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

Abstract

Phylogenetic relationships among 18 isolates in the genus Verticillium, representing 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 nuclear (NS) rRNA regions. The isolates were also assessed for their abilities to infect insect larvae (Galleria mellonella) and to cause necrosis in alfalfa (Medicago sativa), and for their proteolytic, chitinolytic and pectinolytic activities. The phylogenetic data suggested that Verticillium is polyphyletic in origin and is therefore a form genus. However, the phylogenetic tree supported the plant pathogens (V. dahliae, V. albo-atrum and V. nigrescens) 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 with 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 consensus tree, suggesting that the ability to infect insects may have evolved independently many times. Similarly, the nematophagous Verticillium species appear to have evolved independently along several different routes and one isolate, V. chlamydosporium, was able to infect insects. V. albo-atrum, V. nigrescens and V. dahliae all produced high levels of enzymes capable of degrading pectin, a major component of plant cell walls. The ability to excrete pectinase was a broad indicator of the ability to produce lesions on alfalfa. In the plant pathogens, the functions of a broad-spectrum protease were assumed by trypsins which degrade Bz-AA-AA-Arg-NA substrates (Bz, benzoyl; AA, various amino acids; NA, p-nitroanilide). The insect pathogens and mushroom pathogen (V. fungicola) were characterized by production of high levels of subtilisin-like proteases active against a chymotrypsin substrate (succinyl-Ala,-Pro-Phe-NA) and the inability to clear pectin. The insect and mushroom pathogens, and several nematode pathogens, were distinguishable from the plant pathogens in their ability to produce chitinases.

Fungal chitinases and their biological role in the antagonism onto nematode eggs. A review

“…Two opportunistic species have been thoroughly studied and tested as biocontrol agents: Pochonia chlamydosporia (Goddard) Zare and Gams (= Verticillium chlamydosporium) and P. lilacinus. These parasites grow in soil and may colonize the rhizosphere and the root (Kerry 1990;Siddiqui and Mahmood 1996;Bidochka et al 1999;Kerry 2000;Siddiqui et al 2000;Hirsch et al 2001;Khan et al 2001Khan et al , 2006Mukhtar and Pervaz 2003;Brand et al 2004). Both species have been used in the preparation of commercial formulations for the control of cyst and root-knot nematodes (Stirling et al 1998;Kerry 2000, Schenck 2004, Rumbos and Kiewnick 2006.…”

Role of inhibitors of proteolytic enzymes in plant defense against phytopathogenic microorganisms

“…All known serine proteinases of phy topathogens can be divided into trypsin like and subtil isin like enzymes. The first group contains proteinases that are produced by Cochliobolus carbonum [16], Verticillium dahliae [17,18], Stagonospora (Septoria) nodorum [19], and Phytophtora infestans [20] microorgan isms. Subtilisin like enzymes are secreted by C. carbonum [16], P. infestans [20], Acremonium typhium [21], Magnaporthe poae [22], Trichoderma harzianum [23], and Fusarium oxysporum [24].…”

New Insights on the Mode of Action of Fungal Pathogens of Invertebrates for Improving Their Biocontrol Performance

“…It has been suggested that FPI may have arisen from plant pathogens to escape competition via further specialisation in alternative hosts (Barron 1992;Bidochka et al 1999). FPI capabilities of changing hosts between invertebrates (insects or nematodes), endophytism and mycoparasitism could then be the consequences of this evolutionary process (Lopez-Llorca and Jansson 2006).…”

“…Serine proteases from both nematophagous and entomopathogenic fungi seem to have a common ancestor, different from that of plant-parasitic fungi ). Production of subtilisins may be a retained character from a saprophytic ancestor, since subtilisin-like proteases are the principal broad-spectrum proteases produced by many saprophytes (Bidochka et al 1999).…”

“…Subtilisin-like proteases are a large family of endopeptidases present only in fungi and Gram-negative bacteria, in contrast to the trypsin-like family (EC3.4.21.4). Broad-spectrum subtilisin-like proteases are the major proteins produced by the insect pathogens M. anisopliae and B. bassiana during infection processes, and have much greater ability than the trypsin-like enzymes to degrade insect cuticle (Bidochka et al 1999). Collagenase secretion upon penetration of nematode cuticles by Arthrobotrys spp.…”