Jihong ZHANG,
Shijun ZHENG,
Tangming ZHU,
Siguo LIU,
Rusong LI,
Ting ZHU,
Suwen HUANG,
Jianfeng WANG,
Jianbo NI,
Xiuling ZHAO,
Jing SUN,
Shaotang HUANG
3754
Rapid and simultaneous detection of Mycobacterium bovis and Mycobacterium avium subsp. paratuberculosis in invasive flies by duplex PCR
A rapid and effective polymerase chain reaction (PCR) test, specific for the detection of Mycobacterium bovis and Mycobacterium avium subsp. paratuberculosis in invasive flies, was developed. This duplex PCR technique was specifically designed to amplify the sequences of IS6110 of M. bovis and IS900 of M. avium subsp. paratuberculosis. The results of the interference test, simulated contamination test, and several other experiments indicated that this technique had high stability, specificity, and sensitivity with detection limits at 92.98 pg of M. bovis and 110.27 pg of M. avium subsp. paratuberculosis. Therefore, this duplex PCR test is a rapid and effective technique to simultaneously detect M. bovis and M. avium subsp. paratuberculosis.
Rapid and simultaneous detection of Mycobacterium bovis and Mycobacterium avium subsp. paratuberculosis in invasive flies by duplex PCR
A rapid and effective polymerase chain reaction (PCR) test, specific for the detection of Mycobacterium bovis and Mycobacterium avium subsp. paratuberculosis in invasive flies, was developed. This duplex PCR technique was specifically designed to amplify the sequences of IS6110 of M. bovis and IS900 of M. avium subsp. paratuberculosis. The results of the interference test, simulated contamination test, and several other experiments indicated that this technique had high stability, specificity, and sensitivity with detection limits at 92.98 pg of M. bovis and 110.27 pg of M. avium subsp. paratuberculosis. Therefore, this duplex PCR test is a rapid and effective technique to simultaneously detect M. bovis and M. avium subsp. paratuberculosis.
___
- Grant, I.R., Pope, C.M., O’Riordan, L.M., Ball, H.J., Rowe, M.T.: Improved detection of Mycobacterium avium subsp. paratuberculosis in milk by immunomagnetic PCR. Vet. Microbiol., 2000; 77: 369–378.
- Khare, S., Ficht, T.A., Santos, R.L., Romano, J., Ficht, A.R., Zhang, S., Grant, I.R., Libal, M., Hunter, D., Adams, L.G.: Rapid and sensitive detection of Mycobacterium avium subsp. paratuberculosis in bovine milk and feces by a combination of immunomagnetic bead separation-conventional PCR and real-time PCR. J. Clin. Microbiol., 2004; 42: 1075–1081.
- Sreevatsan, S., Bookout, J.B., Ringpis, F., Perumaalla, V.S., Ficht, T.A., Adams, L.G., Hagius, S.D., Elzer, P.H., Bricker, B.J., Kumar, G.K., Rajasekhar, M., Isloor, S., Barathur, R.R.: A multiplex approach to molecular detection of Brucella abortus and/or Mycobacterium bovis infection in cattle. J. Clin. Microbiol., 2000; 38: 2602–2610.
- Miller, J.M., Jenny, A.L., Payeur, J.B.: Polymerase chain reaction detection of Mycobacterium tuberculosis complex and Mycobacterium avium organisms in formalin-fixed tissues from culture-negative ruminants. Vet. Microbiol., 2002; 87: 15–
- Fang, Y., Wu, W.H., Pepper, J.L., Larsen, J.L., Marras, S.A.E., Nelson, E.A., Epperson, W.B., Christopher-Hennings, J.: Comparison of real-time, quantitative PCR with molecular beacons to nested PCR and culture methods for detection of Mycobacterium avium subsp. paratuberculosis in bovine fecal samples. J. Clin. Microbiol., 2002; 40: 287–291.
- Giese, S.B., Ahrens, P.: Detection of Mycobacterium avium subsp. paratuberculosis in milk from clinically affected cows by PCR and culture. Vet. Microbiol., 2000; 77: 291–297.
- Angela, D.P., Giuseppina, C., Tony, F.V., Bijo, B., Fatmira, S., Giuseppina, T.: Detection of Mycobacterium tuberculosis complex in milk using polymerase chain reaction (PCR). Food Control, 2006; 17: 776–780.
- Bhide, M., Chakurkar, E., Tkacikova, L., Barbuddhe, S., Novak, M., Mikula, I.: IS900-PCR-based detection and characterization of Mycobacterium avium subsp. paratuberculosis from buffy coat of cattle and sheep. Vet. Microbiol., 2006; 112: 33–41.
- Chen, R., Liu, Z.Y., Gao, X.B., Liu, Z.L., Wu, X.W., Zeng, B.J., Luo, C.B., Lin, Z.X.: Development and application of a real time PCR detection kit specific for Mycobacterium paratuberculosis. J. Microbiol., 2009; 36: 292–297.
- Englund, S., Bölske, G., Johansson, K.E.: An IS900-like sequence found in a Mycobacterium sp. other than Mycobacterium avium subsp. paratuberculosis. FEMS Microbiol. Lett., 2002; 209: 267– 2
- Ikonomopoulos, J., Gazouli, M., Pavlik, I., Bartos, M., Zacharatos, P., Xylouri, E., Papalambros, E., Gorgoulis, V.: Comparative evaluation of PCR assays for the robust molecular detection of Mycobacterium avium subsp. paratuberculosis. J. Microbiol. Methods, 2004; 56: 315–321.
- Kim, S.G., Shin, S.J., Jacobson, R.H., Miller, L.J., Harpending, P.R., Stehman, S.M., Rossiter, C.A., Lein, D.A.: Development and application of quantitative polymerase chain reaction assay based on the ABI 7700 system (TaqMan) for detection and quantification of Mycobacterium avium subsp. paratuberculosis. J. Vet. Diagn. Invest., 2002; 14: 126–131.
- Millar, D., Ford, J., Sanderson, J., Withey, S., Tizard, M., Doran, T., Hermon-Taylor, J.: IS900 PCR to detect Mycobacterium paratuberculosis in retail supplies of whole pasteurized cows’ milk in England and Wales. Appl. Environ. Microbiol., 1996; 62: 3446–3452.
- Savelkoul, P.H.M., Catsburg, A., Mulder, S., Oostendorp, L., Schirm, J., Wilke, H., van der Zanden, A.G., Noordhoek, G.T.: Detection of Mycobacterium tuberculosis complex with real time PCR: comparison of different primer-probe sets based on the IS6110 element. J. Microbiol. Methods, 2006; 66: 177–180.
- Bakshi, C.S., Shah, D.H., Verma, R., Singh, R.K., Malik, M.: Rapid differentiation of Mycobacterium bovis and Mycobacterium tuberculosis based on a 12.7-kb fragment by a single tube multiplex-PCR. Vet. Microbiol., 2005; 109: 211– 2
- Gao, A., Mutharia, L., Raymond, M., Odumeru, J.: Improved template DNA preparation procedure for detection of Mycobacterium avium subsp. paratuberculosis in milk by PCR. J. Microbiol. Methods, 2007; 69: 417–420.
- Hong, S.K., Kim, B.J., Yun, Y.J., Lee, K.H., Kim, E.C., Park, E.M., Park, Y.G., Bai, G.H., Kook, Y.H.: Identification of Mycobacterium tuberculosis by PCR-linked reverse hybridization using specific rpoB oligonucleotide probes. J. Microbiol. Methods, 2004; 59: 71–79.
- Kurabachew, M., Enger, Ø., Sandaa, R.A., Skuce, R., Bjorvatn, B.: A multiplex polymerase chain reaction assay for genus-, group- and species-specific detection of mycobacteria. Diagn. Microbiol. Infect. Dis., 2004; 49: 99–104.
- Stabel, J.R., Bosworth, T.L., Kirkbride, T.A., Forde, R.L., Whitlock, R.H.: A simple, rapid, and effective method for the extraction of Mycobacterium paratuberculosis DNA from fecal samples for polymerase chain reaction. J. Vet. Diagn. Invest., 2004; 16: 22–30.
- Tobler, N.E., Pfunder, M., Herzog, K., Frey, J.E., Altwegg, M.: Rapid detection and species identification of Mycobacterium spp. using real-time PCR and DNA-microarray. J. Microbiol. Methods, 2006; 66: 116–124.
- Van Soolingen, D., Hermans, P.W., de Haas, P.E., Sol, D.R., van Embden, J.D.: Occurrence and stability of insertion sequences in Mycobacterium tuberculosis complex strains: evaluation of an insertion sequence-dependent DNA polymorphism as a tool in the epidemiology of tuberculosis. J. Clin. Microbiol., 1991; 29: 2578–2586.
- Ellingson, J.L.E., Stabel, J.R., Radcliff, R.P., Whitlock, R.H., Miller, J.M.: Detection of Mycobacterium avium subspecies paratuberculosis in free-ranging bison (Bison bison) by PCR. Mol. Cell. Probes, 2005; 19: 219–225.
- Yang, L.: Detection of deer tuberculosis by PCR technique and the cloning and expression of HSP65 Gene. MSc. Northeast Forestry University, Harbin, China. 2006.
- General Administration of Quality Supervision, Inspection and Quarantine of the People’s Republic of China. Protocol of the PCR for Mycobacterium paratuberculosis (SN/T 19072007). 2007.
- Sampson, S., Warren, R., Richardson, M., van der Spuy, G., van Helden, P., Dunlap, N., Benjamin W.H. Jr: IS6110 Insertions in Mycobacterium tuberculosis: predominantly into coding regions. J. Clin. Microbiol., 2001; 39: 3423–3424.
- Motiwala, A.S., Strother, M., Amonsin, A., Byrum, B., Naser, S.A., Stabel, J.R., Shulaw, W.P., Bannantine, J.P., Kapur, V., Sreevatsan, S.: Molecular epidemiology of Mycobacterium avium subsp. paratuberculosis: evidence for limited strain diversity, strain sharing, and identification of unique targets for diagnosis. J. Clin. Microbiol., 2003; 41: 2015–2026.
- Vansnick, E., De Rijk, P., Vercammen, F., Geysen, D., Rigouts, L., Portaels, F.: Newly developed primers for the detection of Mycobacterium avium subspecies paratuberculosis. Vet. Microbiol., 2004; 100: 197–204.
- Whittington, R.J., Marsh, I., Turner, M.J., McAllister, S., Choy, E., Eamens, G.J., Marshall, D.J., Ottaway, S.: Rapid detection of Mycobacterium paratuberculosis in clinical samples from ruminants and in spiked environmental samples by modified BACTEC 12B radiometric culture and direct confirmation by IS900 PCR. J. Clin. Microbiol., 1998; 36: 701–707.
- Bartos, M., Hlozek, P., Svastova, P., Dvorska, L., Bull, T., Matlova, L., Parmova, I., Kuhn, I., Stubbs, J., Moravkova, M., Kintr, J., Beran, V., Melicharek, I., Ocepek, M., Pavlik, I.: Identification of members of Mycobacterium avium species by Accu-Probes, serotyping, and single IS900, IS901, IS1245 and IS901-flanking region PCR with internal standards. J. Microbiol. Methods, 2006; 64: 333–345.
- Vitale, F., Capra, G., Maxia, L., Reale, S., Vesco, G., Caracappa, S.: Detection of Mycobacterium tuberculosis complex in cattle by PCR using milk, lymph node aspirates, and nasal swabs. J. Clin. Microbiol., 1998; 36: 1050–1055.
- Farkas, T., Antal, M., Sámi, L., Germán, P., Kecskeméti, S., Kardos, G., Belák, S., Kiss, I.: Rapid and simultaneous detection of avian influenza and Newcastle disease viruses by duplex polymerase chain reaction assay. Zoonoses Public Health, 2007; 54: 38–43.
- Liu, S.G., Guo, S.P., Wang, C.L., Shao, M.L., Zhang, X.H., Guo, Y., Gong, Q.: A novel fusion protein-based indirect enzymelinked immunosorbent assay for the detection of bovine tuberculosis. Tuberculosis (Edinb.), 2007; 87: 212–217.