Infection characteristics and classification of a microsporidian isolated from silkworm

Abstract:

[Background] Silkworm pebrine is a serious disease that has caused great damage to the silkworm production and its pathogen is the unique quarantine object that for silkworm egg production. The sources of pathogenic microsporidan of silkworm B. mori are variety and complex, which increases the difficulty of controlling pebrine in the silkworm egg production. [Objective] The present study were conducted to investigate the pathogenicity and classification feature of a microsporidan which was isolated from the silkworm mother moths (name GXM15), and its source was analysed, which will improve the classification and database of silkworm pathogenic microsporidian and will be provide reference basis for the control of silkworm pebrine in silkworm egg production. [Methods] The 50% infectious concentration (IC50) and the germinative infection rate of GXM15 were investigated with biological test methods. The spore morphology of GXM15 was observed with optical microscope, its internal structure was observed under transmission electron microscope, the SSU rRNA gene and ITS sequence of GXM15 were obtained by PCR amplification, clone and sequencing, the phylogenetic tree based on SSU rRNA sequences was constructed with the software of MEGA 5.0, and the genetic distance of SSU rRNA and ITS sequence were analyzed with software of DNAstar. [Results] The IC50 of GXM15 is 8.29×104 spores/mL, which is 2.28 times to that of Nosema bombycis (Nb) and the germinative infection rate of GXM15 is 3.60%, that is significantly lower than that of Nb. The spore of GXM15 is short oval in shape and (2.05±0.20)× (3.25±0.30) μm in size, and the volume of GXM15 is 2.19 times to that of Nb. There are binuclear and has 13 polar filament coils in the spores of GXM15, and its angles of polar filament is 45°, that is conform to the taxonomic features of Nosema. The SSU rRNA of GXM15 is grouped in the Nosema clade, and the genetic distance analysis of SSU rRNA and ITS show that GXM15 and other Nosema microsporidian are belong to different species of the same genus, GXM15 is a new microsporidian. [Conclusion] GXM15 is a new pathogenic microsporidan of Bombyx mori, and study on the pathogenicity and classification feature of GXM15 microsporidan can provide basis for prevention and control of the silkworm pebrine.