شناسایی و تعیین سویه مایکوباکتریوم پاراتوبرکلوزیس (MAP) به روش PCR و REA بر اساس قطعات درون جایگیر IS900 و IS1311

نوع مقاله : مقاله پژوهشی

نویسندگان

1 2دانشیار، گروه پژوهشی بیوتکنولوژی کشاورزی و دامی، پژوهشکده فناوری زیستی، دانشگاه فردوسی مشهد1دانشیار، گروه علوم دامی، دانشکده کشاورزی، دانشگاه فردوسی مشهد

2 3دانشجوی مقطع دکتری، گروه علوم دامی، دانشگاه فردوسی مشهد

3 دانشجوی مقطع دکتری، گروه علوم دامی، دانشگاه فردوسی مشهد

4 جوی مقطع دکتری، گروه علوم دامی، دانشگاه فردوسی مشهد

چکیده

هدف از این مطالعه شناسایی مایکوباکتریوم آویوم زیرگونه پاراتوبرکلوزیس (M. avium paratuberculosis)، بررسی درصد شیوع آلودگی به آن و همچنین بهینه سازی یک روش سریع، دقیق و ساده به کمک PCR برای تشخیص این زیرگونه و جدایه های آن بود. مایکوباکتریوم آویوم زیرگونه پاراتوبرکلوزیس (MAP)، از لحاظ اقتصادی عامل یکی از مهمترین بیماری ها در صنعت دامپروری به نام بیماری یون (Johne's Disease) می باشد، که یکی از عفونت های مزمن نشخوارکنندگان به شمار می رود. به منظور شناسایی این زیرگونه 243 نمونه مدفوع و 56 نمونه شیر خام از دام های مشکوک گاوداری های اطراف مشهد جمع آوری شد. پس از استخراج DNA به منظور شناسایی نمونه های آلوده، قطعه درون جایگیر اختصاصی MAP به نام IS900 به طول 413 جفت باز، با استفاده از پرایمرهای P90 و P91 تکثیر شد. سپس به منظور تعیین جدایه گاوی و گوسفندی MAP، نسبت به تکثیر قطعه درون جایگیر IS1311 با طول 268 جفت باز به کمک دو پرایمر M56 و M94  اقدام شد. قطعات حاصله به کمک آنزیم HinfI هضم شدند. تعداد 107 نمونه از 243 نمونه مدفوع (44%) و 10 نمونه از 56 نمونه شیر خام (18%) مورد مطالعه، آلوده به MAP بودند. از بین 56 نمونه شیر و مدفوع که از دام های یکسانی گرفته شده بودند 19 نمونه مدفوع و 10 نمونه شیر، آلوده شناسایی شدند (18% در مقابل 34%). نتایج هضم آنزیمی نیز نشان داد که تمامی MAP های شناسایی شده از جدایه گاوی می باشند.
 

کلیدواژه‌ها

عنوان مقاله [English]

Identification and strain determination of M. paratuberculosis (MAP) by PCR and REA methods based on IS900 and IS1311 insertion segments

نویسندگان [English]

  • Mohammadreza Nassiri 1
  • Mohammad hassan Jahandar 2
  • Mehdi Soltani 3
  • Morteza Mahdavi 4
  • Mohammad Doosti 3
چکیده [English]

The purpose of this study was to identifying the M. avium paratuberculosis (MAP), investigating its outbreaks and optimizing a fast, accurate and simple method based on PCR and RFLP for detection of this subspecies and their strains. This pathogen cause a chronic infection called Johne's disease. Johne's disease a ruminant chronic infection and economically, is one of the most important disease in livestock industry. 243 fecal samples and 56 raw milk samples of suspected cattle, from some farms of Mashhad were gathered. After DNA extraction, in order to identifying contaminated samples, a specific MAP insertion segment called IS900, with length of 413bp was amplified with PCR using P90 and P91 primers. Then, in order to determining the cow and sheep strains of MAP, 268bp fragment of insertion segment IS1311  was amplified with PCR using M56 and M94 primers. The resulting fragments were digested with HinfI restriction enzyme. 107 of 243 stool samples (44%) and 10 of 56 raw milk samples (18%) were identified as infected with MAP. Among 56 Milk and feces samples taken from the same animal, 19 stools and 10 milk samples were identified as infected (18% vs. 34%). Results of enzymatic digestion also showed that all detected MAP were from bovine isolates.

کلیدواژه‌ها [English]

  • M. paratuberculosis
  • Johne's disease
  • IS900
  • IS1311
  • REA-PCR

اصل مقاله

M. paratuberculosis ,Johne's disease,  IS900, IS1311, REA-PCR.

 

مراجع

 
Ayele WY, Bartos M, Svastova P, Pavlik I (2004). Distribution of Mycobacterium avium subsp. paratuberculosis in organs of naturally infected bull-calves and breeding bulls. Veterinary Microbiology 103: 209-217.
Ayele WY, Machackova M, Pavlik I (2001). The transmission and impact of paratuberculosis infection in domestic and wild ruminants. Veterinary Medicine (Czech) 46: 205-224.
Ayele WY, Svastova P, Roubal P, Bartos M, Pavlik I (2005). Mycobacterium avium subspecies paratuber-culosis cultured from locally and commercially pasteurized cow’s milk in the Czech Republic. Applied and Environmental Microbiology 71: 1210-1214.
Bhide M, Chakurkar E, Tkacikova L, Barbuddhe S, Novak M, Mikula I (2006). IS900-PCR-based detection and characterization of Mycobacterium avium subsp. paratuberculosis from buffy coat of cattle and sheep. Veterinary Microbiology 112: 33-41.
Chaleshtari RS (2009). Detection of Mycobacterium paratuberculosis in Raw Milk Samples from Holstein Dairy Cows of Shahre kord by PCR. Medical Laboratory Journal 3: 15-19.
Chiodini RJ, Van Kruiningen HJ, Merkal RS (1984). Ruminant paratuberculosis Johne’s disease: the current status and future prospects. Cornell Vet 74: 218-262.
Collins M (2003). Update on paratuberculosis: Epidemiology of Johne’s disease and the biology of Mycobacterium paratuberculosis. Veterinary Ireland Journal 56: 565-574.
Corti S, Stephan R (2002). Detection of Mycobacterium avium subspecies paratuberculosis specific IS900 insertion sequences in bulk-tank milk samples obtained from different regions throughout Switzerland. BMC Microbiol 2: 15-22.
D’Haese E, Dumon I, Werbrouck H, J.V D, Herman L (2005). Improved detection of Mycobacterium paratuberculosis in milk. The Journal of Dairy Research 72: 125-128.
De Juan L, Mateos A, Dominguez L, Sharp JM, Stevenson K (2005). Genetic diversity of Mycobacterium avium subspecies paratuberculosis isolates from goats detected by pulsed-field gel electrophoresis. Veterinary Microbiology 106: 249-257.
Diéguez F, Arnaiz I, Sanjuán M, Vilar M, López M, Yus E (2007). Prevalence of serum antibodies to Mycobacterium avium subsp. paratuberculosis in cattle in Galatia (north west spain). Preventive Veterinary Medicine 82: 321-326.
Douarre PE, Cashman W, Buckley J, Coffey A, O’Mahony JM (2010). Isolation and detection of Mycobacterium avium subsp. paratuberculosis (MAP) from cattle in Ireland using both traditional culture and molecular based methods. Gut Pathogens 2: 11.
Ellingson JLE, Anderson JL, Koziczkowski JJ, Radcliff RP, Sloan SJ, Allen SE, Sullivan NM (2005). Detection of Viable Mycobacterium avium subsp. paratuberculosis in Retail Pasteurized Whole Milk by Two Culture Methods and PCR Journal of Food Protection 68:966-972.
Ellingson JLE, Bolin CA, Stabel JR (1998). Identification of a gene unique to Mycobacterium avium subspecies paratuberculosis and application to diagnosis of paratuberculosis. Molecular and Cellular Probes 12: 133-142.
Englund S, Bolske G, Johansson K-E (2002). An IS900-like sequence found in a Mycobacterium sp. other than Mycobacterium avium subsp. paratuberculosis. FEMS Microbiology Letters 209: 267-271.
Eriks IS, Munck KT, Besser TE, Cantor GH, Kapur V (1996). Rapid differentiation of Mycobacterium avium and M. paratuberculosis by PCR and restriction enzyme analysis. Journal of Clinical Microbiology 34: 734-737.
Gonda M, Chang Y, Shook G, Collins M, Kirkpatrick B (2007). Effect of Mycobacterium paratuberculosis infection on production, reproduction, and health traits in US Holsteins. Preventive Veterinary Medicine 80: 103-119.
Good M, Clegg T, Sheridan H, Yearsely D, O’ Brien T, Egan J, Mullowney P (2009). Prevalence and distribution of paratuberculosis (Johne’s disease) in cattle herds in Ireland. Irish Veterinary Journal 62: 597-606.
Grant I, Ball H, Rowe M (2002). Incidence of Mycobacterium paratuberculosis in Bulk Raw and commercially pasteurized cows’ milk from approved dairy processing establishments in the United Kingdom. Applied and Environmental Microbiology 68: 2428-2435.
Grant IR (2005). Zoonotic potential of Mycobacterium avium subsp. paratuberculosis: the current position. Journal of Applied Microbiology 98: 1282-1293.
Haghkhah M, Ansari-Lari M, Novin-Baheran A, Bahramy A (2008). Herd- level prevalence of Mycobacterium avium subspecies paratuberculosis by bulk-tank milk PCR in Fars province (Southern Iran) dairy herds. Preventive Veterinary Medicine 86: 8-13.
Haji Hajikolaei MR, Ghorbanpoor M, Solaymani M (2006). The prevalence of Mycobacterium paratuberculosis infection in ileocecal valve of cattle slaughtered in Ahvaz abattoir, southern Iran. Iranian Journal of Veterinary Research 7: 77-80.
Hasani tabatabai A, Firozi R (2005) Animal bacterial disease vol 1, 3edn. Tehran university pub, Tehran.
Jorgensen JB, Jensen PT (1978). Enzyme-linked immunosorbent assay (ELISA) for detection of antibod-ies to Mycobacterium paratuberculosis in cattle. Acta Veterinaria Scandinavica 19: 310-312.
Khol JL, Kralik P, Slana I, Beran V, Aurich C, Baumgartner W, Pavlik I (2010). Consecutive excretion of Mycobacterium avium subspecies paratuberculosis in semen of a breeding bull compared to the distribution in feces, tissue and blood by IS900 and F57 quantitative real-time PCR and culture examinations. Journal of Veterinary Medical Science 72: 1283-1288.
Mutharia L, K lassen M, Fairles J, Barbut S, Gill C (2010). Mycobacterium avium subsp. paratuberculosis in muscle, lymphatic and organ tissues from cows with advanced Johne’s disease. International Journal of Food Microbiology 136: 340-344.
Nielsen S, Enevoldsen C, Gröhn Y (2002). The Mycobacterium avium subsp. Paratuberculosis ELISA response by parity and stage of lactation. Preventive Veterinary Medicine 54: 1-10.
Nielsen S, Toft N (2009). A review of prevalence of paratuberculosis in farmed animals in Europe. Preventive Veterinary Medicine 88: 1-14.
O’Mahony J, Hill C (2002). Real times PCR assay for the detection and quantitation of Mycobacterium avium subsp. paratuberculosis using SYBR Green and the Light Cycler. Journal of Microbiological Methods 51: 283-293.
O’Mahony J, Hill C (2004). Rapid real-time PCR assay for detection and quantitation of Mycobacterium avium subsp. paratuberculosis DNA in artificially contaminated milk. Applied and Environmental Microbiology 70: 4561-4568.
Olsen I, Sigurgardottir G, Djonne B (2002). Paratuberculosis with special reference to cattle. Veterinary Quarterly 24: 12-28.
Pavlik I, Bartl J, Dvorska L, Svastova P, du MR, Machackova M, Yayo AW (2000). Epidemiology of paratuberculosis in wild ruminants studied by restriction fragment length polymorphism in the Czech Republic during the period 1995-1998. Veterinary Microbiology 77: 231-251.
Pillai SR, Jayarao BM, Gummo JD, Hue EC, Tiwari D, Stabel JR, Whitlock RH (2001). Identification and subtyping of Mycobacterium avium subsp. paratuberculosis and Mycobacterium avium subsp. avium by randomly amplified
polymorphic DNA. Veterinary Microbiology 79: 275-284.
Rademaker JL, Vissers MM, Te Giffel MC (2007). Effective heat inactivation of Mycobacterium avium subsp. paratuberculosis in raw milk contaminated with naturally infected feces. Applied and Environmental Microbiology 73: 4185-4190.
Sadeghi-Garmarodi F (1993). An abattoir study of mycobacterium paratuberculosis infection in cattle and buffalo. Doctorate Veterinary Medicine. Tehran, Tehran.
Shahmoradi AH (2009). Study of John’s disease in industrlal & semi industrial cattle husbandry of Esfahan province. Veterinary Researches in Pajouhesh & Sazandegi 6: 13-17.
Sharma G, Singh S, Sevilla I, Singh A, Whittington R, Juste R (2008). Evaluation of indigenous milk ELISA with m-culture and m-PCR for the diagnosis of Bovine Johne’s disease (BJD) in lactating Indian dairy cattle. Research in Veterinary Science 84: 30-37.
Singh S, Singh A, Singh R, Sandhu K, Singh P, Sohal J (2007). Evaluation of highly sensitive indigenous milk ELISA kit with fecal culture, milk culture and fecal-PCR for the diagnosis of bovine Johne’s disease (BJD) in India Comparative Immunology, Microbiology & Infectious Diseases 30: 175-186.
Slana I, Kralik P, Kralova A, Babak V, Pavlik I (2012). Examination of milk filters by real-time PCR as a herd-level indicator of the presence of Mycobacterium avium subspecies paratuberculosis in dairy herds. Journal of Dairy Science 95: 1162-1165.
Slana I, Kralik P, Kralova A, Pavlik I (2008a). On-farm spread of Mycobacterium avium ssp. paratuberculosis in raw milk studied by IS900 and F57 competitive real time quantitative PCR and culture examination. International Journal of Food Microbiology 128: 250-257.
Slana I, Paolicchi F, Janstova B, Navratilova P, Pavlik I (2008b). Detection methods for Mycobacterium avium subsp. paratuberculosis in milk and milk products: a review. Veterinarni Medicina 53: 283-306.
Soltani M, Nassiry MR, Eftekhari Shahroudi F, Bassami MR (2008). Detection of Mycobacterium paratuberculosis in Feces and Milk Samples from Holstein Dairy Cows by PCR. Biotechnology 7: 582-585.
Soltani M, Nassiry MR, Eftekhari Shahroudi F, Bassami MR (2010). PCR-Restriction Endonuclease Analysis for strain Typing of Mycobacterium avium Subsp. Paratuberculosisbased on Polymorphisms in IS 1311. Middle east journal of scienticfic research 5: 311-315.
Stabel J, Bannantine J (2005). Development of a nested PCR method targeting a unique multicopy element, ISMap02, for detection of Mycobacterium avium subsp. Paratuberculosis in fecal samples. Journal of Clinical Microbiology 43: 4744-4750.
Stable J, Wells S, Wagner B (2002). Relationships between fecal culture, ELISA, and bulk-tank milk test results for john`s disease in US dairy herds. Journal of Dairy Science 85: 525-533.
Strommenger B, Stevenson K, Gerlach GF (2001). Isolation and diagnostic potential of ISMav2, a novel insertion sequence-like element from Mycobacterium avium subspecies paratuberculosis. FEMS Microbiology Letters 196: 31-37.
Taylor TK, Wilks CR, McQueen DS (1981). Isolation of Mycobacterium paratuberculosis from the milk of a cow with Johne’s disease. The Veterinary Record 109: 532-533.
Van Weering H, Van Schaik G, Van der Meulen A, Waal M, Franken P, van Maanen K (2007). Diagnostic performance of pourquier ELISA for detection antibodies against Mycobacterium avium subspecies paratuberculosis in individual milk and bulk milk samples of dairy herds. Veterinary Microbiology 125: 49-58.
Vary PH, Andersen PR, Green E, Hermon-Taylor J, Mc-Fadden JJ (1990). Use of highly specific DNA probes and the polymerase chain reaction to detect Mycobacterium paratuberculosis in Johne’s disease. Journal of Clinical Microbiology 28: 933-937.
Wells S, Collins M, Faaberg K, Wees C, Tavornpanich S, Petrini K (2006). Evaluation of a rapid fecal PCR test for detection of Mycobacterium avium subsp paratuberculosis in dairy cattle. Clinical and Vaccine Immunology 13: 1125-1130.
Whittington R, Sergeant E (2001). Progress towards understanding the spread, distribution and control of Mycobacterium avium subsp. paratuberculosis in animal populations. ustralian Veterinary Journal 79: 267-278.
Whittington RJ, Taragel CA, Ottaway S, Marsh I, Seaman J, Fridriksdottir V (2001). Molecular epidemiological confirmation and circumstances of occurrence of sheep (S) strains of Mycobacterium avium subsp. paratuberculosis in cases of paratuberculosis in cattle in Australia and sheep and cattle in Iceland. Veterinary Microbiology 79: 311-322.
Yousof-beygi G, Ramin AG, Faraji-vand AR (2003). Study on the Prevalence of Subclinical Cattle Johne’s Disease in the Urmia Abattoir Archives of Razi Institute 55: 63-70.
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
مجله بیوتکنولوژی کشاورزی
دوره 4، شماره 1 - شماره پیاپی 1
سال 4 شماره 1 بهارو تابستان 1391
شهریور 1391
صفحه 83-96

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