تعیین فراوانی آلل های تیپ های آمیزشی در جمعیت های آلترناریا آلترناتا مرکبات جنوب کشور با روش واکنش زنجیره ای پلی مراز
محورهای موضوعی : میکروب شناسی گیاهیعلیرضا نیازمند 1 , شهاب حاج منصور 2
1 - استادیار، گروه بیماری شناسی گیاهی، واحد جهرم، دانشگاه آزاد اسلامی، جهرم
2 - کارشناس ارشد، گروه بیماری شناسی
گیاهی، واحد علوم و تحقیقات تهران، دانشگاه آزاد اسلامی، تهران
کلید واژه: واکنش زنجیره ای پلی مراز, تیپ آمیزشی, تولید مثل جنسی, ژن MAT1-1, ژن MAT1-2,
چکیده مقاله :
سابقه و هدف: تاکنون فرم جنسی برای گونه آلترناریا آلترناتا گزارش نشده است. اما سطوح بالایی از تنوع ژنتیکی و ریخت شناسی مشاهده شده در بین جمعیت های این قارچ بیانگر وجود مکانیسم های ناشناخته و مرموز تولید مثل جنسی است. در قارچ های آسکومیستی مشابه با این قارچ، تولید مثل جنسی توسط یک ژن گاه منفرد به نام MAT1 کنترل می شود که دارای دو آلل MAT1-1 و MAT1-2 می باشند. این مطالعه با هدف تعیین فراوانی و نحوه پراکنش آلل های ژن MAT1 در جمعیت قارچ آلترناریا آلترناتا مرکبات جنوب کشور انجام شد. مواد و روش ها: در این مطالعه مقطعی- توصیفی، استخراج DNA با روش CTAB، از ریسه های جوان نمونه های برگی 45 جدایه قارچ آلترناریا آلترناتا که قبلاً از گونه ها و ارقام مختلف باغات مرکبات جنوب کشور جمع آوری و خالص سازی شده بودند، انجام گردید. به منظور تعیین تیپ های آمیزشی، از آغازگرهای اختصاصی برای تکثیر ناحیه های MAT1-1 و MAT1-2 در واکنش زنجیره ای پلی مراز استفاده شد. یافته ها: جدایه های تیپ های آمیزشی MAT1-1 و MAT1-2 به ترتیب تکثیر قطعات 642 و 882 جفت بازی را داشتند. به طور کلی تیپ های آمیزشی در این ناحیه از فراوانی مساوی برخوردار نبودند. در مجموع، 16 جدایه دارای تیپ آمیزشی MAT1-1 و 29 جدایه دارای تیپ آمیزشی MAT1-2 بودند. فراوانی های تیپ های آمیزشی در شهرستان های مختلف و در گونه های مختلف مرکبات متفاوت بود. نتیجه گیری: نتایج نشان داد که در برخی از مناطق جنوب کشور و در برخی از میزبان ها عوامل ناشناخته ای در القای مرحله جنسی قارچ آلترناریا آلترناتا دخالت دارند.
Background & Objectives: Sexual stage of Alternaria alternata has not been discovered yet. However, high levels of phenotypic and genetic diversity have been reported in different populations of A. alternata. In other Ascomycetes similar to this species, sexual reproduction is controlled by a single locus, named MAT1, with two alleles MAT1-1 and MAT 1-2. The aims of this research were identification of frequency and distribution of both alleles of MAT1in A. alternata population of citrus collected from South of Iran. Materials & Methods: In this cross-sectional study, DNA was extracted from young mycelia of 45 isolates of A. alternata that previously were collected and isolated directly from different species and cultivars of the citrus plants showing leaf spot disease symptoms in different orchards of Southern region of Iran using CTAB method. Mating types were determined using specific primer sets for MAT1-1 and MAT1-2 idiomorphs and PCR assay. Results: A 642 bp amplicon and a 882 bp amplicon were amplified pecificlly for Mat1-1Mat1-2 isolates, respectively. No equal frequency of both mating type idiomorphs were observed in studied samples. Totally 16 and 29 isolates showed the presence of MAT1-1 and MAT1-2 idiomorphs, respectively. Unequal frequency of both idiomorphs was observed in the citrus species and cultivars collected from different regions. Conclusion: It seems that unknown factors are involved in sexual induction of A. alternata in some regions and citrus hosts in South of Iran.
1. Timmer LW, Solel Z, Gottwald TR, Ibanez AM, Zitko SE. Environmental factors affecting production, release, and field populations of conidia of Alternaria alternata, the cause of brown spot of citrus. Phytopathol. 1998; 88(11): 1218-1223.
2. Metzenberg RL, Glass NL. Mating type and mating strategies in Neurospora. Bioessays. 1990; 12(2): 53-59.
3. Sun Y, Corcoran P, Menkis A, Whittle, CA, Andersson SGE, Johannesson H. Large scale introgression shapes the evolution of the mating-type chromosomes of the filamentous ascomycete Neurospora tetrasperma. PLoS Genet. 2012; 8(7): 1002820.
4. Thrall PH, Laine AL, Ravensdale M, Nemri A, Dodds PN, Barrett LG, Burdon JJ. Rapid genetic change underpins antagonistic coevolution in a natural host pathogen metapopulation. Ecol Lett 2012; 15(5): 425-435.
5. Stewart JE, Timmer LW, Lawrence CB, Pryor BM, Peever TL. Discord between morphological and phylogenetic species boundaries: incomplete lineage sorting and recombination results in fuzzy species boundaries in an asexual fungal pathogen. BMC Evol Biol. 2014; 14(1): 38.
6. Simmons EG. Alternaria themes and variations (22-26). Mycotaxon 1986; 25(1): 287-308.
7. Hawksworth DL, Kirk PM, Sutton BC, Pegler DN. Ainsworth and Bisby's dictionary of the fungi. Rev I Med Trop. 1996; 38(4): 272-272.
8. Berbee ML, Payne BP, Zhang G, Roberts RG, Turgeon BG. Shared ITS DNA substitutions in isolates of opposite mating type reveal a recombining history for three presumed asexual species in the filamentous ascomycete genus Alternaria. Mycol Res. 2003; 107(02): 169-182.
9. Chelkowski J, Visconti A. Alternaria: biology, plant diseases and metabolites: Elsevier Science Publishers; 1992.
10. Bock CH, Thrall PH, Burdon JJ. Genetic structure of populations of Alternaria brassicicola suggests the occurrence of sexual recombination. Mycol Res. 2005; 109(02): 227-236.
11. Linde CC, Liles JA, Thrall PH. Expansion of genetic diversity in randomly mating founder populations of Alternaria brassicicola infecting Cakile maritima in Australia. Appl Environ Microb 2010; .1954-1946 :(6) 76
12. O‘Gorman CM, Fuller HT, Dyer PS. Discovery of a sexual cycle in the opportunistic fungal pathogen Aspergillus fumigatus. Nature. 2009; 457(7228): 471-474.
13. Peever TL, Su G, Carpenter-Boggs L, Timmer LW. Molecular systematics of citrus-associated Alternaria species. Mycologia. 2004; 96(1): 119-134.
14. Peever TL, Carpenter-Boggs L, Timmer LW, Carris LM, Bhatia A. Citrus black rot is caused by phylogenetically distinct lineages of Alternaria alternata. Phytopathol. 2005; 95(5): 512-518.
15. Peever TL, Olsen L, Ibanez A, Timmer LW. Genetic differentiation and host specificity among populations of Alternaria spp. causing brown spot of grapefruit and tangerine x grapefruit hybrids in Florida. Phytopathol. 2000; 90(4): 407-414.
16. Peever TL, Canihos Y, Olsen L, Ibanez A, Liu YC, Timmer LW. Population genetic structure and host specificity of Alternaria spp. causing brown spot of Minneola tangelo and rough lemon in Florida. Phytopathol. 1999; 89(10): 851-860.
17. Sommerhalder RJ, McDonald BA, Mascher F, Zhan J. Sexual recombinants make a significant contribution to epidemics caused by the wheat pathogen Phaeosphaeria nodorum. Phytopathol. 2010; 100(9): 855-862.
18. Doyle J, Doyle JL. Genomic plant DNA preparation from fresh tissue-CTAB method. Phytochem Bull. 1987; 19(11): 11-15.
19. Cullings KW. Design and testing of a plant- specific PCR primer for ecological and evolutionary studies. Mol Ecol. 1992; 1(4): 233-240.
20. Stewart JE, Thomas KA, Lawrence CB, Dang HA, Pryor BM, Timmer LM, Peever TL. Signatures of recombination in clonal lineages of the citrus brown spot pathogen, Alternaria alternata sensu lato. Phytopathol. 2013; 103(7): 741-749.
21. Zheng HH, Zhao J, Wang TY, Wu XH. Characterization of Alternaria species associated with potato foliar diseases in China. Plant Pathol. 2014; 64(2): 425-433.
22. Goodwin SB, Waalwijk C, Kema GHJ, Cavaletto JR, Zhang G. Cloning and analysis of the mating-type idiomorphs from the barley pathogen Septoria passerinii. Mol Genet Genomics 2003; 269(1): 1-12.
23. Bakhshi M, Arzanlou M, Babai Ahari A. Determining of mating type alleles in Cercospora beticola, the causal agent of Cercospora leaf spot on sugar beet using specific primers and induction of sexual phase under laboratory and greenhouse conditions. J Appl Res Plant. 2012; 1(1): 15-27.
24. Groenewald M, Groenewald JZ, Harrington TC, Abeln ECA, Crous PW. Mating type gene analysis in apparently asexual Cercospora species is suggestive of cryptic sex. Fungal Gene Biol. 2006; 43(12): 813-825.
25. Ghasemloee M, Niazmand AR. Genetic Diversity of Alternaria alternata Associated with Citrus spp. in Southern Iran Based on RAPD-PCR. J Pure Appl Microb. 2015; 9(1): 473-482.
26. Morris PF, Connolly MS, St Clair DA. Genetic diversity of Alternaria alternata isolated from tomato in California assessed using RAPDs. Mycol Res. 2000; 104(03): 286-292.
27. Cubry P, De Bellis F, Avia K. An initial assessment of linkage disequilibrium (LD) in coffee trees: LD patterns in groups of Coffea canephora Pierre using microsatellite analysis. BMC genomics. 2013; 14(1): 1.
28. Guo L-D, Xu L, Zheng WH, Hyde KD. Genetic variation of Alternaria alternata, an endophytic fungus isolated from Pinus tabulaeformis as determined by random amplified microsatellites (RAMS). Fungal Divers. 2004; 16: 53-65.
29. Pegg KG. Studies of a strain of Alternaria citri Pierce, the causal organism of brown spot of Emperor mandarin. Queensland J Agric Animal Sci. 1996; 23: 15-28.
30. Kohmoto K, Scheffer RP, Whiteside JO. Host-selective toxins from Alternaria citri. Phytopathol. 1979; 69(6): 667-671.
31. Mmbaga MT, Shi A, Kim M-S. Identification of Alternaria alternata as a causal agent for leaf blight in Syringa species. Plant Pathol J. 2011; 27(2): 120-127.
32. Debuchy R, Turgeon BG. Mating-type structure, evolution, and function in Euascomycetes. In: Growth, Differentiation and Sexuality: Springer; 2006: 293-323.
33. Leslie JF, Klein KK. Female fertility and mating type effects on effective population size and evolution in filamentous fungi. Genetics 1996; 144(2): 557-567.
34. Stewart JE, Timmer LW, Lawrence CB, Pryor BM, Peever TL. Discord between morphological and phylogenetic species boundaries: incomplete lineage sorting and recombination results in fuzzy species boundaries in an asexual fungal pathogen. BMC Evol Biol. 2014; 14(1): 38.
35. Stewart JE, Kawabe M, Abdo Z, Arie T, Peever TL. Contrasting codon usage patterns and purifying selection at the mating locus in putatively asexual Alternaria fungal species. PloS one. 2011; 6(5): e20083.
36. Meng JW, Zhu W, He MH, Wu EJ, Yang LN, Shang LP, Zhan J. High genotype diversity and lack of isolation by distance in the Alternaria solani populations from China. Plant Pathol 2014; 64(2): 434-441.
37. Meng JW, Zhu W, He MH, Wu, EJ, Duan GH, Xie YK, Jin YJ, Yang LN, Shang LP, Zhan J. Population genetic analysis reveals cryptic sex in the phytopathogenic fungus Alternaria alternata. Scientific Rep. 2015; 5: 18250.
_||_1. Timmer LW, Solel Z, Gottwald TR, Ibanez AM, Zitko SE. Environmental factors affecting production, release, and field populations of conidia of Alternaria alternata, the cause of brown spot of citrus. Phytopathol. 1998; 88(11): 1218-1223.
2. Metzenberg RL, Glass NL. Mating type and mating strategies in Neurospora. Bioessays. 1990; 12(2): 53-59.
3. Sun Y, Corcoran P, Menkis A, Whittle, CA, Andersson SGE, Johannesson H. Large scale introgression shapes the evolution of the mating-type chromosomes of the filamentous ascomycete Neurospora tetrasperma. PLoS Genet. 2012; 8(7): 1002820.
4. Thrall PH, Laine AL, Ravensdale M, Nemri A, Dodds PN, Barrett LG, Burdon JJ. Rapid genetic change underpins antagonistic coevolution in a natural host pathogen metapopulation. Ecol Lett 2012; 15(5): 425-435.
5. Stewart JE, Timmer LW, Lawrence CB, Pryor BM, Peever TL. Discord between morphological and phylogenetic species boundaries: incomplete lineage sorting and recombination results in fuzzy species boundaries in an asexual fungal pathogen. BMC Evol Biol. 2014; 14(1): 38.
6. Simmons EG. Alternaria themes and variations (22-26). Mycotaxon 1986; 25(1): 287-308.
7. Hawksworth DL, Kirk PM, Sutton BC, Pegler DN. Ainsworth and Bisby's dictionary of the fungi. Rev I Med Trop. 1996; 38(4): 272-272.
8. Berbee ML, Payne BP, Zhang G, Roberts RG, Turgeon BG. Shared ITS DNA substitutions in isolates of opposite mating type reveal a recombining history for three presumed asexual species in the filamentous ascomycete genus Alternaria. Mycol Res. 2003; 107(02): 169-182.
9. Chelkowski J, Visconti A. Alternaria: biology, plant diseases and metabolites: Elsevier Science Publishers; 1992.
10. Bock CH, Thrall PH, Burdon JJ. Genetic structure of populations of Alternaria brassicicola suggests the occurrence of sexual recombination. Mycol Res. 2005; 109(02): 227-236.
11. Linde CC, Liles JA, Thrall PH. Expansion of genetic diversity in randomly mating founder populations of Alternaria brassicicola infecting Cakile maritima in Australia. Appl Environ Microb 2010; .1954-1946 :(6) 76
12. O‘Gorman CM, Fuller HT, Dyer PS. Discovery of a sexual cycle in the opportunistic fungal pathogen Aspergillus fumigatus. Nature. 2009; 457(7228): 471-474.
13. Peever TL, Su G, Carpenter-Boggs L, Timmer LW. Molecular systematics of citrus-associated Alternaria species. Mycologia. 2004; 96(1): 119-134.
14. Peever TL, Carpenter-Boggs L, Timmer LW, Carris LM, Bhatia A. Citrus black rot is caused by phylogenetically distinct lineages of Alternaria alternata. Phytopathol. 2005; 95(5): 512-518.
15. Peever TL, Olsen L, Ibanez A, Timmer LW. Genetic differentiation and host specificity among populations of Alternaria spp. causing brown spot of grapefruit and tangerine x grapefruit hybrids in Florida. Phytopathol. 2000; 90(4): 407-414.
16. Peever TL, Canihos Y, Olsen L, Ibanez A, Liu YC, Timmer LW. Population genetic structure and host specificity of Alternaria spp. causing brown spot of Minneola tangelo and rough lemon in Florida. Phytopathol. 1999; 89(10): 851-860.
17. Sommerhalder RJ, McDonald BA, Mascher F, Zhan J. Sexual recombinants make a significant contribution to epidemics caused by the wheat pathogen Phaeosphaeria nodorum. Phytopathol. 2010; 100(9): 855-862.
18. Doyle J, Doyle JL. Genomic plant DNA preparation from fresh tissue-CTAB method. Phytochem Bull. 1987; 19(11): 11-15.
19. Cullings KW. Design and testing of a plant- specific PCR primer for ecological and evolutionary studies. Mol Ecol. 1992; 1(4): 233-240.
20. Stewart JE, Thomas KA, Lawrence CB, Dang HA, Pryor BM, Timmer LM, Peever TL. Signatures of recombination in clonal lineages of the citrus brown spot pathogen, Alternaria alternata sensu lato. Phytopathol. 2013; 103(7): 741-749.
21. Zheng HH, Zhao J, Wang TY, Wu XH. Characterization of Alternaria species associated with potato foliar diseases in China. Plant Pathol. 2014; 64(2): 425-433.
22. Goodwin SB, Waalwijk C, Kema GHJ, Cavaletto JR, Zhang G. Cloning and analysis of the mating-type idiomorphs from the barley pathogen Septoria passerinii. Mol Genet Genomics 2003; 269(1): 1-12.
23. Bakhshi M, Arzanlou M, Babai Ahari A. Determining of mating type alleles in Cercospora beticola, the causal agent of Cercospora leaf spot on sugar beet using specific primers and induction of sexual phase under laboratory and greenhouse conditions. J Appl Res Plant. 2012; 1(1): 15-27.
24. Groenewald M, Groenewald JZ, Harrington TC, Abeln ECA, Crous PW. Mating type gene analysis in apparently asexual Cercospora species is suggestive of cryptic sex. Fungal Gene Biol. 2006; 43(12): 813-825.
25. Ghasemloee M, Niazmand AR. Genetic Diversity of Alternaria alternata Associated with Citrus spp. in Southern Iran Based on RAPD-PCR. J Pure Appl Microb. 2015; 9(1): 473-482.
26. Morris PF, Connolly MS, St Clair DA. Genetic diversity of Alternaria alternata isolated from tomato in California assessed using RAPDs. Mycol Res. 2000; 104(03): 286-292.
27. Cubry P, De Bellis F, Avia K. An initial assessment of linkage disequilibrium (LD) in coffee trees: LD patterns in groups of Coffea canephora Pierre using microsatellite analysis. BMC genomics. 2013; 14(1): 1.
28. Guo L-D, Xu L, Zheng WH, Hyde KD. Genetic variation of Alternaria alternata, an endophytic fungus isolated from Pinus tabulaeformis as determined by random amplified microsatellites (RAMS). Fungal Divers. 2004; 16: 53-65.
29. Pegg KG. Studies of a strain of Alternaria citri Pierce, the causal organism of brown spot of Emperor mandarin. Queensland J Agric Animal Sci. 1996; 23: 15-28.
30. Kohmoto K, Scheffer RP, Whiteside JO. Host-selective toxins from Alternaria citri. Phytopathol. 1979; 69(6): 667-671.
31. Mmbaga MT, Shi A, Kim M-S. Identification of Alternaria alternata as a causal agent for leaf blight in Syringa species. Plant Pathol J. 2011; 27(2): 120-127.
32. Debuchy R, Turgeon BG. Mating-type structure, evolution, and function in Euascomycetes. In: Growth, Differentiation and Sexuality: Springer; 2006: 293-323.
33. Leslie JF, Klein KK. Female fertility and mating type effects on effective population size and evolution in filamentous fungi. Genetics 1996; 144(2): 557-567.
34. Stewart JE, Timmer LW, Lawrence CB, Pryor BM, Peever TL. Discord between morphological and phylogenetic species boundaries: incomplete lineage sorting and recombination results in fuzzy species boundaries in an asexual fungal pathogen. BMC Evol Biol. 2014; 14(1): 38.
35. Stewart JE, Kawabe M, Abdo Z, Arie T, Peever TL. Contrasting codon usage patterns and purifying selection at the mating locus in putatively asexual Alternaria fungal species. PloS one. 2011; 6(5): e20083.
36. Meng JW, Zhu W, He MH, Wu EJ, Yang LN, Shang LP, Zhan J. High genotype diversity and lack of isolation by distance in the Alternaria solani populations from China. Plant Pathol 2014; 64(2): 434-441.
37. Meng JW, Zhu W, He MH, Wu, EJ, Duan GH, Xie YK, Jin YJ, Yang LN, Shang LP, Zhan J. Population genetic analysis reveals cryptic sex in the phytopathogenic fungus Alternaria alternata. Scientific Rep. 2015; 5: 18250.
