S. Sreenivasaprasad

DNA sequence variation and interrelationships among Colletotrichum species causing strawberry anthracnose

Abstract Restriction fragment length polymorphisms (RFLPs) of the ribosomal DNA (rDNA) and mitochondrial DNA (mtDNA) of isolates of the strawberry anthracnose pathogens Colletotrichum acutatum, C. fragariae and C. gloeosporioides were analysed using rDNA from Saccharomyces carlsbergensis and mtDNA extracted from C. acutatum, C. fragariae and C. gloeosporioides as probes. These analyses revealed considerable heterogeneity within C. acutatum from diverse hosts. The European strawberry isolates formed a discrete group while the American strawberry isolates fell into a broad group which included isolates from other hosts. No polymorphisms in either rDNA or mtDNA were observed among C. gloeosporioides isolates from strawberry. C. fragariae isolates divided into two groups with distinct rDNA and mtDNA patterns. Random amplified polymorphic DNA (RAPD) analysis grouped isolates in a similar manner to the mtDNA RFLP analysis. From sequencing data, the internally transcribed spacer (ITS) 1 region of the rDNA repeat unit of C. gloeosporioides, C. fragariae and C. acutatum was shown to be 171, 171 or 172 and 180 or 181 bases, respectively. C. gloeosporioides and C. fragariae differed from one another by only three to seven bases compared with C. acutatum which differed from C. gloeosporioides and C. fragariae by approx. 36–37 bases. ITS 1 data for C. acutatum generally support RFLP and RAPD taxonomies; differences of eight-eleven bases between European strawberry isolates and all other isolates studied being the greatest.

Nucleotide sequence of the rDNA spacer 1 enables identification of isolates of Colletotrichum as C. acutatum.

Nucleotide sequence of the rDNA internally transcribed spacer (ITS) 1 of twelve isolates of Colletotrichum , originally identified as either C. fructigenum, C. gloeosporioides or C. musae , was determined. Comparison of the sequence data with that from previously characterized isolates of Colletotrichum species revealed 97–100% homology with C. acutatum and only 79–81% homology with C. gloeosporioides and C. musae . We conclude that the isolates belong to C. acutatum . The value of sequence data from ITS 1 in species assignment is demonstrated.

Cross-infection potential of genetic groups of Colletotrichum gloeosporioides on tropical fruits

Cross-inoculation experiments demonstrated variation in the level of host preference among Colletotrichum gloeosporioides isolates from seven tropical fruit crops and also variation in the susceptibility of the hosts. In general, isolates were much more pathogenic on leaves of the original host than on those of alternative crops. Isolates from avocado, durian and mango showed the greatest degree of host preference while isolates from mangosteen and pini jambu were the most pathogenic on alternative crops. Avocado, mango and rambutan were the most susceptible and mangosteen and pini jambu were the least susceptible to C. gloeosporioides isolates from the other crops. The extent of cross-infection appeared dependent on inoculum density. Molecular markers based on restriction fragment length polymorphisms (RFLPs) in ribosomal DNA (rDNA), mitochondrial DNA (mtDNA) and random amplified polymorphic DNA (RAPD) enabled genetic grouping of the C. gloeosporioides isolates. In some instances, isolates from different hosts, e.g. two of the avocado isolates, all three durian isolates and one rambutan isolate, had identical rDNA and very similar mtDNA banding patterns (category 3) indicating a common ancestry, but differed in their host preference, implying adaptation to different crops.

Coffee berry disease pathogen in Africa: genetic structure and relationship to the group species Colletotrichum gloeosporioides

Restriction fragment length polymorphisms of the ribosomal DNA (rDNA) and mitochondrial DNA (mtDNA) of isolates of the coffee berry disease pathogen in Africa, Colletotrichum kahawae , were analysed using rDNA from Saccharomyces carlsbergensis and mtDNA extracted from C. kahawae and C. gloeosporioides as probes. These analyses revealed homogeneity among C. kahawae isolates. The estimated sizes of the rDNA repeat unit and the mtDNA of C. kahawae were 8·4–9·18 kb and 60 kb, respectively. Polymorphisms were observed in rDNA and mtDNA when C. kahawae was compared with C. gloeosporioides from coffee, avocado and mango. However, an avocado isolate JIA1, from Australia, had an identical rDNA restriction pattern to C. kahawae when digested with Bam H I and C. kahawae showed greater than 96% similarity to two C. gloeosporioides avocado isolates (918 and 1072) from New Zealand in mtDNA restriction fragment pattern. Random amplified polymorphic DNA analysis also grouped the C. kahawae isolates together. Nucleotide sequence of the internally transcribed spacer 1 region of the rDNA repeat unit of C. kahawae and C. gloeosporioides isolates from Coffea spp. differed by only two to three bases (98·8–98·2 % homology). Results obtained confirmed the close genetic relationship of C. kahawae to the group species C. gloeosporioides .

Genetic characterization of Colletotrichum gloeosporioides isolates obtained from mango

Abstract Most isolates of Colletotrichum gloeosporioides (>80%) obtained from anthracnose lesions on mango leaves, inflorescences and fruits from different regions in Sri Lanka showed the same ribosomal DNA (rDNA) and very similar mitochondrial DNA (mtDNA) restriction fragment banding patterns when genomic DNA was digested with endonuclease EcoR I. This rDNA restriction fragment banding pattern was the same as that revealed by C. gloeosporioides isolates from mango in other tropical countries. Isolates obtained from inflorescences all showed the same rDNA and mtDNA restriction patterns but a few isolates from fruits and leaves from mango grown in mixed orchards in areas of Sri Lanka with two monsoons and year‐round rainfall had rDNA restriction patterns associated with isolates from other fruit crops. This suggested that some cross‐infection onto mango occurs. However, it is suggested that there is a strain of C. gloeosporioides specific to mango and the clonal nature of this strain implies that it has di...

Selection of a genetic variant within Colletotrichum gloeosporioides isolates pathogenic on mango by passaging through wounded tomato fruits

Abstract A genetic variant virulent on tomato was selected by passaging field isolates of Colletotrichum gloeosporioides, obtained from mango, at least twice through wounded tomato fruits. The field isolates and isolates virulent on tomato showed ribosomal DNA restriction fragment length polymorphisms (RFLP). They also had different random amplified polymorphic DNA (RAPD) banding patterns. The mango field isolates produced conidia of sizes ranging from 12 to 22 μm and approx. 5% of these conidia were multinucleate. Cultures of the genetic variant produced uniformly-sized conidia (approx. 14 μm) which were uninucleate. The genetic variant was isolated in culture, only with difficulty, as a single conidium (L2t3) from a culture (L2) which, in turn, was grown from a single approx. 20 μm conidium obtained from one of the mango field isolates (M11). It is suggested that two genetically different nuclei residing within individual multinucleate conidia gave rise to the observed variation. The importance of heterokaryosis in relation to the considerable genetic variation and apparent adaptability seen in C. gloeosporioides is discussed.