ABSTRACT: The association between insects and fungi has evolved over millions of years and is ubiquitous in nature. This symbiotic relationship holds critical implications for both partners, the insects and the associated microbes. Numerous fungi are externally allied with bark beetles and form a close symbiosis, but the community structures of these fungi are largely unknown. In Yunnan Province in southwestern China, the beetles Tomicus yunnanensis, T. minor, and T. brevipilosus are major forest pests that cause large losses of two indigenous pines, Pinus yunnanensis and P. kesiya. In this study, we used the Illumina MiSeq PE300 platform to process 48 samples of epibiotic fungal communities pooled from 1348 beetles; the beetles were collected during both the branch- and trunk-infection sections from five locations across Yunnan Province. Considerably greater species richness was detected using high-throughput sequencing of amplified internal transcribed spacer 1 (ITS1) ribosomal DNA (rDNA) libraries than previously documented by using culture-dependent methods. In total, 1,413,600 reads were generated, and a 97% sequence-similarity cutoff produced eight phyla, 31 classes, 83 orders, 181 families, 331 genera, 471 species, and 1157 operational taxonomic units (OTUs), with 659, 621, and 609 OTUs being confined to T. yunnanensis, T. minor, and T. brevipilosus, respectively. Tomicus yunnanensis, T. minor, and T. brevipilosus had the similar OTUs richness and evenness of fungal communities in Yunnan Province; nevertheless, the two fungal community compositions associated with T. yunnanensis and T. minor were structurally similar to each other but distinct from that associated with T. brevipilosus. Lastly, the results of principal co-ordinates analysis suggested that epibiotic fungal community structures of the three Tomicus spp. were conditioned strongly by the locations and pine hosts but weakly by beetle species and infection sections. Our findings provide baseline knowledge regarding the epibiotic fungal communities of three major Tomicus spp. in southwestern China.
