Phylogeny, Biology And Ecology Of Ophiostomatoid Fungi Associated With Dendroctonus Valens In Northern China

Biological invasions have been thought to be a significant component of globalenvironmental change. Invasive alien species can cause considerable irreversibleenvironmental and socio-economic impact at the genetic, species and ecosystem levels. InChina, a numerous well-know invasive forest pests have caused extensively economical, socialand ecological damage. Dendroctonus valens is a recent introduced pest in coniferous forestsof northern China. On the contrary to the situation in North America where the beetle hasexhibited as a secondary pest originally, the outbreak of this introduced beetle has beensuspected to be responsible for the death of more than 3 million P. tabuliformis trees in China.This abnormal phenomenon has been thought closely related to the changes in the beetleautecology and synecology resulting in an increase of aggressiveness.Bark beetles infesting conifers are commonly associated with ophiostomatoid fungi,which contribute to success or failure of beetle establishment, tree disease, and/or wooddiscoloration, and thus to economical losses. The relationships between bark beetles andophiostomatoid fungi are often thought as a mutualistic symbiosis resulting in tree mortality.The association of D. valens with ophiostomatoid fungi, and their role in the decline of P.tabuliformis in northern China remained unknown, however. The present study aimed atinventorying the ophiostomatoid species associated with the D. valens attack, and to evaluatetheir possible role in tree mortality.Fungi were recovered from and around the gallery system of D. valens attacking P.tabuliformis and P. bungeana. Over 30 genera of fungi were isolated during the sampling.Except Penicillium the ophiostomatoid fungi were isolated most frequently. In order to identifythese fungi, morphological and physiological observation, mating compatibility, multiple DNAsequences comparisons and phylogenetic analyses were conducted. The pathogenicity of someof these fungi was assessed based on inoculation on mature trees in forest stands.Seven species of ophiostomatoid fungi were isolated from and around D. valens galleries.They are Leptographium alethinum, Grosmannia koreana comb. nov. (teleomorph of L. koreanum), L. procerum, L. sinoprocerum sp. nov., L. truncatum, Pesotum aureum and P. pini.Besides of the new species, all are recorded for the first time in China.Leptographium sinoprocerum is a species closely related to L. procerum morphologicallyand phylogenetically. Although the phylogenetic analysis based on ITS&LSU sequence doesnot support the L. sinoprocerum clade confidentially (only one base difference from L.procerum), the phylogenetic analyses ofβ-tubulin gene and EF 1-αgene sequences, as well ascritical morphological comparisons, clearly evidence L. sinoprocerum as a new species. Thisspecies can be distinguished from resembling species in a combination of morphologicalcharacters.As a dominant species of ophiostomatoid fungi, Leptographium procerum was also foundassociated with D. valens in P. tabuliformis ecosystem and its distribution is across the beetleoccurrence. The identity of the fungus was confused by its extra long stipe of conidiophore,reaching up to 3900μm frequently. And confusion also partly resulted from the reference DNAsequences deposited at GenBank, where the identical sequences of multiple genes are in nameof L. terebrantis, another virulent pathogen closely associated with D. valens in North America.However, the morphological characters corresponding to the species description and theidentical sequences ofβ-tubulin gene and EF 1-αgene to an authentic strain made the decisionof L. procerum recorded in China. Because the fungus has been documented commonly inassociation with D. valens in North America, it is suspected introduced to China by the meansof D. valens vector from North America. As an important pathogen of root disease andinvolving in pine decline, the introduced L. procerum is noteworthy to inspect extensively andintensively in future.The community of ophiostomatoid fungi associated with D. valens in P. tabuliformisecosystem in northern China is positively linked to attack intensities of D. valens, whichperhaps indicates a mutualistic symbiotic relationship between the fungi and D. valens.The pathogenicity of four ophiostomatoid fungi was tested on mature P. tabuliformis treesin low-density stem inoculation. All inoculated strains caused significant necrotic lesion ofinner bark. Grosmannia koreanum and L. truncatum were more virulent than L. procerum andL. sinoprocerum, which correlated with their growth rate differences in vitro. Meanwhile, thelower vigor of the stand is also linked to stronger pathogenicity expression of the studied fungi. A preliminary study on the antagonism between the ophiostomatoid fungi and othermember strains of mycobiota associated with D. valens showed several strains effectivelyreduced the growth of the ophiostomatoid fungi in duel cultures. This suggests it is worth infuture to explore the interaction among fungal communities and potential biological controlagency to D. valens and its associated fungi.In conclusion, drier climatic events during late 90ies in northern China might have placedP. tabuliforrnis under stress and predisposed them to attack by the RTB. The subsequentdevelopment of ophiostomatoid fungi could have contributed further to decrease the resistancecapacity of the stressed P. tabuliformis to both D. valens and soil humidity level. Thealternation of attack pattern to extensive root system in China was thought as favoring thebeetle's population establishment and its aggressiveness. Because this change could also bebeneficial to increasing the pathogens' virulence, such as L. procerurn, these ophiostomatoidfungi might play more important roles in the pine mortality in northern China.