IGS Minisatellites Useful for Race Differentiation in Colletotrichum lentis and a Likely Site of Small RNA Synthesis Affecting Pathogenicity.
Colletotrichum lentis is a fungal pathogen of lentil in Canada but rarely reported elsewhere. Two races, Ct0 and Ct1, have been identified using differential lines. Our objective was to develop a PCR-probe differentiating these races. Sequences of the translation elongation factor 1α (tef1α), RNA polymerase II subunit B2 (rpb2), ATP citrate lyase subunit A (acla), and internal transcribed spacer (ITS) regions were monomorphic, while the intergenic spacer (IGS) region showed length polymorphisms at two minisatellites of 23 and 39 nucleotides (nt). A PCR-probe (39F/R) amplifying the 39 nt minisatellite was developed which subsequently revealed 1-5 minisatellites with 1-12 repeats in C. lentis. The probe differentiated race Ct1 isolates having 7, 9 or 7+9 repeats from race Ct0 having primarily 2 or 4 repeats, occasionally 5, 6, or 8, but never 7 or 9 repeats. These isolates were collected between 1991 and 1999. In a 2012 survey isolates with 2 and 4 repeats increased from 34% to 67%, while isolated with 7 or 9 repeats decreased from 40 to 4%, likely because Ct1 resistant lentil varieties had been grown. The 39 nt repeat was identified in C. gloeosporioides, C. trifolii, Ascochyta lentis, Sclerotinia sclerotiorum and Botrytis cinerea. Thus, the 39F/R PCR probe is not species specific, but can differentiate isolates based on repeat number. The 23 nt minisatellite in C. lentis exists as three length variants with ten sequence variations differentiating race Ct0 having 14 or 19 repeats from race Ct1 having 17 repeats, except for one isolate. RNA-translation of 23 nt repeats forms hairpins and has the appropriate length to suggest that IGS could be a site of small RNA synthesis, a hypothesis that warrants further investigation. Small RNA from fungal plant pathogens able to silence genes either in the host or pathogen thereby aiding infection have been reported.
Agriculture and Agri-Food Canada, Saskatoon Research Centre, Saskatoon, Canada.
Jonathan
J
Durkin
Agriculture and Agri-Food Canada, Eastern Cereal and Oilseed Research Centre, Ottawa, Canada.
John
J
Bissett
Department of Agronomy and Plant Breeding, Gorgan University of Agricultural Sciences, Gorgan, Iran.
Mohammadhadi
M
Pahlavani
Agriculture and Agri-Food Canada, Saskatoon Research Centre, Saskatoon, Canada.
Brent
B
Mooney
Agriculture and Agri-Food Canada, Saskatoon Research Centre, Saskatoon, Canada.
Lone
L
Buchwaldt
D001275:ATP Citrate (pro-S)-Lyase / Q000235:genetics; D001203:Ascomycota / Q000235:genetics / Q000472:pathogenicity; D001483:Base Sequence; D020171:Botrytis / Q000235:genetics / Q000472:pathogenicity; D020231:Colletotrichum / Q000235:genetics / Q000472:pathogenicity*; D015342:DNA Probes; D021903:DNA, Ribosomal Spacer / Q000235:genetics; D039642:Eukaryotic Initiation Factors / Q000235:genetics*; D005656:Fungal Proteins / Q000235:genetics*; D054884:Host-Pathogen Interactions; D027806:Lens Plant / Q000382:microbiology; D018598:Minisatellite Repeats; D008969:Molecular Sequence Data; D016533:Mycological Typing Techniques; D010935:Plant Diseases / Q000382:microbiology; D011110:Polymorphism, Genetic; D021122:Protein Subunits / Q000235:genetics; D012319:RNA Polymerase II / Q000235:genetics; D012342:RNA, Small Nuclear / Q000096:biosynthesis / Q000235:genetics*; D017422:Sequence Analysis, DNA; D013045:Species Specificity; D014774:Virulence
D015342:DNA Probes; D021903:DNA, Ribosomal Spacer; D039642:Eukaryotic Initiation Factors; D005656:Fungal Proteins; D021122:Protein Subunits; D012342:RNA, Small Nuclear; D001275:ATP Citrate (pro-S)-Lyase; D012319:RNA Polymerase II