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3 Biotech

, 8:386 | Cite as

Molecular characterization of Nosema bombycis methionine aminopeptidase 2 (MetAP2) gene and evaluation of anti-microsporidian activity of Fumagilin-B in silkworm Bombyx mori

  • Vijaya Gowri Esvaran
  • Tania Gupta
  • A. R. Narasimha Nayaka
  • Vankadara Sivaprasad
  • Kangayam M. Ponnuvel
Original Article
  • 47 Downloads

Abstract

Nosema bombycis is a spore-forming parasite causing microsporidiosis in silkworm Bombyx mori. Methionine aminopeptidase 2 (MetAP2), an essential gene of N. bombycis, is a target for the anti-microsporidian drug Fumagillin, an antibiotic derived from Aspergillus fumigatus. In this study, a 1077 bp full-length cDNA of the MetAP2 gene of N. bombycis was cloned and characterized. Furthermore, the expression study of the MetAP2 gene revealed a ubiquitous expression during all the developmental stages of the silkworm B. mori. The phylogenetic analysis of the MetAP2 gene of N. bombycis revealed the MetAP2 gene sequences to be highly conserved in nature. The present study also includes the validation of the anti-microsporidian drug Fumagillin against the MetAP2 gene of N. bombycis. The findings revealed that Fumagilin-B could also suppress the N. bombycis multiplication in the silkworm B. mori, thereby proving the therapeutic role of Fumagillin against microsporidian infection. This is the first-ever report regarding the characterization of the MetAP2 gene in the Indian isolate of N. bombycis and also towards the usage of Fumagillin in the control of microsporidiosis in B. mori.

Keywords

Microsporidiosis MetAP2 Fumagillin Nosema bombycis Anti-microsporidian 

Notes

Acknowledgements

The authors would like to thank Central Silk Board, Ministry of Textiles, Govt. of India, Bangalore, for providing the financial support and necessary facilities to carry out the research work and Dr. Ursula Da Rugna, Medivet Pharmaceuticals Ltd, High River, Canada, for the generous supply of Fumagilin-B for the research.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflicts of interest.

Ethical approval

This article does not contain any studies with human participants or animals.

Supplementary material

13205_2018_1411_MOESM1_ESM.tif (118 kb)
Fig. S1 Schematic representation of primer-binding sites and PCR amplicon size of the MetAP2 gene of Nosema bombycis (TIF 117 KB)
13205_2018_1411_MOESM2_ESM.jpg (44 kb)
Fig. S2 Nosema bombycis MetAP2 gene expression in different developmental stages of silkworm, B. mori. The β-actin gene is used as an internal control. (JPG 44 KB)
13205_2018_1411_MOESM3_ESM.jpg (173 kb)
Fig. S3 Quantification of Nosema spore load in infected and Fumagillin-treated silkworms. The spore load was determined using qRT-PCR targeting the β-tubulin gene of Nosema bombycis using genomic DNA isolated from serially diluted spores. Data are expressed as relative-fold expression to the spore load of infected sample at 24 h post-infection, which defined as 1.0-fold. ***= P< 0.001. (Infected, Infected with Nosema bombycis; FT 2 mg, FT 20 mg, and FT 120 mg infected silkworms treated with Fumagillin at concentrations of 2 mg, 20 mg, and 120 mg, respectively) (JPG 172 KB)
13205_2018_1411_MOESM4_ESM.docx (12 kb)
Supplementary material 4 (DOCX 12 KB)

References

  1. Alvarado JJ, Nemkal A, Sauder JM, Roshell M, Akiyoshi DE, Shi W, Almo SE, Weiss LM (2009) Structure of a microsporidian methionine aminopeptidase type 2 complexed with Fumagillin and TNP-470. Mol Biochem Parasitol 168:158–167.  https://doi.org/10.1016/j.molbiopara.2009.07.008 CrossRefPubMedPubMedCentralGoogle Scholar
  2. Didier PJ, Phillips JN, Kuebler DJ (2006) Anti microsporidial activities of Fumagillin, TNP-470, ovalicin, and ovalicin derivatives in vitro and in vivo. Antimicrob Agents Chemother 50:2146–2155.  https://doi.org/10.1128/AAC.00020-06 CrossRefPubMedPubMedCentralGoogle Scholar
  3. Griffith EC, Su Z, Turk BE (1997) Methionine aminopeptidase (type 2) is the common target for angiogenesis inhibitors AGM-1470 and ovalicin. Chem Biol 4:461–471CrossRefPubMedGoogle Scholar
  4. Grob U (2003) Treatment of microsporidiosis including albendazole. Parasitol Res 90:14–18.  https://doi.org/10.1007/s00436-002-0753 CrossRefGoogle Scholar
  5. Higes M, Nozal MJ, Alvaro A, Barrios L, Meana A (2011) The stability and effectiveness of Fumagillin in controlling Nosema ceranae (microsporidia) infection in honey bees (Apis mellifera) under laboratory and field conditions. Apidologie 42:364–377.  https://doi.org/10.1007/s13592-011-0003-2 CrossRefGoogle Scholar
  6. Katinka MD, Duprat S, Cornillo E, Brottier P, Wincker P, Delbac F, Alaoui HE, Peyret P, Saurin W, Gouy M, Weissenbach J, Vivarès CP (2001) Genome sequence and gene compaction of the eukaryotic parasite Encephalitozoon cuniculi. Nature 184:29–42.  https://doi.org/10.1038/35106579 CrossRefGoogle Scholar
  7. Klein C, Folkers G (2003) Understanding the selectivity of Fumagillin for the methionine aminopeptidase type II. Oncol Res 13:513–520CrossRefPubMedGoogle Scholar
  8. Kumar S (2004) MEGA3: integrated software for molecular evolutionary genetics analysis and sequence alignment. Brief Bioinform 5(2):150–163CrossRefPubMedGoogle Scholar
  9. Liu S, Widom J, Kemp CW, Crews CM, Clardy J (1998) Structure of human methionine aminopeptidase-2 complexed with Fumagillin. Science 282:1324–1327CrossRefPubMedGoogle Scholar
  10. Lowther WT, McMillen DA, Orville AM, Matthews BW (1998) The anti-angiogenic agent Fumagillin covalently modifies a conserved active-site histidine in the Escherichia coli methionineaminopeptidase. Proc Natl Acad Sci 95:12153–12157.  https://doi.org/10.1073/pnas.95.21.12153 CrossRefPubMedGoogle Scholar
  11. Molina JM, Tourneur M, Sarfati C, Gouvello AD, Gobert JG, Balkan S, Deroin F (2002) Fumagillin treatment of intestinal microsporidiosis. N Engl J Med 346:1963–1969.  https://doi.org/10.1056/NEJMoa012924 CrossRefPubMedGoogle Scholar
  12. Pandrea I, Mittleider D, Brindley PJ, Didier E, Robertson D (2005) Phylogenetic relationships of methionine aminopeptidase 2 among Encephalitozoon species and genotypes of microsporidia. Mol Biochem Parasitol 140:141–152CrossRefPubMedGoogle Scholar
  13. Reddy DK, Nagaraju J, Abraham EG (1999) Genetic characterization of the silkworm Bombyx mori by simple sequence repeat (SSR)-anchored PCR. Heredity 83:681–687.  https://doi.org/10.1046/j.1365-2540.1999.00607.x CrossRefPubMedGoogle Scholar
  14. Selvakumar T, Nataraju B, Chandrasekaran K, Sharma D (2005) Isolation of a new Microsporidian sp. (NIK-5hm) forming spores within the haemocytes of silkworm, B. mori L. Int J Ind Entomol 11:63–66Google Scholar
  15. Stevanovic J, Simeunovic P, Gajic B, Lakic N, Radovic D, Fries I, Stanimirvoc Z (2013) Characteristics of Nosema ceranae infection in Serbian honey bee colonies. Apidologie 44:522–536CrossRefGoogle Scholar
  16. Turk BE, Su Z, Liu JL (1998) Synthetic analogues of TNP-470 and ovalicin reveal a common molecular basis for inhibition of angiogenesis and immunosuppression. Bioorg Med Chem 6:1163–1169.  https://doi.org/10.1016/S0968-0896(98)00078-9 CrossRefPubMedGoogle Scholar
  17. Van den Heever JP, Thompson TS, Curtis JM, Ibrahim A, Pernal SF (2014) Fumagillin: an overview of recent scientific advances and their significance for apiculture. J Agric Food Chem 62:2728–2737CrossRefPubMedGoogle Scholar
  18. Williams GR, Shutler D, Little CM, Maclellan KL, Rogers EL (2011) The microsporidian Nosema ceranae, the antibiotic Fumagilin-B, and Western honey bee (Apis mellifera) colony strength. Apidologie 42:15–22.  https://doi.org/10.1051/apido/2010030 CrossRefGoogle Scholar
  19. Zhang H, Huang H, Cali A, Takworian P, Feng X, Zhou G, Weiss L (2005) Investigations into microsporidian methionine aminopeptidase type 2: a therapeutic target for microsporidiosis. Folia Parasitol 52:182–192.  https://doi.org/10.14411/fp.2005.023 CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2018

Authors and Affiliations

  1. 1.Genomics DivisionSeribiotech Research LaboratoryBangaloreIndia
  2. 2.Silkworm Pathology DivisionCentral Sericulture Training and Research InstituteMysoreIndia

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