Virulence attenuation frequently occurs in in vitro culturing of pathogenic microbes. In this study, we investigated the total putative long noncoding RNAs (lncRNAs) in an aphid-obligate pathogen, Conidiobolus obscurus , and screened the differentially expressed (DE) lncRNAs and protein-coding genes involved in the virulence decline. The virulence was significantly attenuated after eight subculturing events, in which the median lethal concentration of the conidia ejected from mycelial mats relative to the bamboo aphid, Takecallis taiwanus , increased from 36.1 to 126.1 conidia mm –2 , four days after inoculation. In total, 1,252 lncRNAs were identified based on the genome-wide transcriptional analysis. By characterizing their molecular structures and expression patterns, we found that the lncRNAs possessed shorter transcripts, lower expression, and fewer exons than did protein-coding genes in C. obscurus . A total of 410 DE genes of 329 protein-coding genes and 81 lncRNAs were identified. The functional enrichment analysis showed the DE genes were enriched in peptidase activity, protein folding, autophagy, and metabolism. Moreover, target prediction analysis of the 81 lncRNAs revealed 3,111 cis -regulated and 23 trans -regulated mRNAs, while 121 DE lncRNA-mRNA pairs were possibly involved in virulence decline. Moreover, the DE lncRNA-regulated target genes mainly encoded small heat shock proteins, secretory proteins, transporters, autophagy proteins, and other stress response-related proteins. This implies that the decline in virulence regulated by lncRNAs was likely associated with the environmental stress response of C. obscurus . Hence, these findings can provide insights into the lncRNA molecules of Entomophthoromycotina, with regards to virulence regulators of entomopathogens.

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Fungi in the Entomophthorales order can cause insect disease and epizootics in nature, contributing to biological pest control in agriculture and forestry. Most Entomophthorales have narrow host ranges, limited to the arthropod family level; however, rare genomic information about host-specific fungi has been reported. Conidiation is crucial for entomopathogenic fungi to explore insect resources owing to the important roles of conidia in the infection cycle, such as dispersal, adhesion, germination, and penetration into the host hemocoel. In this study, we analyzed the whole genome sequence of the aphid-obligate pathogen Conidiobolus obscurus strain ARSEF 7217 (Entomophthoromycotina), using Nanopore technology from Biomarker Technologies (Beijing, China). The genome size was 37.6 Mb, and encoded 10,262 predicted genes, wherein 21.3% genes were putatively associated to the pathogen–host interaction. In particular, the serine protease repertoire in C. obscurus exhibited expansions in the trypsin and subtilisin classes, which play vital roles in the fungus’ pathogenicity. Differentially expressed transcriptomic patterns were analyzed in three conidiation stages (pre-conidiation, emerging conidiation, and post-conidiation), and 2915 differentially expressed genes were found to be associated with the conidiation process. Furthermore, a weighted gene co-expression network analysis showed that 772 hub genes in conidiation are mainly involved in insect cuticular component degradation, cell wall/membrane biosynthesis, MAPK signaling pathway, and transcription regulation. Our findings of the genomic and transcriptomic features of C. obscurus help reveal the molecular mechanism of the Entomophthorales pathogenicity, which will contribute to improving fungal applications in pest control.

Filamentous fungi in the order Entomophthorales are the main natural regulators of insect populations. Conidiation is crucial for entomopathogenic fungi to explore host resources due to the multifunction of conidia such as growth, infection, and stress resistance; however, the molecular mechanisms underlying the conidial functions in Entomophthorales is unknown. This study analyzed the differentially expressed transcriptomic patterns in three conidiation stages (pre-conidiation, emerging conidiation, and post-conidiation, respectively) of the aphid-obligate pathogen Conidiobolus obscurus (Entomophthoromycotina). The emerging conidiation stage vs. pre- or post- conidiation stage had 3,091 and 3,235 differentially expressed genes (DEGs), respectively, wherein 2,915 upregulated DEGs were putatively related to the conidial functions. A weighted gene co-expression network analysis showed that 772 hub genes in conidiation, which were related to cuticular component degradation, oxidative phosphorylation, ribosomal biogenesis, cell wall/membrane biosynthesis, MAPK signaling pathway, secondary metabolite biosynthesis, and other metabolic processes. This implied that the conidia of Entomophthorales have abundant transcripts with various functions to favor a quick response to the surrounding environment and effectively explore the host resources.