR processing: Strategies to generate PCR-compatible samples. Mol Biotechnol. 26: 133-146.

Schmidt, T. 2008. Detection of Campylobacter fetus in bovine preputial scrapings using PCR and culture assay. Dissertation University of Pretoria, South Africa.

Schulze, F., A. Bagon A. M. Wolfgang and H. Hotzel. 2006. Identification of Campylobacter fetus subspesies by phenotypic differentiation and PCR. J. Clin. Microbiol. 44: 2019-2024.

Surzycky, R. 2000. Molecular and Cellular Biology. Wadsworth Inc., Belmon.

Theophilus, B. D. M. 2008. Principles and Medical Applications of the Polymerase Chain Reaction. In: Molecular Biomethods Handbook Second Edition. J.M. Walker and N.J. Rapley (Eds.). Humana Press. USA. Pp. 33-19.

Thunberg, R. L., T. T. Tran, and M. O. Walderhau. 2000. Detection of thermophilic Campylobacter spp. in blood-free enriched samples of inoculated foods by the polymerase chain reaction. J. Food Prot. 63: 299-303.

Truyers, I., T. Luke, D. Wilson and N. Sargison. 2014. Diagnosis and management of venereal campylobacteriosis in beef cattle. BMC Veterinary Research 10: 280.

Van Bergen, M. A., K. E. Dingle, M. C. Maiden, D. G. Newell, L. van der Graaf-Van Bloois, J. P.  and J. A. Waagenar. 2005. Clonal nature of Campylobacter fetus as defined by multilocus sequence typing. J. Clin. Microbiol. 43: 5888-5898.

Van der Graaf-van Bloois, L., M. A van Bergen, F. J. van der Wal, A. G. de Boer, B. Duim, T. Schmidt, and J. A. Wagenaar. 2013. Evaluation of molecular assay for identification Campylobacter fetus species and subspecies and development of a C. fetus spesific real time PCR Assay. J. Microbiol. Methods. 95: 93-97.

Van der Graaf-van Bloois, L., W. G. Miller, E. Yee, M. Rijnsburger, J. A. Wagenaar, and B. Duim. 2014. Inconsistency of phenotypic and genomic characteristics of campylobacterfetus subspecies requires reevaluation of current diagnostics. J. Clin. Microbiol. 52: 4183-4188.

Wilson, I. G. 1997. Inhibition and facilitation of nucleic acid amplification. Appl. Environ. Microbiol. 63: 3741-3751.

 



DOI: https://doi.org/10.21059/buletinpeternak.v42i1.26114

Article Metrics

Abstract views : 867 | views : 618

Refbacks

  • There are currently no refbacks.




Creative Commons License
This work is licensed under a Creative Commons Attribution-ShareAlike 4.0 International License.

Buletin Peternakan (Bulletin of Animal Science) Indexed by:

 Harvard Library
Creative Commons License
This work is licensed under a Creative Commons Attribution-ShareAlike 4.0 International License.