Molecular and pathological differentiation of colletotrichum truncatum from scentless chamomile and legume crops

The fungus Colletotrichum truncatum is a potential biocontrol agent (BA) against the noxious weed scentless chamomile (Metricaria perforata Mérat; syn.: Tripleurospermum perforatum (Mérat) Lainz) in western Canada. This potential BA, however, is taxonomically related to the anthracnose pathogen on lentil, raising questions about crop safety. Ribosomal DNA (rDNA) internal transcribed space (ITS) regions of C. truncatum isolates collected from different plant hosts were examined, and compared with additional Colletotrichum species. Sequences were amplified with the universal primers its4 and its5, and C. truncatum isolates from scentless chamomile and selected legume crops were differentiated consistently. All scentless chamomile isolates fell within a single cluster in phylogenetic trees, regardless of their geographic origins. These isolates were more closely related to lentil isolates of C. truncatum than to isolates from the other host species. Soybean isolates, with more falcate and slender conidia and slightly bigger appressoria, were distinguishable from other C. truncatum isolates, while the isolates from scentless chamomile, lentil and pea were morphologically more similar. Based on sequence information, strain-specific PCR primers were designed for C. truncatum isolates from these hosts and used to amplify specific DNA bands (markers) from isolates of C. truncatum. This technique may be used for rapid detection and differentiation of C. truncatum, from scentless chamomile and designated legume species, as well as for tracking the BA after release. Inoculation trials were conducted using detached leaves and whole plants to determine potential cross infection of these C. truncatum isolates. Isolates from scentless chamomile caused disease only on their original host, but not on lentil, pea, soybean or alfalfa. In contrast, lentil isolates caused severe disease on lentil and pea, light symptoms on alfalfa, but no disease on the other hosts tested. Potential penetration of lentil leaves by scentless chamomile isolates was tested, with 2-23% incidence of the fungus from inoculated detached, senescence leaves but disease symptoms were not observed on either detached leaves or whole plants. Examination of the infection process revealed that scentless chamomile and lentil isolates had a similar pattern of infection and disease development on their respective hosts; infection vesicles were produced 24 h after inoculation, both primary and secondary infection hyphae were present, and the onset of disease symptoms tended to coincide with the development of secondary hyphae. The current study provided molecular and pathological evidence that differentiates the potential BA of scentless chamomile from C. truncatum isolates from lentil, pea and soybean