Comparative phylogenetic analysis of intergenic spacers and small subunit rRNA gene sequences of two microsporidian isolates from Antheraea myllita

Two microsporidian isolates extracted from infected tasar silkworms (Antheraea mylitta) collected from forest area in Deoghar district, Jharkhand, India were subjected to PCR amplification using intergenic spacer (IGS) region and small subunit rRNA (SSU-rRNA) gene specific primers followed by cloning and sequencing. The IGS and SSU-rRNA gene sequences were analysed to derive the identity of the microspoidian isolates and establish their phylogenetic relationships. The phylogenetic analysis of test isolates included assessment of variation in sequences and length of IGS and SSU-rRNA genes with reference to 16 different microsporidian sequences. The results proved that IGS sequences have more variation than SSU-rRNA gene sequences. Analysis of phylogenetic trees reveal that both test isolates have very close relationship with each other as well as with three Nosema reference species viz., N. philosamia and N. antheraea isolated from Philosamia cynthia ricini and Antheraea pernyi in China respectively, and N. disstriae isolated from Malacosma disstriae in Canada. The test microsporidian isolates revealed closer relationship with other Nosema reference strains compared to Nosema sp. (NIK-1s_mys) from India. The study results indicate that the IGS or/and SSU rRNA sequence based analysis is suitable and valuable to ascertain phylogenetic relationships between various microsporidian strains/species.Key words: Microsporidia, Antheraea mylitta, small subunit rRNA, intergenic spacer, phylogenetic relationship

Transcriptomic and proteomic profile of Aspergillus fumigatus on exposure to artemisinin

Artemisinin, an antimalarial drug, and its derivatives are reported to have antifungal activity against some fungi. We report its antifungal activity against Aspergillus fumigatus (A. fumigatus), a pathogenic filamentous fungus responsible for allergic and invasive aspergillosis in humans, and its synergistic effect in combination with itraconazole (ITC), an available antifungal drug. In order to identify its molecular targets, we further analyzed transcript and proteomic profiles of the fungus on exposure to the artemisinin. In transcriptomic analysis, a total of 745 genes were observed to be modulated on exposure to artemisinin, and some of them were confirmed by real-time polymerase chain reaction analysis. Proteomic profiles of A. fumigatus treated with artemisinin showed modulation of 175 proteins (66 upregulated and 109 downregulated) as compared to the control. Peptide mass fingerprinting led to the identification of 85 proteins — 29 upregulated and 56 downregulated, 65 of which were unique proteins. Consistent with earlier reports of molecular mechanisms of artemisinin and that of other antifungal drugs, we believe that oxidative phosphorylation pathway (64 kDa mitochondrial NADH dehydrogenase), cell wall-associated proteins and enzymes (conidial hydrophobin B protein, cell wall phiA protein, extracellular thaumatin domain protein, 1,3-beta-glucanosyltransferase Gel2) and genes involved in ergosterol biosynthesis (ERG6 and coproporphyrinogen III oxidase, HEM13) are potential targets of artemisinin for further investigations