Direct detection of Mycobacterium avium subsp. Paratuberculosis in bovine milk by multiplex real-time PCR

• Autorzy: Selim A. , El-haig M. , Galila E. S. , Gaede W.
• Języki publikacji: EN

Abstrakty

EN

The study aimed at direct detection of Mycobacterium avium subsp. Paratuberculosis (MAP) in milk by evaluating a multiplex real-time PCR assay targeting IS900 and ISMAV2 sequences including the amplification of PUC19-plasmid as internal control. The sensitivity of the assays was evaluated by testing MAP isolates in broad linear range of DNA (50 ng – 5 fg/μl). For the validation of the specificity, 6 MAP isolates and 22 isolates of genus Mycobacteriaceae were tested.Results revealed that reproducible detection limit for real-time PCR targeting IS900 and ISMAV2 was 5 fg/μl and 50 fg/μl respectively. By targeting ISMAV2 sequence, 100% specificity was detected. However, a cross reaction with 5 ng/μl of genome of 3 M. avian subspecies avium strains was detected by targeting IS900 and negative in lower genome quantity (5pg/μl). To maximize the assay’s detection sensitivity,an efficient strategy for MAP-DNA extraction from spiked milk was assessed. Targeting of IS900 was sensitive and targeting ISMAV2 was very specific. Therefore, a multiplex real-time PCR assay targeting IS900 and ISMAV2 in combination with two commercial DNA extraction kits could be an ideal sensitive and specific protocol for routine large-scale analysis of milk samples and other clinical specimens from man and animals.

Słowa kluczowe

EN

detection; Mycobacterium avium ssp.paratuberculosis; milk; bovine milk; multiplex real-time polymerase chain reaction; paratuberculosis

• Wydawca: -
• Czasopismo: Animal Science Papers and Reports
• Rocznik: 2013
• Tom: 31
• Numer: 4
• Opis fizyczny: p.291-302,ref.

Twórcy

autor

Selim A.

• Faculty of Veterinary Medicine, Benha University, P.O.Box 13736, Toukh, Egypt

autor

El-haig M.

• Department of Internal Medicine and Infectious Diseases, Faculty of Veterinary Medicine, Suez Canal University, Ismailia, Egypt

autor

Galila E. S.

• Faculty of Veterinary Medicine, Benha University, P.O.Box 13736, Toukh, Egypt

autor

Gaede W.

• Department of Veterinary Medicine, State Institute for Consumer Protection of Saxony-Anhalt, Stendal, Germany

Bibliografia

• Cocito C., Gilot P., Coene M., De Kesel M., Poupart P., Vannuffel P., 1994 –Paratuberculosis. Journal of Clinical Microbiology Review 7, 823-345.
• Cousins D.V., Whittington R., Marsh I., Masters A., Evans R.J., Kluver P., 1999 – Mycobacteria distinct from Mycobacterium avium subsp. paratuberculosis isolated from the faeces of ruminants possess IS900-like sequences detectable by IS900 polymerase chain reaction:Implications for diagnosis. Molecular Cellular Probes 13, 431-442.
• DALZIEL T., 1913 – Chronic intestinal enteritis. British Medical Journal. 1068-70
• Englund S., Ballagi -Pordany A., Bölske G., Johansson K.E., 1999 – Single PCR and nested PCR with a mimic molecule for detection of Mycobacterium avium subsp. paratuberculosis.Diagnostic Microbiology Infectious Disease 33, 163-171.
• Englund S., Bölske G., Ballagi -Pordany A., Johansson K.E., 2001 – Detection of Mycobacterium avium subsp. paratuberculosis in tissue samples by single, flourescent and nested PCR based on the IS900 gene. Veterinary Microbiology 81, 257-271.
• Giese S.B., Ahrens P., 2000 – Detection of Mycobacterium avium subsp. paratuberculosis in milk from clinically affected cows by PCR and culture. Veterinary Microbiology 77, 291–297.
• Grant I.R., Ball H.J. Rowe M.J., 1998 – Isolation of Mycobacterium paratuberculosis from milk by immunomagnetic separation. Applied Environmental Microbiology 64, 3153-3158.
• Grant I.R., Pope C.M., O’Riordan L.M., Ball H.J., Rowe M.T., 2000 – Improved detection of Mycobacterium avium subsp. paratuberculosis in milk by immunomagnetic PCR. Veterinary Microbiology 77, 369-378.
• Harris N.B., Barletta R.G., 2001 – Mycobacterium avium subsp. paratuberculosis in veterinary medicine. Clinical Microbiology Review 14, 489-512.
• Hughes V.M., Stevenson K., Sharp J.M., 2001 – Improved preparation of high molecular weight DNA for pulsed-field gel electrophoresis from mycobacteria. Journal of Microbiological Methods 44, 209-215.
• 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., 2004 – 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 PCR. Journal of Clinical Microbiology 42, 1075-1081.
• Kim S.G., Shin S.J., Jacobson R.H., Miller L.J., Harpending P.R., Stehman S.M.,Rossiter C.A., Lein D.A., 2002 – 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. Journal of Veterinary Diagnostic Investigation 14,126-131.
• Köhler H., Burkert B., Pavilk I., Diller R., Geue L., Conraths F.J., Martin G., 2008 – Evaluation of five ELISA test kits for the measurment of antibodies against M.avium subsp.paratuberculosis in bovine serum. Berliner und Müncher Tierärztliche Wochenschrift 121, 203-210.
• Lantz P.G., Hahnhagerdal B., Radstrom P., 1994 – Sample preparation methods in PCR-based detection of food pathogens. Trends in Food Science Technology 5, 384-389.
• Mäde D., Reiting R., Strauch E., Ketteritzsch K., Wicke A., 2008 – A real-time PCR for Detection of Pathogenic Yersinia enterocolitica in food combined with an Universal Internal Amplification Control System. Journal für Verbraucherschutz und Lebensmittelsicherheit 3, 141-151.
• Möbius P., Hotzel H., Rassbach A., Köhler H., 2008 – Comparison of 13 single-rounds and nested PCR assays targeting IS900, ISMAV2, F57 and locus 255 for detection of Mycobacterium avium subsp. Paratuberculosis. Veterinary Microbiology 126, 324-333.
• Odumeru J., Gao A., Chen S., Raymond M., Mutharia L., 2001 – Use of the bead beater for preparation of Mycobacterium paratuberculosis template DNA in milk. Canadian Journal of Veterinary Research 65, 201-205.
• Ravva S.V., Stanker L.H., 2005 – Real-time quantitative PCR detection of Mycobacterium avium subsp. paratuberculosis and differentiation from other mycobacterium using SYBR Green and TaqMan assays. Journal of Microbiological Methods 63, 305-317.
• Rodriguez -Lazaro D., D´Agostino M., Herrewegh A., Pla M., Cook N.,Ikonomopulos J., 2005 – Real-time PCR-based methods for detection of Mycobacterium avium subsp. paratuberculosis in water and milk. International Journal of Food Micorbiology 101, 93-104.
• Schönenbrücher H., Abdulmawjood A., Failing K., Bülte M., 2008 – New Triplex Real-time PCR Assay for detection of Mycobacterium avium subsp. Paratuberculosis in Bovine faces. Applied and Environmental Microbiology 47, 2751-2758.
• Selim A. 2011 – Evaluation of detection of Mycobacterium avium subsp. paratuberculosis in bovine feces by real-time PCR. (Dissertation med. Vet). Leipzig University. Leipzig.
• Stabel J.R., 1998 – Johne´s disease: a hidden threat. Journal of Dairy Science 81, 283-288.
• Stabel J.R., Bannantine J.P., 2005 – Development of nested PCR method targeting a unique multicopy element, ISMAV02, for detection of Mycobacterium avium subsp. paratuberculosis in faecal samples. Journal of Clinical Microbiology 43, 4744-4750.
• Stratmann J., Stevenson K., Gerlach G.F., 2006 – Peptide aMptD-mediated capture PCR for detection of Mycobacterium avium subsp. paratuberculosis in bulk milk samples. Applied Environmental Microbiology 72, 5150-5158.
• Tasara T., Hoelzle L.E., Stephan R., 2005 – Development and evaluation of a Mycobacterium avium subsp. paratuberculosis specific multiplex PCR assay. International Journal of Food Microbiology 104, 279-287.
• Van Soolingen D., Hermans P.W.M., de Haas P.E., Soll D.R., Van Embden J.D., 1991 – 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. Journal of Clinical Microbiology 29, 2578-2586.
• Vansnick E., Derijik , P., Vercammen F., Geysen D., Rigouts L., Portaels F.,2004 – Newly developed primers for detection of Mycobacterium avium subsp. Paratuberculosis.Veterinary Microbiology 100, 197-204.