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. 2018 Sep;8(9):386.
doi: 10.1007/s13205-018-1411-z. Epub 2018 Aug 24.

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  1 , Tania Gupta  1 , A R Narasimha Nayaka  2 , Vankadara Sivaprasad  2 , Kangayam M Ponnuvel  1
Affiliations

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 et al. 3 Biotech. 2018 Sep.

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: Anti-microsporidian; Fumagillin; MetAP2; Microsporidiosis; Nosema bombycis.

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Conflict of interest statement

The authors declare that they have no conflicts of interest.This article does not contain any studies with human participants or animals.

Figures

Fig. 1
Fig. 1
Phylogenetic tree based on amino acid sequence of Nosema bombycis MetAP2 gene compared with different microsporidian sequences retrieved from GenBank. The numbers near nodes indicate the number of bootstrap replicates; out of 100, bootstrap results greater than 95 are shown and the values lesser than 95 were removed. The scale indicates amino acid replacements per site
Fig. 2
Fig. 2
Amino acid sequence alignment of MetAP2 of eight different microsporidians: the conserved regions are shown in black boxes. The Fumagilin-B interacting amino acid residues at active site of MetAP2 are indicated with black triangles and appear to be conserved completely in the microsporidian-infecting insects
Fig. 3
Fig. 3
Expression level of Nosema bombycis MetAP2 gene in different tissues of silkworm B. mori infected with N. bombycis
Fig. 4
Fig. 4
Nosema bombycis MetAP2 gene expression pattern in infected midgut tissues at different hours of post-infection (hpi). The β-actin gene is used as an internal control
Fig. 5
Fig. 5
Effect of Fumagilin-B against the microsporidian, Nosema bombycis infection: the pathogen N. bombycis proliferation in silkworm B. mori was evaluated at 24 h interval up to 120 hpi. Fumagilin-B at three different concentrations (2, 20, and 120 mg/mL) was given and the experiment was performed under the laboratory conditions. The mRNA transcripts of pathogen genes were quantified using cDNA synthesized from total RNA of the infected and treated midgut tissues. The p < 0.01 are considered significant and are represented by asterisk. p < 0.01*, p < 0.001**, p < 0.001***. a Effect of Fumagillin-B on MetAP2 gene expression of N. bombycis. b Effect of Fumagilin-B on β-tubulin gene expression of N. bombycis. c Effect of Fumagilin-B on PTP1 gene expression of N. bombycis
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