Molecular analysis of genetic diversity and variability in Colletotrichum gloeosporioides

Results from this study suggest two distinct genetic subpopulations of Colletotrichum gloeosporioides from citrus based on DNA variation, cultural morphology, and growth. Type 1 strains are slow growing, morphologically stable, benomyl tolerant, and contain a single form of ribosomal DNA (rDNA) as detected by common HindIII, PstI, SphI and SstI fragments hybridizing to cloned Neurospora crassa rDNA. Type 2 strains are faster growing, morphologically less stable, benomyl sensitive, and have rDNA distinct from type 1 strains. The rDNA from type 1 and type 2 strains were cloned and mapped for 10 restriction enzyme sites and genes coding for large subunit, small subunit and 5.8S rRNA. A subclone constructed from the non-transcribed spacer region of type 1 rDNA clone hybridizes only to rDNA from type 1 strains. DNA polymorphisms detected by heterologous hybridization with cloned N. crassa genes for glutamate dehydrogenase, anthranilate synthetase, histidinol dehydrogenase, and {dollar}beta{dollar}-tubulin corresponded to type 1 or type 2 strains. All strains liberate free fatty acid from ({dollar}sp3{dollar}H) -labelled cutin and hydrolyze cutin model substrates. Serine esterases from extracellular fluids of cutin-grown C. gloeosporioides strains were detected by labelling proteins separated by sodium dodesyl sulfate polyacrylamide gel electrophoresis with {dollar}sp3{dollar}H-diisofluorophosphate. The two major esterases from type 1 strains have molecular weights of 26 and 20 kilodaltons (kd) whereas the type 2 esterases were 24 and 22 kd. A DNA probe containing the cloned cutinase gene from C. gloeosporioides hybridized strongly to DNA from type 2 strains but poorly to type 1 strains. Distinct cutinase genes may be present in the two types of C. gloeosporioides strains from citrus. Chromosome-sized DNAs separated by pulsed-field gel electrophoresis corresponded to type 1 or type 2 strains. Type 1 strains had five large chromosomal DNAs 7.6, 7.0, 4.7, 3.7, and 3.3 (or 2.8) million base pairs (Mb) in size and one or two smaller chromosomes (1.6 to 0.63 Mb). Type 2 strains had three large chromosomal DNAs (7.8, 4.7, and 3.7 Mb) and two to four smaller chromosomal DNAs (0.52-0.28 Mb).