Biocontrol potential of a native species of Trichoderma longibrachiatum against Meloidogyne incognita

Highlights

• T. longibrachiatum was found for the first time to be an effective bio-control agent against M. incognita.

• T. longibrachiatum has significant potential against second stage juveniles of M. incognita.

• The best T. longibrachiatum concentration against M. incognita was at 1.5 × 10⁷ conidia ml⁻¹.

• Main mechanism of T. longibrachiatum against M. incognita was via parasitism and inhibition.

• T. longibrachiatum has a great ability to colonize the rhizosphere of cucumber.

Abstract

Meloidogyne incognita is one of the most important soil-borne and plant parasitic pathogens in cucumber worldwide. An ideal strategy is to develop and use effective and environmentally-friendly bio-control agents to control the plant parasitic pathogen M. incognita. This study determined the biocontrol efficacy of Trichoderma longibrachiatum, a fungal species native to China, against M. incognita. In vitro, T. longibrachiatum at the concentrations of 1.5 × 10⁵ to 1.5 × 10⁷ conidia ml⁻¹ had a strong lethal and parasitic effect on the second stage juveniles of M. incognita, with the concentration of 1.5 × 10⁷ conidia ml⁻¹ inhibited and parasitized the second stage juveniles >88% 14 days after treatments. Microscope observations revealed that after the mutual recognition with the second stage juveniles, the conidia of T. longibrachiatum adhered or parasitized on the surface of second stage juveniles, germinated with a large number of hyphae and penetrated the integument, and was reproduced on the surface of second stage juveniles. Meanwhile, the surface of second stage juveniles became deformed and some were completely dissolved by the metabolite of T. longibrachiatum. The maximum protease activity of T. longibrachiatum was 3.35 U min⁻¹ ml⁻¹ at the 5^th day after the treatment of second stage juveniles. In greenhouse experiments, the concentrations of T. longibrachiatum ranging from 1.5 × 10⁵ to 1.5 × 10⁷ conidia ml⁻¹ all decreased M. incognita infection and increased plant height, root length, shoot and root fresh weights in cucumber significantly compared to the control. T. longibrachiatum can be considered to be a promising bio-control agent against M. incognita with a high efficacy.

Introduction

Plant parasitic nematodes are one of the most important soil-borne pathogens (Bird and Kaloshian, 2003), and annual yield losses due to plant parasitic nematodes have been estimated to be $100 billion USD worldwide (Sikora and Fernández, 2005). Root-knot nematode species (Meloidogyne spp.) are among the most widely spread agricultural pests in the world, and their extensive distribution cause various types of root-knot diseases in many field crops and horticultural species (Sikora and Fernández, 2005).

In recent years, cucumber (Cucumis sativus L.) has been grown as a vegetable crop on a large scale in China. However, its production is seriously threatened by root-knot nematodes Meloidogyne spp. (Sikora and Fernández, 2005). Four Meloidogyne species are frequently found in the major cucumber production area of eastern China, with Meloidogyne incognita being the most dominant species (Wang et al., 2001). M. incognita is parasitic, and its second-stage juveniles (J2s) can penetrate and enter the crop roots to establish permanent feeding sites. Infected plants exhibit the formation of knots in root tissues, and the abundance of knots subsequently affects the uptake of nutrients and water (Milligan et al., 1998), leading to increased plant sensitivity to pathogen infection and reduced yield (Castagnone-Sereno et al., 1992).

A number of strategies have been developed to control root-knot nematode in crop production, including the use of chemical and biological nematicides. Chemical nematicides are effective in most cases, but they create environmental pollution and can be toxic to humans (El-Alfy and Schlenk, 2002). Therefore, the development of alternative methods such as biological control is of great importance. Among biological control measures is the use of nematophagous fungi because these fungi have the ability to trap or hyperparasitize the nematodes (Huang et al., 2004). For example, the egg-parasitic fungi Paecilomyces sp. (Khan et al., 2004) and Pochonia sp. (Tikhonov et al., 2002) can infect nematode eggs and repress the hatching of juveniles, thereby reducing nematode populations. Although egg-parasitic fungi could be used for biological control of root-knot nematode, only a limited number of fungal control agents are commercially available (Kerry, 1997). Most importantly, some species of control agents are not safe to human health. For example, Paecilomyces spp. has been reported to be involved in onychomycoses (Hilmioglu-Polat et al., 2005). The fungus Paecilomyces lilacinus causes hyalohyphomycosis in renal transplant patients (Castro et al., 1990). Therefore, more effective strategies for the control of M. incognita are needed.

Trichoderma spp. is a class of fungi widely distributed in soil and active mycoparasites, and the fungi have been considered for biocontrol of foliar (Mora and Earle, 2001) and soil-borne diseases (Larralde-corona et al., 2008, Monga, 2012). Among the antagonisms, Trichoderma harzianum can provide excellent control (Sharon et al., 2001), and it is viewed as a strong contender for the development of bio-control agents to control some plant diseases in vitro and fields. Some bio-control agents have been broadly used in the United State and Israel (Elad, 2000, Alessandro et al., 2012), and attempts have been made to use Trichoderma spp. to control plant parasitic nematodes. T. harzianum is one of the bio-agents effective for the management of the citrus nematode (Sharon et al., 2001). Trichoderma longibrachiatum has been shown to have a strong parasitic and lethal effect on the cysts of Heterodera avenae in vitro (Zhang et al., 2014a). Also, AL-Shammari et al. (2013) revealed that T. longibrachiatum and Mortierella alpina can be used as a bio-agent for the sustainable management of root-knot nematodes on selected crops. Several factors may limit the efficacy of fungal bio-control agents, including temperature, moisture, texture and structure of soil, nematode density and multiplication, and fungal isolates (Kerry, 1997). However, there is little information available on the effectiveness of using Trichoderma spp. to control plant parasitic nematodes, and specially lack of knowledge about the process of T. longibrachiatum in the control of M. incognita second stage juveniles. In the present study, we used both laboratory and greenhouse experiments to (i) evaluate the ability and effectiveness of T. longibrachiatum in the control of M. incognita second stage juveniles, and (ii) determine the probable mechanism with which T. longibrachiatum functions against the second stage juveniles of M. incognita.