title: The Ifchit1 chitinase gene acts as a critical virulence factor in the insect pathogenic fungus Isaria fumosorosea.
abstract: The filamentous fungus, Isaria fumosorosea, is a promising insect biological control agent. Chitinases have been implicated in targeting insect cuticle structures, with biotechnological potential in insect and fungal control. The I. fumosorosea chitinase gene, Ifchit1, was isolated and determined to encode a polypeptide of 423 amino acids (46 kDa, pI = 6.53), present as a single copy in the I. fumosorosea genome. A split marker transformation system was developed and used to construct an Ifchit1 gene knockout. The ΔIfchit1 strain displayed minor alterations in mycelial growth on diverse media at 26 °C compared to the wild type and complemented (ΔIfchit1::Ifchit1) strains; however, colony morphology was affected, and the mutant strain had a temperature sensitive phenotype (32 °C). Although sporulation was delayed for the mutant, overall conidial production was almost twice than that of wild type. Biochemical assays indicated decreased chitinase activity during growth in Czapek-Dox liquid media for the ΔIfchit1 strain. Insect bioassays using diamondback moth, Plutella xylostella, larvae revealed decreased infectivity, i.e., increased LC 50 (threefold to fourfold) and a significantly delayed time to death, LT 50 from 3 to 6 days, for the ΔIfchit1 strain compared to the wild type and complemented strains. These data indicate an important role for the Ifchit1 chitinase as a virulence factor in I. fumosorosea.
authors.affiliation: Engineering Research Center of Biological Control, Ministry of Education, College of Agriculture, South China Agricultural University, Guangzhou, 510642, China. hzscau@126.com.|Department of Microbiology and Cell Science, Institute of Food and Agricultural Sciences, University of Florida, Bldg 981, Museum Rd, Gainesville, FL, 32611, USA. hzscau@126.com.
authors.forename: Zhen
authors.initials: Z
authors.lastname: Huang
authors.affiliation: Engineering Research Center of Biological Control, Ministry of Education, College of Agriculture, South China Agricultural University, Guangzhou, 510642, China.
authors.forename: Yongfen
authors.initials: Y
authors.lastname: Hao
authors.affiliation: Engineering Research Center of Biological Control, Ministry of Education, College of Agriculture, South China Agricultural University, Guangzhou, 510642, China.
authors.forename: Tianni
authors.initials: T
authors.lastname: Gao
authors.affiliation: Engineering Research Center of Biological Control, Ministry of Education, College of Agriculture, South China Agricultural University, Guangzhou, 510642, China.
authors.forename: Yü
authors.initials: Y
authors.lastname: Huang
authors.affiliation: Engineering Research Center of Biological Control, Ministry of Education, College of Agriculture, South China Agricultural University, Guangzhou, 510642, China. shxren@scau.edu.cn.
authors.forename: Shunxiang
authors.initials: S
authors.lastname: Ren
authors.affiliation: Department of Microbiology and Cell Science, Institute of Food and Agricultural Sciences, University of Florida, Bldg 981, Museum Rd, Gainesville, FL, 32611, USA. keyhani@ufl.edu.
authors.forename: Nemat O
authors.initials: NO
authors.lastname: Keyhani
mesh_terms: D000818:Animals; D002688:Chitinases / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D055786:Gene Knockout Techniques; D016681:Genome, Fungal; D006999:Hypocreales / Q000235:genetics* / Q000254:growth & development / Q000472:pathogenicity / Q000502:physiology; D007300:Insect Control; D007814:Larva / Q000382:microbiology; D009036:Moths / Q000254:growth & development; D010572:Pest Control, Biological; D010641:Phenotype; D013172:Spores, Fungal; D014170:Transformation, Genetic; D037521:Virulence Factors / Q000235:genetics* / Q000378:metabolism
chemical_list: D037521:Virulence Factors; D002688:Chitinases
keywords: Chitinase gene; Entomopathogenic fungi; Ifchit1; Isaria fumosorosea; Transformation system; Virulence