Abstract
The ascomycete Leptographium qinlingensis is one of the major externally living fungal associates of Dendroctonus armandi, which can kill host trees when it invades Chinese white pine (Pinus armandii). We identified and phylogenetically analysed the cytochrome P450 (CYP) genes in the transcriptome of L. qinlingensis. Furthermore, the expression profiles of six CYPs in the mycelium of L. qinlingensis grown on different media or treated with terpenoids were detected, as well as their growth rates on different nutritional media and inhibition by terpenoids. The CYP evolution predicted that most of the CYPs occurred in a putative common ancestor shared between L. qinlingensis and Grosmannia clavigera. The growth rates and inhibition test result shows that L. qinlingensis has more similarity with G. clavigera, which can retrieve nutrition from pine wood and utilize monoterpenes as the sole carbon source. CYP582C and CYP52Z4 of L. qinlingensis might be involved in the metabolism of fatty acids and detoxification of terpenes and phenolics after the analysis of their transcription levels with different treatments.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Altschul S (1997) Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucl Acids Res 25:3389–3402. https://doi.org/10.1093/nar/25.17.3389
Ayres M, Wilkens R, Ruel J, Lombardero M (2000) Nitrogen budgets of phloem feeding bark beetles with and without symbiotic fungi. Ecology 8:2198–2210. https://doi.org/10.1890/0012-9658(2000)081[2198:NBOPFB]2.0.CO;2
Balibar CJ, Walsh CT (2006) GliP, a multimodular nonribosomal peptide synthetase in Aspergillus fumigatus, makes the diketopiperazine scaffold of gliotoxin. Biochem J 45:15029–15038. https://doi.org/10.1021/bi061845b
Bentz BJ, Six DL (2006) Ergosterol content of fungi associated with Dendroctonus ponderosae and Dendroctonus rufipennis (Coleoptera: Curculionidae, Scolytinae). Ann Entomol Soc Am 99:189–194. https://doi.org/10.1603/0013-8746(2006)099[0189:ECOFAW]2.0.CO;2
Boone CK, Aukema BH, Bohlmann J, Carroll AL, Raffa KF (2011) Efficacy of tree defense physiology varies with bark beetle population density: a basis for positive feedback in eruptive species. Can J for Res 41:1174–1188. https://doi.org/10.1139/x11-041
Briones-Roblero CI, Rodríguez-Díaz R, Santiago-Cruz JA, Zúñiga G, Rivera-Orduña FN (2017) Degradation capacities of bacteria and yeasts isolated from the gut of Dendroctonus rhizophagus (Curculionidae: Scolytinae). Folia Microbiol 62:1–9. https://doi.org/10.1007/s12223-016-0469-4
Cano-Ramírez C, Santiago-Hernández A, Rivera-Orduña FN, García-Huante Y, Zúñiga G, Hidalgo-Lara ME (2016) Expression, purification and characterization of an endoglucanase from Serratia proteamaculans CDBB-1961, isolated from the gut of Dendroctonus adjunctus (Coleoptera: Scolytinae). AMB Express 6:63. https://doi.org/10.1186/s13568-016-0233-9
Chakraborty A, Roy A (2021) Microbial influence on plant-insect interaction. In: Singh A, Singh IK (eds) Plant-pest interactions: from molecular mechanisms to chemical ecology. Springer, Singapore, pp 337–363
Chen H, Tang M (2002) Microstructure of blue-stain fungi (Leptographium terebrantis) associated with Dendroctonus. armandi in the xylem tissue of Pinus armandi. Acta Bot Boreal Occident Sin 22:1391–1395
Chen H, Tang M (2007) Spatial and temporal dynamics of bark beetles in Chinese white pine in Qinling Mountains of Shaanxi Province, China. Environ Entomol 36:1124–1130. https://doi.org/10.1093/ee/36.5.1124
Chen H, Tang M, Gao JM, Chen X, Li ZB (2006) Changes in the compositions of volatile monoterpenes and sesquiterpenes of Pinus armandi, P. tabulaeformis and P. bungeana in northwest China. Chem Nat Comp 42:430–433. https://doi.org/10.1007/s10600-006-0208-1
Chen H, Tang M, Liu L, Wang HZ, Li ZB (2007) Cytochemical localization of acid phosphatase activity in tissues of Pinus armandi infected by Leptographium qinlingensis. Symbiosis 43(2):65–70
Chen H, Li Z, Tang M (2010) Laboratory evaluation of flight activity of Dendroctonus armandi (Coleoptera: Curculionidae: Scolytinae). Can Entomol 142:378–387. https://doi.org/10.4039/n10-018
Cheng C, Xu L, Xu D, Lou Q, Lu M, Sun J (2016) Does cryptic microbiota mitigate pine resistance to an invasive beetle-fungus complex? Implications for Invasion Potential. Sci Rep 6:33110. https://doi.org/10.1038/srep33110
Clark EL, Huber DPW, Carroll AL (2012) The legacy of attack: implications of high phloem resin monoterpene levels in lodgepole pines following mass attack by mountain pine beetle, Dendroctonus ponderosae Hopkins. Environ Entomol 41:392–398. https://doi.org/10.1603/EN11295
Dai L, Ma M, Wang C, Shi Q, Zhang R, Chen H (2015a) Cytochrome P450s from the Chinese white pine beetle, Dendroctonus armandi (Curculionidae: Scolytinae): expression profiles of different stages and responses to host allelochemicals. Insect Biochem Mol Biol 65:35–46. https://doi.org/10.1016/j.ibmb.2015.08.004
Dai L, Li ZM, Yu JM, Ma MY, Zhang RR, Chen H, Pham T (2015b) The CYP51F1 gene of Leptographium qinlingensis: sequence characteristic, phylogeny and transcript levels. Inter J Mol Sci 16(6):12014–12034. https://doi.org/10.3390/ijms160612014
Davies PJ (2010) Plant hormones. Springer, Dordrecht
Davis TS, Crippen TL, Hofstetter RW, Tomberlin JK (2013) Microbial volatile emissions as insect semiochemicals. J Chem Ecol 39:840–859. https://doi.org/10.1007/s10886-013-0306-z
de Mendonça AL, da Silva CE, de Mesquita FLT, da Campos RS, Do Nascimento RR, de Ximenes ECPA, Sant’Ana AEG (2009) Antimicrobial activities of components of the glandular secretions of leaf cutting ants of the genus Atta. Antonie Van Leeuwenhoek 95:295–303. https://doi.org/10.1007/s10482-009-9312-0
DiGuistini S, Wang Y, Liao NY, Taylor G, Tanguay P, Feau N, Henrissat B, Chan SK, Hesse-Orce U, Alamouti SM (2011) Genome and transcriptome analyses of the mountain pine beetle-fungal symbiont Grosmannia clavigera, a lodgepole pine pathogen. Proc Natl Acad Sci USA 108:2504–2509. https://doi.org/10.1073/pnas.1011289108
Duetz WA, Bouwmeester H, van Beilen JB, Witholt B (2003) Biotransformation of limonene by bacteria, fungi, yeasts, and plants. App Microbiol Biotechnol 61:269–277. https://doi.org/10.1007/s00253-003-1221-y
Erbilgin N, Powell JS, Raffa KF (2003) Effect of varying monoterpene concentrations on the response of Ips pini (Coleoptera: Scolytidae) to its aggregation pheromone: implications for pest management and ecology of bark beetles. Agric for Entomol 5:269–274. https://doi.org/10.1046/j.1461-9563.2003.00186.x
Faber BW, van Gorcom RFM, Duine JA (2001) Purification and characterization of benzoate-para-hydroxylase, a cytochrome P450 (CYP53A1), from Aspergillus niger. Arch Biochem Biophys 394:245–254. https://doi.org/10.1006/abbi.2001.2534
Ferrer-Sevillano F, Fernández-Cañón JM (2007) Novel phacB-encoded cytochrome P450 monooxygenase from Aspergillus nidulans with 3-hydroxyphenylacetate 6-hydroxylase and 3,4-dihydroxyphenylacetate 6-hydroxylase activities. Eukaryot Cell 6:514–520. https://doi.org/10.1128/EC.00226-06
Fischer C, Holl W (1992) Food reserves of scots pine (Pinus sylvestris L.). 2. Seasonal-changes and radial-distribution of carbohydrate and fat reserves in pine wood. Trees-Struct Funct 6(3):147–155. https://doi.org/10.1007/BF00202430
Fleet C, Breuil C, Uzunovic A (2001) Nutrient consumption and pigmentation of deep and surface colonizing sapstaining fungi in Pinus contorta. Holzforschung 55(4):340–346. https://doi.org/10.1515/HF.2001.057
García-Fraile P (2018) Roles of bacteria in the bark beetle holobiont—how do they shape this forest pest? Ann Appl Biol 172:111–125. https://doi.org/10.1111/aab.12406
Grabherr MG, Haas BJ, Yassour M et al (2011) Full-length transcriptome assembly from RNA-Seq data without a reference genome. Nat Biotechnol 29(7):644-U130. https://doi.org/10.1038/nbt.1883
Haridas S, Wang Y, Lim L, Massoumi Alamouti S, Jackman S, Docking R, Robertson G, Birol I, Bonlmann J, Breuil C (2013) The genome and transcriptome of the pine saprophyte Ophiostoma piceae, and a comparison with the bark beetle associated pine pathogen Grosmannia clavigera. BMC Genomics 14:373. https://doi.org/10.1186/1471-2164-14-373
Hernández-Martínez F, Briones-Roblero CI, Nelson DR, Rivera-Orduña FN, Zúñiga G (2016) Cytochrome P450 complement (CYPome) of Candida oregonensis, a gut-associated yeast of bark beetle, Dendroctonus rhizophagus. Fungal Biol 120:1077–1089. https://doi.org/10.1016/j.funbio.2016.06.005
Hesse-Orce U, DiGuistini S, Keeling CI, Wang Y, Li M, Henderson H, Docking TR, Liao NY, Robertson G, Holt RA, Jones SJM, Bohlmann J, Breuil C (2010) Gene discovery for the bark beetle-vectored fungal tree pathogen Grosmannia clavigera. BMC Genomics 11:536. https://doi.org/10.1186/1471-2164-11-536
Hofstetter RW, Mahfouz JB, Klepzig KD, Ayres MP (2005) Effects of tree phytochemistry on the interactions among endophloedic fungi associated with the southern pine beetle. J Chem Ecol 31:539–560. https://doi.org/10.1007/s10886-005-2035-4
Howe M, Keefover-Ring K, Raffa KF (2018) Pine engravers carry bacterial communities whose members reduce concentrations of host monoterpenes with variable degrees of redundancy, specificity, and capability. Environ Entomol 47:638–645. https://doi.org/10.1093/ee/nvy032
Hu X, Wang C, Wang L, Zhang R, Chen H (2014) Influence of temperature, pH and metal ions on guaiacol oxidation of purified laccase from Leptographium qinlingensis. World J Microbiol Biotechnol 30:1285–1290. https://doi.org/10.1007/s11274-013-1554-3
Ide M, Ichinose H, Wariishi H (2012) Molecular identification and functional characterization of cytochrome P450 monooxygenases from the brown-rot basidiomycete Postia placenta. Arch Microbiol 194:243–253. https://doi.org/10.1007/s00203-011-0753-2
Kalb VF, Woods CW, Turi TG, Dey CR, Sutter TR, Loper JC (1987) Primary structure of the P450 lanosterol demethylase gene from Saccharomyces cerevisiae. DNA 6:529–537. https://doi.org/10.1089/dna.1987.6.529
Kelly SL, Lamb DC, Corran AJ, Baldwin BC, Parks LW, Kelly DE (1995) Purification and reconstitution of activity of Saccharomyces cerevisiae P450 61, a sterol delta 22-desaturase. FEBS Lett 377:217–220. https://doi.org/10.1016/0014-5793(95)01342-3
Kimura M, Tokai T, Takahashi-Ando N, Ohsato S, Fujimura M (2007) Molecular and genetic studies of fusarium trichothecene biosynthesis: pathways, genes, and evolution. Biosci Biotechnol Biochem 71:2105–2123. https://doi.org/10.1271/bbb.70183
Kitazume T, Takaya N, Nakayama N, Shoun H (2000) Fusarium oxysporum fattyacid subterminal hydroxylase (CYP505) is a membrane-bound eukaryotic counterpart of Bacillus megaterium cytochrome P450BM3. J Biol Chem 275:39734–39740. https://doi.org/10.1074/jbc.M005617200
Kitazume T, Tanaka A, Takaya N, Nakamura A, Matsuyama S, Suzuki T, Shoun H (2002) Kinetic analysis of hydroxylation of saturated fatty acids by recombinant P450foxy produced by an Escherichia coli expression system. Eur J Biochem 269:2075–2082. https://doi.org/10.1046/j.1432-1033.2002.02855.x
Klepzig KD, Moser JC, Lombardero MJ, Ayres MP, Hofstetter RW, Walkinshaw CJ (2001) Mutualism and antagonism: ecological interactions among bark beetles, mites and fungi. In: Jeger MJ, Spence NJ (eds) Biotic interactions in plant-pathogen associations. CABI Publishing, Wallingford, pp 237–267
Kligun E, Ostretsov B, Titievsky A, Farkov M, Alamouti SM, Brodsky L (2017) Adaptation of the pine fungal pathogen Grosmannia clavigera to monoterpenes: biochemical mechanisms revealed by RNA-seq analysis. For Pathol 47(6):e12372. https://doi.org/10.1111/efp.12372
Kopper BJ, Illman BL, Kersten PJ, Klepzig KD, Raffa KF (2005) Effects of diterpene acids on components of a conifer bark beetle-fungal interaction: tolerance by Ips pini and sensitivity by its associate Ophiostoma ips. Environ Entomol 34:486–493. https://doi.org/10.1603/0046-225X-34.2.486
Kull FC, Eisman PC, Sylwestrowicz HD, Mayer RL (1961) Mixtures of quaternary ammonium compounds and long-chain fatty acids as antifungal agents. Appl Microbiol 9:538–541. https://doi.org/10.1128/am.9.6.538-541.1961
Lah L, Haridas S, Bohlmann J, Breuil C (2013) The cytochromes P450 of Grosmannia clavigera: genome organization, phylogeny, and expression in response to pine host chemicals. Fungal Genet Biol 50:72–81. https://doi.org/10.1016/j.fgb.2012.10.002
Lee S, Kim JJ, Breuil C (2006) Pathogenicity of Leptographium longiclavatum associated with Dendroctonus ponderosae to Pinus contorta. Can J for Res 36:2864–2872. https://doi.org/10.4039/n08-CPA01
Li XJ, Gao JM, Chen H, Zhang AL, Tang M (2012) Toxins from a symbiotic fungus, Leptographium qinlingensis associated with Dendroctonus armandi and their in vitro toxicities to Pinus armandi seedling. Eur J Plant Pathol 134:239–247. https://doi.org/10.1007/s10658-012-9981-9
Liu F, Wickham JD, Cao Q, Lu M, Sun JH (2020) An invasive beetle–fungus complex is maintained by fungal nutritional-compensation mediated by bacterial volatiles. ISME J 14:2829–2842. https://doi.org/10.1038/s41396-020-00740-w
Livak KJ, Schmittgen TD (2008) Analyzing real-time PCR data by the comparative CT method. Nat Protoc 3:1101–1108. https://doi.org/10.1038/nprot.2008.73
Matsuzaki F, Wariishi H (2005) Molecular characterization of cytochrome P450 catalyzing hydroxylation of benzoates from the white-rot fungus Phanerochaete chrysosporium. Biochem Biophys Res Commun 334:1184–1190. https://doi.org/10.1016/j.bbrc.2005.07.013
Matsuzaki F, Shimizu M, Wariishi H (2008) Proteomic and metabolomic analyses of the white-rot fungus Phanerochaete chrysosporium exposed to exogenous benzoic acid. J Proteome Res 7:2342–2350. https://doi.org/10.1021/pr700617s
Nakayama N, Takemae A, Shoun H (1996) Cytochrome P450foxy, a catalytically self-sufficient fatty acid hydroxylase of the fungus Fusarium oxysporum. J Biochem 119:435–440. https://doi.org/10.1093/oxfordjournals.jbchem.a021260
Nasir H, Noda H (2003) Yeast-like symbiotes as a sterol source in anobiid beetles (Coleoptera, Anobiidae): possible metabolic pathways from fungal sterols to 7-dehydrocholesterol. Arch Insect Biochem Physiol 52:175–182. https://doi.org/10.1002/arch.10079
Nelson DR (2009) The cytochrome p450 homepage. Hum Genomics 4:59–65. https://doi.org/10.1186/1479-7364-4-1-59
Nelson DR (2011) Progress in tracing the evolutionary paths of cytochrome P450. Biochim Biophys Acta 1814:14–18. https://doi.org/10.1016/j.bbapap.2010.08.008
Pham T, Chen H, Yu J, Dai L, Zhang R, Trang Vu TQ (2014) The Differential effects of the blue-stain fungus Leptographium qinlingensis on monoterpenes and sesquiterpenes in the stem of Chinese white pine (Pinus armandi) saplings. Forests 5:2730–2749. https://doi.org/10.3390/f5112730
Podobnik B, Stojan J, Lah L, Krasevec N, Seliskar M, Lanisnik Rizner T, Rozman D, Komel R (2008) CYP53A15 of Cochliobolus lunatus, a target for natural antifungal compounds. J Med Chem 51:3480–3486. https://doi.org/10.1021/jm800030e
Proctor RH, Brown DW, Plattner RD, Desjardins AE (2003) Co-expression of 15 contiguous genes delineates a fumonisin biosynthetic gene cluster in Gibberella moniliformis. Fungal Genet Biol 38:237–249. https://doi.org/10.1016/S1087-1845(02)00525-X
Raffa K, Smalley E (1995) Interaction of pre-attack and induced monoterpene concentrations in host conifer defense against bark beetle fungal complexes. Oecologia 102(3):285–295. https://doi.org/10.1007/BF00329795
Reid ML, Purcell JRC (2011) Condition-dependent tolerance of monoterpenes in an insect herbivore. Arthropod Plant Int 5:331–337. https://doi.org/10.1007/s11829-011-9137-4
Schirp A, Farrell R, Kreber B, Singh A (2003) Advances in understanding the ability of sapstaining fungi to produce cell wall-degrading enzymes. Wood Fiber Sci 35(3):434–444. https://doi.org/10.1177/004051750307300715
Schmidt K, Engel P (2021) Mechanisms underlying gut microbiota–host interactions in insects. J Exp Biol 224:jeb207696. https://doi.org/10.1242/jeb.207696
Scriber JM, Slansky F Jr (1981) The nutritional ecology of immature insects. Annu Rev Entomol 26:183–211. https://doi.org/10.1146/annurev.en.26.010181.001151
Shin J, Kim JE, Lee YW, Son H (2018) Fungal cytochrome P450s and the P450 complement (CYPome) of Fusarium graminearum. Toxins 10:112. https://doi.org/10.3390/toxins10030112
Six DL (2012) Ecological and evolutionary determinants of bark beetle-fungus symbioses. Insects 3:339–366. https://doi.org/10.3390/insects3010339
Six DL, Wingfield MJ (2011) The role of phytopathogenicity in bark beetle-fungus symbioses: a challenge to the classic paradigm. Annu Rev Entomol 56:255–272. https://doi.org/10.1146/annurev-ento-120709-144839
Skaggs BA, Alexander JF, Pierson CA, Schweitzer KS, Chun KT, Koegel C, Barbuch R, Bard M (1996) Cloning and characterization of the Saccharomyces cerevisiae C-22 sterol desaturase gene, encoding a second cytochrome P-450 involved in ergosterol biosynthesis. Gene 169:105–109. https://doi.org/10.1016/0378-1119(95)00770-9
Tamura K, Peterson D, Peterson N, Stecher G, Nei M, Kumar S (2011) MEGA5: molecular evolutionary genetics analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods. Mol Biol Evol 28:2731–2739. https://doi.org/10.1093/molbev/msr121
Tang M, Chen H, Zhao J, Zhu C (2004) Leptographium qinlingensis sp. nov. associated with Dendroctonus armandi in Pinus armandi. J Huazhong Agric Univ 23:5–6
Van Der Werf MJ, Boot AM (2000) Metabolism of carveol and dihydrocarveol in Rhodococcus erythropolis DCL14. Microbiology 146(Pt 5):1129–1141. https://doi.org/10.1099/00221287-146-5-1129
Wang Y, DiGuistini S, Wang TCT, Bohlmann J, Breuil C (2010) Agrobacterium meditated gene disruption using split-marker in Grosmannia clavigera, a mountain pine beetle associated pathogen. Curr Genet 56(3):297–307. https://doi.org/10.1007/s00294-010-0294-2
Wang B, Salcedo C, Lu M, Sun JH (2012) Mutual interactions between an invasive bark beetle and its associated fungi. Bull Entomol Res 102:71–77. https://doi.org/10.1017/S000748531100037X
Wang B, Lu M, Cheng CH, Salcedo C, Sun JH (2013a) Saccharidemediated antagonistic effects of bark beetle fungal associates on larvae. Biol Lett 9:20120787. https://doi.org/10.1098/rsbl.2012.0787
Wang Y, Lim L, DiGuistini S, Robertson G, Bohlmann J, Breuil C (2013b) A specialized ABC efflux transporter GcABC-G1 confers monoterpene resistance to Grosmannia clavigera, a bark beetle-associated fungal pathogen of pine trees. New Phytol 197(3):886–898. https://doi.org/10.1111/nph.12063
Wang Y, Lim L, Lina M, Ljerka L, Joerg B, Colette B (2014) Gene discovery for enzymes involved in limonene modification or utilization by the mountain pine beetle-associated pathogen Grosmannia clavigera. Appl Environ Microbiol 80:4566–4576. https://doi.org/10.1128/AEM.00670-14
Weisskopf L, Schulz S, Garbeva P (2021) Microbial volatile organic compounds in intra-kingdom and inter-kingdom interactions. Nat Rev Microbiol 19:391–404. https://doi.org/10.1038/s41579-020-00508-1
Yu JH, Keller N (2005) Regulation of secondary metabolism in filamentous fungi. Annu Rev Phytopathol 43:437–458. https://doi.org/10.1146/annurev.phyto.43.040204.140214
Yu J, Chang PK, Ehrlich KC, Cary JW, Bhatnagar D, Cleveland TE, Payne GA, Linz JE, Woloshuk CP, Bennett JW (2004) Clustered pathway genes in aflatoxin biosynthesis. Appl Environ Microbiol 70:1253–1262. https://doi.org/10.1128/aem.70.3.1253-1262.2004
Zabel R, Morrell J (1992) Wood stains and discolorations. In: Zabel R, Morrell J (eds) Wood microbiology: decay and its prevention. Academic Press Inc., San Diego, pp 326–343
Zwart Voorspuij AJ, Nass CA (1957) Some aspects of the notions additivity, synergism and antagonism in the simultaneous activity of two antibacterial agents in vitro. Arch Int Pharmacodyn Ther 109:211–228
Acknowledgements
The authors thank Dr. David Nelson for assigning CYP family names to newly identified L. qinlingensis CYPs. This work was supported with funds from the National Natural Science Foundation of China (31700572, 31870636) and the Natural Science Basic Research Plan in Shaanxi Province of China (2018JQ3055).
Author information
Authors and Affiliations
Corresponding author
Ethics declarations
Conflict of interest
The authors report no conflicts of interest.
Additional information
Communicated by Erko Stackebrandt.
Publisher's Note
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Supplementary Information
Below is the link to the electronic supplementary material.
Rights and permissions
About this article
Cite this article
Dai, L., Zheng, J., Ye, J. et al. Phylogeny of Leptographium qinlingensis cytochrome P450 genes and transcription levels of six CYPs in response to different nutrition media or terpenoids. Arch Microbiol 204, 16 (2022). https://doi.org/10.1007/s00203-021-02616-9
Received:
Revised:
Accepted:
Published:
DOI: https://doi.org/10.1007/s00203-021-02616-9