Out in the Cold: Identification of Genomic Regions Associated With Cold Tolerance in the Biocontrol Fungus Clonostachys rosea Through Genome-Wide Association Mapping

Abstract

There is an increasing importance for using biocontrol agents in combating plant diseases sustainably and in the long term. As large scale genomic sequencing becomes economically viable, the impact of single nucleotide polymorphisms (SNPs) on biocontrol-associated phenotypes can be easily studied across entire genomes of fungal populations. Here, we improved a previously reported genome assembly of the biocontrol fungus Clonostachys rosea strain IK726 using the PacBio sequencing platform, which resulted in a total genome size of 70.7 Mbp and 21,246 predicted genes. We further performed whole-genome re-sequencing of 52 additional C. rosea strains isolated globally using Illumina sequencing technology, in order to perform genome-wide association studies in conditions relevant for biocontrol activity. One such condition is the ability to grow at lower temperatures commonly encountered in cryic or frigid soils in temperate regions, as these will be prevalent for protecting growing crops in temperate climates. Growth rates at 10°C on potato dextrose agar of the 53 sequenced strains of C. rosea were measured and ranged between 0.066 and 0.413 mm/day. Performing a genome wide association study, a total of 1,478 SNP markers were significantly associated with the trait and located in 227 scaffolds, within or close to (< 1000 bp distance) 265 different genes. The predicted gene products included several chaperone proteins, membrane transporters, lipases, and proteins involved in chitin metabolism with possible roles in cold tolerance. The data reported in this study provides a foundation for future investigations into the genetic basis for cold tolerance in fungi, with important implications for biocontrol.

LysM Proteins Regulate Fungal Development and Contribute to Hyphal Protection and Biocontrol Traits in Clonostachys rosea

“…The higher expression of lysm2 during growth on chitincontaining media hints on a possible function of LYSM2 in protecting C. rosea hyphae against endogenous chitinases secreted during external chitin degradation. This hypothesis gain further support by the fact that lysm2 is located adjacent to the killer toxin-like chitinase gene chiC1 in the C. rosea genome Broberg et al, 2018), and that the transcriptional analyses of lysm2 in this study and chiC1 in a previous study showed that both genes are co-induced in medium containing chitin as the sole carbon source. There are several examples of chitinase genes located physically close to co-regulated LysM protein-encoding genes (Gruber et al, 2011;Martinez et al, 2012), indicating an interrelated function of chitinases and LysM proteins (Gruber and Seidl-Seiboth, 2012).…”

Natural variation of root lesion nematode antagonism in the biocontrol fungus Clonostachys rosea and identification of biocontrol factors through genome‐wide association mapping

“…Predicted NRPS proteins were retrieved from the C. rosea IK726 genome sequence (Broberg et al., 2018; Karlsson et al., 2015). Identification of conserved protein modules and features was made using the conserved domain database (CDD) (Marchler‐Bauer et al., 2017) and the simple modular architecture research tool (SMART) (Letunic & Bork, 2018).…”

Comparative genomics highlights the importance of drug efflux transporters during evolution of mycoparasitism in Clonostachys subgenus Bionectria (Fungi, Ascomycota, Hypocreales)

“…Abbreviation: n.a., ot annotated. a In comparison with the C. rosea strain IK726 reference genome (Broberg et al, 2018).…”

“…As part of a large-scale C. rosea genome sequencing initiative (Broberg et al, 2018) Figure S2). However, strains identified as C. rhizophaga clustered in two polyphyletic subgroups in the tef1 tree and the tub sequence-based analysis inferred C. byssicola as a paraphyletic group with respect to C. chloroleuca and C. rhizophaga Figure 2b).…”