Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2021 Dec 16:2:778882.
doi: 10.3389/ffunb.2021.778882. eCollection 2021.

Increased Pathogenicity of the Nematophagous Fungus Drechmeria coniospora Following Long-Term Laboratory Culture

Damien Courtine  1 , Xing Zhang  1 , Jonathan J Ewbank  1
Affiliations

Increased Pathogenicity of the Nematophagous Fungus Drechmeria coniospora Following Long-Term Laboratory Culture

Damien Courtine et al. Front Fungal Biol. .

Abstract

Domestication provides a window into adaptive change. Over the course of 2 decades of laboratory culture, a strain of the nematode-specific fungus Drechmeria coniospora became more virulent during its infection of Caenorhabditis elegans. Through a close comparative examination of the genome sequences of the original strain and its more pathogenic derivative, we identified a small number of non-synonymous mutations in protein-coding genes. In one case, the mutation was predicted to affect a gene involved in hypoxia resistance and we provide direct corroborative evidence for such an effect. The mutated genes with functional annotation were all predicted to impact the general physiology of the fungus and this was reflected in an increased in vitro growth, even in the absence of C. elegans. While most cases involved single nucleotide substitutions predicted to lead to a loss of function, we also observed a predicted restoration of gene function through deletion of an extraneous tandem repeat. This latter change affected the regulatory subunit of a cAMP-dependent protein kinase. Remarkably, we also found a mutation in a gene for a second protein of the same, protein kinase A, pathway. Together, we predict that they result in a stronger repression of the pathway for given levels of ATP and adenylate cyclase activity. Finally, we also identified mutations in a few lineage-specific genes of unknown function that are candidates for factors that influence virulence in a more direct manner.

Keywords: C. elegans; domestication; evolution; genome; model organisms; virulence.

PubMed Disclaimer

Conflict of interest statement

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Figures

Figure 1
Figure 1
Phenotypic differences between Swe1 and Swe3. (A) Survival of C. elegans worms (strain IG463) following infection at 25°C with 1 × 109 spores of Swe1 or Swe3; n = 100 for each strain, ****p < 0.0001, one-sided log rank test. The results are representative of three independent trials. (B) Comparison of the size (left) and prevalence of adhesive buds (right), for spores from Swe1 and Swe3. A minimum of 100 spores were scored at each time point. (C) The number of spores at the mouth and vulval regions of C. elegans was counted after 15 h of infection with 1 × 109 spores of Swe1 or Swe3. Each dot represents the mean of an experiment with at least 10 worms; ns = not significant, two tailed unpaired t-test. (D) Left hand panels: Representative fluorescence images of age-matched IG463 worms uninfected (NI) or infected for 16 h (16 hpi) with Swe1 or Swe3. The worm strain carries the integrated transgene frIs7 that includes nlp-29p::GFP and col-12p::dsRed transgenes; red and green fluorescence is visualised simultaneously; scale bar: 200 μm. Quantification of the green/red fluorescence ratio (in arbitrary units) of IG463 worms at 24 hpi, compared to aged-matched non-infected (NI) worms; n > 250 worms for each, ****p < 0.0001, unpaired t-test. (E) Representative images of Swe1 or Swe3 infected worms at 48, 96 and 144 hpi. The 2nd panels from the left are a higher magnification of the indicated vulval regions. The arrowhead highlights Swe1 spores attached to the vulva (top), and hyphae growing out from the Swe3-infected worm (bottom); scale bar: 50 μm. The righthand panels show hyphae growing from dead worms; scale bar: 1 mm. (F) Spores of Swe1 or Swe3 (harvested 6 dpi) were seeded on NGM plates, incubated at 25°C without worms and images taken at the indicated times. The extent of hyphal growth from a single spore is delimited by dotted lines in the images at 69 h. The two right-hand columns illustrate the degree of spore germination with or without OP50 after 96 h; scale bar: 100 μm.
Figure 2
Figure 2
MAT loci in D. coniospora. The Swe2 region syntenic to the Dan2 MAT locus is shown. The previously annotated Dan2 MAT genes are indicated with green arrows, labelled with their idiomorph identifiers. The newly identified Swe2 MAT1-2-1 gene is shown in blue. The red shapes highlight the extent of the sequence synteny, and the black arrows represent a pair of neighbouring orthologous genes.
Figure 3
Figure 3
Classification of mutations identified between the genomes of the Swe strains. The overview includes the number of genes and mutations identified at each step of the workflow (see section Methods for details). Mutations identified in introns were individually examined; none affected splice donor or acceptor sites.
Figure 4
Figure 4
Comparison of the growth of Swe1 and Swe3 in hypoxia-like conditions. (A) Quantification of growth rates for Swe1 and Swe3 on standard NGM plates (0 mM), or plates supplemented with CoCl2 to a final concentration of 1 or 2 mM. As illustrated in Figure 1F, measurement beyond 44 h of Swe1 and Swe3 growth in the absence of CoCl2 is confounded by the overgrowth of mycelia derived from separate spores. The average size and 95% confidence interval for 8–35 individual spores per condition, followed at each time point, are shown. (B) Representative images of spores of Swe1 and Swe3 growing in the presence of 1 or 2 mM CoCl2. A direct comparison can be made with Figure 1F, as the experiments were conducted in parallel.
Figure 5
Figure 5
Two mutations potentially affect the same pathway. Simplified diagram illustrating the regulation of protein kinase A (PKA) by cAMP. Under low cAMP condition, the PKA regulatory and catalytic subunits dimerize, leading to an inactive PKA. In Swe1, as the regulatory subunit is constitutively inactive, PKA would be predicted to be permanently active. In Swe2, the inactivating mutation of the regulatory subunit is reverted to wild-type (as indicated by the green cross), so PKA would again come under the control of cAMP levels. Its activity is also regulated by a cAMP phosphodiesterase (PDE), as this converts cAMP to AMP. The PDE bears a non-synonymous mutation in Swe2 (red cross). Assuming that this corresponds to a loss of function, the two mutations would act synergistically, as cAMP would accumulate, thereby accentuating the suppression of repression of PKA activity. Thus PKA would be predicted to be more active in Swe2 (and Swe3) than in Swe1 under the same culture conditions.
See this image and copyright information in PMC

Similar articles

See all similar articles

References

    1. Altschul S. F., Madden T. L., Schaffer A. A., Zhang J., Zhang Z., Miller W., et al. . (1997). Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res. 25, 3389–3402 10.1093/nar/25.17.3389 - DOI - PMC - PubMed
    1. Andersson K. M., Kumar D., Bentzer J., Friman E., Ahren D., Tunlid A. (2014). Interspecific and host-related gene expression patterns in nematode-trapping fungi. BMC Genomics 15:968. 10.1186/1471-2164-15-968 - DOI - PMC - PubMed
    1. Brunk M., Sputh S., Doose S., van de Linde S., Terpitz U. (2018). HyphaTracker: an ImageJ toolbox for time-resolved analysis of spore germination in filamentous fungi. Sci. Rep. 8:605. 10.1038/s41598-017-19103-1 - DOI - PMC - PubMed
    1. Chung D., Barker B. M., Carey C. C., Merriman B., Werner E. R., Lechner B. E., et al. . (2014). ChIP-seq and in vivo transcriptome analyses of the Aspergillus fumigatus SREBP SrbA reveals a new regulator of the fungal hypoxia response and virulence. PLoS Pathog. 10:e1004487. 10.1371/journal.ppat.1004487 - DOI - PMC - PubMed
    1. Courtine D., Provaznik J., Reboul J., Blanc G., Benes V., Ewbank J. J. (2020). Long-read only assembly of Drechmeria coniospora genomes reveals widespread chromosome plasticity and illustrates the limitations of current Nanopore methods. GigaScience 9:giaa099. 10.1093/gigascience/giaa099 - DOI - PMC - PubMed

LinkOut - more resources

-