EST mining identifies proteins putatively secreted by the anthracnose pathogen Colletotrichum truncatum

Abstract

BackgroundColletotrichum truncatum is a haploid, hemibiotrophic, ascomycete fungal pathogen that causes anthracnose disease on many economically important leguminous crops. This pathogen exploits sequential biotrophic- and necrotrophic- infection strategies to colonize the host. Transition from biotrophy to a destructive necrotrophic phase called the biotrophy-necrotrophy switch is critical in symptom development. C. truncatum likely secretes an arsenal of proteins that are implicated in maintaining a compatible interaction with its host. Some of them might be transition specific.ResultsA directional cDNA library was constructed from mRNA isolated from infected Lens culinaris leaflet tissues displaying the biotrophy-necrotrophy switch of C. truncatum and 5000 expressed sequence tags (ESTs) with an average read of > 600 bp from the 5-prime end were generated. Nearly 39% of the ESTs were predicted to encode proteins of fungal origin and among these, 162 ESTs were predicted to contain N-terminal signal peptides (SPs) in their deduced open reading frames (ORFs). The 162 sequences could be assembled into 122 tentative unigenes comprising 32 contigs and 90 singletons. Sequence analyses of unigenes revealed four potential groups: hydrolases, cell envelope associated proteins (CEAPs), candidate effectors and other proteins. Eleven candidate effector genes were identified based on features common to characterized fungal effectors, i.e. they encode small, soluble (lack of transmembrane domain), cysteine-rich proteins with a putative SP. For a selected subset of CEAPs and candidate effectors, semiquantitative RT-PCR showed that these transcripts were either expressed constitutively in both in vitro and in planta or induced during plant infection. Using potato virus X (PVX) based transient expression assays, we showed that one of the candidate effectors, i. e. contig 8 that encodes a cerato-platanin (CP) domain containing protein, unlike CP proteins from other fungal pathogens was unable to elicit a hypersensitive response (HR).ConclusionsThe current study catalogues proteins putatively secreted at the in planta biotrophy-necrotrophy transition of C. truncatum. Some of these proteins may have a role in establishing compatible interaction with the host plant.

Achievements and Challenges in Legume Breeding for Pest and Disease Resistance

“…Secreted phytotoxins contribute to virulence or pathogenicity by disrupting host cells and inducing the release of nutrients to facilitate colonization of host tissues (Berestetskiy, 2008). The hemi-biotrophic lentil anthracnose pathogen, Colletotrichum truncatum, expresses an effector gene, CtNUDIX, exclusively during the later biotrophic phase before switching to necrotophic phase, and elicits a hypersensitive response from the host plant (Bhadauria et al, 2011). Overexpression of CtNUDIX in the pathogen causes incompability with the host plant (Bhadauria et al, 2013).…”

Overexpression of a Novel Biotrophy-Specific Colletotrichum truncatum Effector, CtNUDIX, in Hemibiotrophic Fungal Phytopathogens Causes Incompatibility with Their Host Plants

“…Leaflets from 3-week-old lentil plants were detached and inoculated in petri dishes lined with wet filter paper by droplet inoculation with a C. truncatum CT-21 isolate, as described previously (17). The inoculated leaflets were incubated with a 12-h photoperiod at room temperature.…”

“…Lentil plants of the Canadian cultivar Eston and the compatible C. truncatum CT-21 isolate were routinely maintained as described by Bhadauria et al (17). Cell walls were extracted from 3-week-old lentil leaflets following the protocol described previously (18).…”

“…2A and B). Thin secondary hyphae started to differentiate from the biotrophic primary hyphae in infected lentil tissues at 48 hpi, signaling the necrotrophic phase (17). CtNUDIX is therefore the first effector gene identified from any hemibiotrophic phytopathogen that shows explicit expression exclusively during and specific to the late biotrophic phase of Bhadauria et al infection.…”

“…In a previous study, we constructed an in planta BNS-specific cDNA plasmid library from the susceptible Canadian lentil cultivar Eston infected with C. truncatum isolate CT-21 and identified 122 unigenes encoding proteins with putative signal peptides, including effectors (17). In this study, we characterized one of the effector proteins, Ct21-1373, now named CtNUDIX.…”