Nuclear rDNA phylogeny in the fungal genus Verticillium and its relationship to insect and plant virulence, extracellular proteases and carbohydrases

Phylogenetic relationships among 18 isolates in the genus Verticillium, rep resenting 13 species of diverse econutritional groups (pathogens of insects , plants, mushrooms, nematodes and spiders, and saprobes), were examined by using sequences from the internal transcribed spacer (ITS) and small nucle ar (NS) rRNA regions. The isolates were also assessed for their abilities t o infect insect larvae (Galleria mellonella) and to cause necrosis in alfal fa (Medicago sativa), and for their proteolytic, chitinolytic and pectinoly tic activities. The phylogenetic data suggested that Verticillium is polyph yletic in origin and is therefore a form genus. However, the phylogenetic t ree supported the plant pathogens (V. dahliae, V. albo-atrum and V. nigresc ens) as a clade. The alfalfa isolate of V. albo-atrum (isolate 595) was an interesting outlier to the main body of plant pathogens as it clustered wit h the insect pathogen V. indicum. Strains of V. lecanii and V. indicum were able to infect insects and are present in divergent groups in the consensu s tree, suggesting that the ability to infect insects may have evolved inde pendently many times. Similarly, the nematophagous Verticillium species app ear to have evolved independently along several different routes and one is olate, V. chlamydosporium, was able to infect insects. V. albo-atrum, V. ni grescens and V. dahliae all produced high levels of enzymes capable of degr ading pectin, a major component of plant cell walls. The ability to excrete pectinase was a broad indicator of the ability to produce lesions on alfal fa. In the plant pathogens, the functions of a broad-spectrum protease were assumed by trypsins which degrade Bz-AA-AA-Arg-NA substrates (Br, benzoyl; AA, various amino acids; NA, p-nitroanilide). The insect pathogens and mus hroom pathogen (V. fungicola) were characterized by production of high leve ls of subtilisin-like proteases active against a chymotrypsin substrate (su ccinyl-Ala(2)-Pro-Phe-NA) and the inability to clear pectin. The insect and mushroom pathogens, and several nematode pathogens, were distinguishable f rom the plant pathogens in their ability to produce chitinases.