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.