AUTHOR=Zhou Rong , Zhou Xiazhi , Fan Ali , Wang Zhangxun , Huang Bo 

TITLE=Differential Functions of Two Metalloproteases, Mrmep1 and Mrmep2, in Growth, Sporulation, Cell Wall Integrity, and Virulence in the Filamentous Fungus Metarhizium robertsii

JOURNAL=Frontiers in Microbiology

VOLUME=9

YEAR=2018

URL=https://www.frontiersin.org/journals/microbiology/articles/10.3389/fmicb.2018.01528

DOI=10.3389/fmicb.2018.01528

ISSN=1664-302X

ABSTRACT=<p>The <italic>Metarhizium</italic> genus of filamentous entomopathogenic fungi plays a pivotal role in regulating insect populations. Metalloproteases (MEPs) are a widely distributed and diverse family of hydrolytic enzymes that are important toxicity factors in the interactions between fungi and their hosts. Herein, we characterized two MEPs, Mrmep1 and Mrmep2, in <italic>Metarhizium robertsii</italic> using gene deletion. Growth rates of the resulting Δ<italic>Mrmep1</italic> and Δ<italic>Mrmep2</italic> mutants decreased by 16.2 and 16.5%, respectively, relative to the wild-type (WT) strain. Both mutants were less sensitive to cell wall-perturbing agents, sodium dodecyl sulfate and Congo red than the WT strain, whereas did not show any obvious changes in fungal sensitivity to ultraviolet B irradiation or heat stress. The conidial yield of Δ<italic>Mrmep1</italic>, Δ<italic>Mrmep2</italic>, and Δ<italic>Mrmep1</italic>Δ<italic>Mrmep2</italic> mutants decreased by 56.0, 23, and 53%, respectively. Insect bioassay revealed that median lethal time values against <italic>Galleria mellonella</italic> increased by 25.5% (Δ<italic>Mrmep1</italic>), 19% (Δ<italic>Mrmep2</italic>), and 28.8% (Δ<italic>Mrmep1</italic>Δ<italic>Mrmep2</italic>) compared with the WT strain at a concentration of 1 × 10<sup>7</sup> conidia mL<sup>-1</sup>, suggesting attenuated fungal virulence in the Δ<italic>Mrmep1</italic>, Δ<italic>Mrmep2,</italic> and Δ<italic>Mrmep1</italic>Δ<italic>Mrmep2</italic> strains. During fungal infection, transcription levels of <italic>Mrmep1</italic> was 1.6-fold higher than <italic>Mrmep2</italic> at 36 h post inoculation. Additionally, transcription levels of gallerimycin gene were 1.2-fold, 2.18-fold, and 2.5-fold higher in insects infected with the Δ<italic>Mrmep1</italic>, Δ<italic>Mrmep2,</italic> or Δ<italic>Mrmep1</italic>Δ<italic>Mrmep2</italic> mutant than those infected with the WT strain, respectively. Our findings suggest that <italic>Mrmep1</italic> and <italic>Mrmep2</italic> are differentially contributed to the growth, sporulation, cell wall integrity, and virulence of <italic>M. robertsii</italic>.</p>