Colonization of Beauveria bassiana 08F04 in root-zone soil and its biocontrol of cereal cyst nematode (Heterodera filipjevi).

Jie Zhang,Qitong Lin,Linlin Chen,Ian T Riley,Bo Fu,Jiangkuan Cui,Honglian Li,Lirong Yang,Shengli Ding,Zonghua Wang

doi:10.1371/journal.pone.0232770

Abstract

Cereal cyst nematodes cause serious yield losses of wheat in Hunaghuai winter wheat growing region in China. Beauveria bassiana 08F04 isolated from the surface of cysts is a promising biological control agent for cereal cyst nematodes. As the colonization capacity is a crucial criteria to assess biocontrol effectiveness for a microbial agent candidate, we aimed to label B. bassiana 08F04 for efficient monitoring of colonization in the soil. The binary pCAM-gfp plasmid containing sgfp and hph was integrated into B. bassiana 08F04 using the Agrobacterium tumefaciens-mediated transformation. The transformation caused a significant change in mycelial and conidial yields, and in extracellular chitinase activity in some transformants. The cultural filtrates of some transformants also decreased acetylcholinesterase activity and the survival of Heterodera filipjevi second-stage juveniles relative to the wild-type strain. One transformant (G10) had a growth rate and biocontrol efficacy similar to the wild-type strain, so it was used for a pilot study of B. bassiana colonization conducted over 13 weeks. Real-time PCR results and CFU counts revealed that the population of G10 increased quickly over the first 3 weeks, then decreased slowly over the following 4 weeks before stabilizing. In addition, the application of wild-type B. bassiana 08F04 and transformant G10 significantly reduced the number of H. filipjevi females in roots by 64.4% and 60.2%, respectively. The results of this study have practical applications for ecological, biological and functional studies of B. bassiana 08F04 and for bionematicide registration.

Highlights

Cereal cyst nematodes (CCNs) are recognized as the most important group of plant-parasitic nematodes of cereal crops and cause substantial yield losses in combination with other biotic and abiotic factors [1,2]
Colonization of Beauveria bassiana 08F04and its biocontrol of cereal cyst nematode containing the pCAM-gfp plasmid resulted in the formation of positive colonies on the selective medium, whereas no colonies appeared when 08F04 was incubated with A. tumefaciens without plasmids
Colonization of Beauveria bassiana 08F04and its biocontrol of cereal cyst nematode isolated from the CCN females in wild-type 08F04 treatment and control on the potato dextrose agar (PDA) plate supplemented with hygromycin B (500 μg/mL), streptomycin (100 μg/mL) and kanamycin (50 μg/mL)

Summary

Introduction

Cereal cyst nematodes (CCNs) are recognized as the most important group of plant-parasitic nematodes of cereal crops and cause substantial yield losses in combination with other biotic and abiotic factors [1,2]. Of the 12 recognized CCN species, Heterodera filipjevi is one of the most damaging, causing yield losses that can reach 50% in individual fields [3,4]. Colonization of Beauveria bassiana 08F04and its biocontrol of cereal cyst nematode

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