تشخیص مولکولی و میکروسکوپی گونه‌های آناپلاسما در شترهای کشتار شده در استان گلستان

نوع مقاله : مقاله کامل

نویسنده

عضو هیات علمی موسسه تحقیقات واکسن و سرم‌سازی رازی، سازمان تحقیقات و آموزش کشاورزی، کرج، ایران

چکیده

آناپلاسما میکروارگانیسم درون سلولی اجباری است که در سلول‌های خونی میزبان زندگی می‌کند و منجر به بیماری و تحمیل هزینه‌های مراقبتی زیادی می‌شود. هدف از مطالعه حاضر بررسی مولکولی و میکروسکوپی حضور جنس و گونه‌های آناپلاسما در شترهای استان گلستان بود. نمونه‌های خون از شترهای کشتار شده در کشتارگاه صنعتی اینچه برون برای مدت دو سال جمع‌آوری شد. بررسی میکروسکوپی با تهیه گسترش‌های خونی رنگ‌آمیزی شده با گیمسا صورت گرفت. آزمون مولکولی با تکثیر اولیه قطعات DNA و تأیید آن‌ها با انجام nested-PCR صورت گرفت. از پرایمرهای اختصاصی بر روی ژن‌های 16S rRNA و msp4 برای تکثیر گونه‌های مختلف آناپلاسما استفاده شد. قطعات تکثیری حاصل از PCR اولیه و nested-PCR حضور جنس آناپلاسما در 33 نمونه از 90 نمونه (67/36 درصد) را مثبت نشان داد. در میان نمونه‌های حاوی جنس آناپلاسما، 22/2 درصد، 22/22 درصد و 33/13 درصد با PCR اختصاصی به‌ترتیب گونه‌های آناپلاسما مارجیناله، آناپلاسما فاگوسیتوفیلم و آناپلاسما بویس شناسایی شدند. هیچ کدام از نمونه‌ها با PCR اختصاصی حضور گونه آناپلاسما سنتراله را نشان ندادند. مشاهده میکروسکوپی اجرام آناپلاسمایی درون گلبول‌های قرمز حاکی از حضور جنس آناپلاسما در 33/33 درصد از گسترش‌های خونی بود. با مقایسه آزمون مولکولی و میکروسکوپی حساسیت، اختصاصیت و صحت برای شناسایی جنس آناپلاسما به‌ترتیب 21/21 درصد، 67/66 درصد و 50 درصد بدست آمد. نتایج مطالعه حاکی از حضور آناپلاسما در میان شترهای کشتار شده در استان گلستان بود. بر طبق نتایج، بررسی مولکولی آناپلاسما نتایج دقیق‌تری نسبت به بررسی میکروسکوپی بدست می‌دهد.

کلیدواژه‌ها

عنوان مقاله [English]

Molecular and microscopy detection of Anaplasma species in camel of Golestan province

نویسنده [English]

  • parastoo poorghafoor langroodi
Member of Razi Vaccine and Serum Research Institute, Agricultural Research, Education and Extension Organization (AREEO), Karaj, Iran
چکیده [English]

Anaplasma is an intracellular microorganism that lives in blood cells of host, causing illness and impose care costs. This study was aimed to investigate molecular and microscopic presence of genus and Anaplasma species in camels of Golestan province. Blood samples were collected from adult slaughtered camels in Inche Bourun industrial slaughterhouse for two years. Microscopic examination was performed by preparing Giemsa-stained blood smear. Molecular test was performed by primary amplification of DNA fragments and then they confirmed by performing the nested-PCR. The specific primers of 16S rRNA and msp4 genes were used to detect different species of Anaplasma. The proliferation of the primary and the nested PCR products showed that genus Anaplasma in 33 out of 90 samples (36.37 percent) were positive. Among samples including Anaplasma 2.22, 22,22 and 13.33 percentage were detected by specific PCR for A. marginale, A. phagocytophilum and A. bovis, respectively. None of the samples showed the presence of Anaplasma centrale. Microscopic observation of anaplasmic bodies in red blood cells (RBC) indicated the presence of Anaplasma genus in 33.33 percent of blood smears. By comparing molecular and microscopic tests, the sensitivity, specificity and accuracy to identify the genus Anaplasma were 21.21, 66.67 and 50 percent, respectively. The results of this study showed that Anaplasma was present among camels in Golestan province. According to the results, molecular study of Anaplasma yields more accurate results than microscopic examination.

کلیدواژه‌ها [English]

  • Anaplama
  • 16S rRNA gene
  • msp4 gene
  • Blood smear
  • Golestan

مراجع

1- Abdel-Shafy, S., N. A. T. Allam and M. S. Mahmoud. 2016. Molecular Description of Anaplasma Biodiversity regarding 16SrDNA, msp4, hsp60, and rpoB Profiles in Ixodid Ticks Infesting Animals from Some Egyptian Provinces. Bull NRC 41: 121-136.
2- AJ. Al-Gharban, H. and H. SR. AL-Taee. 2016. Seroclinical Diagnosis of Anaplasma Marginale Bacteria in Carrier Arabian One-Humped Camels. Basrah Journal of Veterinary Research 15: 346-359.
3- Alsaad, K. M. 2009. Clinical, hematological and biochemical studies of anaplasmosis in Arabian one-humped camels (Camelus dromedarius). Journal of Animal and Veterinary Advances 8: 2106-2109.
4- Bala, A. E., A. D. Abakar, M. S. Mohammed and M. A. Abbas. 2018. Prevalence of Trypanosoma evansi in camels in four states of Great Butana, Sudan. Journal of Entomology and Zoology Studies 6: 1207-1211.
5- Bastos, A. D. S., O. B. Mohammed, N. C. Bennett, C. Petevinos and A. N. Alagaili. 2015. Molecular detection of novel Anaplasmataceae closely related to Anaplasma platys and Ehrlichia canis in the dromedary camel (Camelus dromedarius). Veterinary Microbiology 179: 310-314.
6- Carelli, G., N. Decaro, A. Lorusso, G. Elia, E. Lorusso, V. Mari, L. Ceci and C. Buonavoglia. 2007. Detection and quantification of Anaplasma marginale DNA in blood samples of cattle by real-time PCR. Veterinary Microbiology 124: 107-114.
7- de Echaide, S. T., M. F. Bono, C. Lugaresi, N. Aguirre, A. Mangold, R. Moretta, M. Farber and C. Mondillo. 2005. Detection of antibodies against Anaplasma marginale in milk using a recombinant MSP5 indirect ELISA. Veterinary Microbiology 106: 287-292.
8- Dumler, J. S., A. F. Barbet, C. P. Bekker, G. A. Dasch, G. H. Palmer, S. C. Ray, Y. Rikihisa and F. R. Rurangirwa. 2001. Reorganization of genera in the families Rickettsiaceae and Anaplasmataceae in the order Rickettsiales: unification of some species of Ehrlichia with Anaplasma, Cowdria with Ehrlichia and Ehrlichia with Neorickettsia, descriptions of six new species combinations and designation of Ehrlichia equi and 'HGE agent' as subjective synonyms of Ehrlichia phagocytophila. International Journal of Systematic and Evolutionary Microbiology 51: 2145-2165.
9- El-Naga, T. R. A. and S. Barghash. 2016. Blood Parasites in Camels (Camelus dromedarius) in Northern West Coast of Egypt. Journal of Bacteriology and Parasitology 7.
10- Ge, N. L., K. M. Kocan, G. L. Murphy and E. F. Blouin. 1995. Detection of Anaplasma marginale DNA in bovine erythrocytes by slot-blot and in situ hybridization with a PCR-mediated digoxigenin-labeled DNA probe. Journal of Veterinary Diagnostic Investigation 7: 465-472.
11- Ghashghaei, O., S. R. Nourollahi Fard, M. Khalili and H. Sharifi. 2016. Abundance and associated risk factors of ixodid ticks (Acari: Ixodidae) collected from one-humped camels ( Camelus dromedarius ) in Sistan and Balouchestan region, southeast of Iran. Persian Journal of Acarology 5.
12- Ghazvinian, K. and T. Khodaiean. 2016. Anaplasmosis among Camels in Iran and Observation of Abnormalities in Infected Blood Films. International Journal of Animal and Veterinary Sciences 10: 475-478.
13- Ibrahim, A. M., A. A. H. Kadle and H. S. Nyingilili. 2017. Microscopic and Molecular Detection of Camel Piroplasmosis in Gadarif State, Sudan. Veterinary Medicine International 2017: 5.
14- Ismael, A. and A. A. A. Swelum. 2016. First evidence of natural anaplasmosis in Camelus dromedarius in Saudi Arabia. Journal of Camel Practice and Research 23: 1-6.
15- Jacobson, R. H. 1996. rinciples of validation of diagnostic assays for infectious disease. In manual of standards for diagnostic tests and vaccines. Office International des Epizooties. Paris.
16- Kocan, K. M., J. de la Fuente, E. F. Blouin and J. C. Garcia-Garcia. 2004. Anaplasma marginale (Rickettsiales: Anaplasmataceae): recent advances in defining host-pathogen adaptations of a tick-borne rickettsia. Parasitology 129 Suppl: S285-300.
17- Kocan, K. M., J. de la Fuente and A. Cabezas-Cruz. 2015. The genus Anaplasma: new challenges after reclassification. Revue Scientifique et Technique 34: 577-586.
18- Lbacha, A. H., Z. Zouagui, S. Alali, A. Rhalem, E. Petit, M. J. Ducrotoy, H. J. Boulouis and R. Maillard. 2017. "Candidatus anaplasma camelii" in one-humped camels (Camelus dromedarius) in Morocco: a novel and emerging anaplasma species? Infect Dis Poverty 6: 1.
19- Liu, Z., J. Luo, Q. Bai, M. Ma, G. Guan and H. Yin. 2005. Amplification of 16S rRNA genes of Anaplasma species in China for phylogenetic analysis. Veterinary Microbiology 107: 145-148.
20- Martin, S. W., A. H. Meek and P. Willeberg. 1987. Veterinary Epidemiology: Principles and Methods. Iowa State University Press, Ames IA.
21- McQuiston, J. H., C. L. McCall and W. L. Nicholson. 2003. Ehrlichiosis and related infections. Journal of the American Veterinary Medical Association 223: 1750-1756.
22- Molad, T., M. L. Mazuz, L. Fleiderovitz, L. Fish, I. Savitsky, Y. Krigel, B. Leibovitz, J. Molloy, F. Jongejan and V. Shkap. 2006. Molecular and serological detection of A. centrale- and A. marginale-infected cattle grazing within an endemic area. Veterinary Microbiology 113: 55-62.
23- Monteverde, G. 1937. Anaplasmosi nei cammelli in Cirenaica. Clinic Veterinary (Milano) 60: 73-77.
24- Noaman, V. 2016. A review of anaplasmosis and the prevalence of Anaplasma marginale in cattle in Iran and the world. Veterinary Researches and Biological Products 30: 2-15.
25- Noaman, V. 2018. Molecular Detection of Novel Genetic Variants Associated to Anaplasma ovis among Dromedary Camels in Iran. Arch Razi Inst 73: 11-18.
26- Noaman, V. 2019. A review on Anaplasma phagocytophilum as a zoonotic agent: review article (In Farsi). Tehran University Medical Journal 76: 778-785.
27- Noaman, V. and P. Shayan. 2019. Molecular Detection of Anaplasma Phagocytophilum in Carrier Cattle of Iran - First Documented Report. Iranian Journal of Microbiology 1: 37-42.
28- Noaman, V., P. Shayan and A. H. Shahmoradi. 2009. Detection of Anaplasma ovis based on 16S rRNA gene by PCR-RFLP in sheep from central part of Iran (In Farsi). Journal of Veterinary Laboratory Research 1: 25-35.
29- Poorghafoor-Langroodi, P., V. Noaman, S. Esmaeilkhanian, A. H. Shahmoradi, M. R. Heidari, A. R. Hatami, A. Rahimi and A. A. Fouisi-Yousofi. 2015. Molecular detection of Anaplasma species in cattle and sheep of Golestan (In Farsi). Final Report of Research Project Department of Education and Research. Golestan Research Center and Education of Agriculture and Natural Resources.
30- Rabana, J. L., H. A. Kumshe, J. Kamani, G. Hafsat, U. A. Turaki and H. K. Dilli. 2011. Effects of Parasitic Infections on Erythrocyte Indices of Camels in Nigeria. Veterinary Research Forum 2: 59-63.
31- Ranjbar Bahadori, S., M. Aghaei Ghamsari and A. Safdari. 2015. Investigation of blood parasites of slaughtered camels in Mashhad abattoir (In Farsi). 15th Iranian Veterinary Congress. Tehran.
32- Sazmand, A., J. Harl, B. Eigner, A. Hodzic, R. Beck, S. Hekmatimoghaddam, M. Mirzaei, H. P. Fuehrer and A. Joachim. 2019. Vector-borne bacteria in blood of camels in Iran: New data and literature review. Comparative Immunology, Microbiology and Infectious Diseases 65: 48-53.
33- Setayeshgar, F. 2019. Opportunities for camel breeding in Golestan (In Farsi). Hamshahri Online.
34- Sharifiyazdi, H., S. Jafari, M. Ghane, S. Nazifi and A. Sanati. 2017. Molecular investigation of Anaplasma and Ehrlichia natural infections in the dromedary camel (Camelus dromedarius) in Iran. Comparative Clinical Pathology 26: 99-103.
35- Sudan, V., R. L. Sharma and M. K. Borah. 2014. Subclinical anaplasmosis in camel (Camelus dromedarius) and its successful therapeutic management. Journal of Parasitic Diseases 38: 163-165.
36- Wahba, A. A., S. G. Ghattas, R. S. Fadly and K. A. M. Shokier. 2017. Serological and microscopical diagnosis of anaplasmosis in some farm animals. Animal Health Research Journal 5: 68-75.

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  • تاریخ دریافت: 03 شهریور 1398
  • تاریخ بازنگری: 29 مهر 1398
  • تاریخ پذیرش: 09 آبان 1398

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