Elsevier

Rhizosphere

Volume 19, September 2021, 100390
Rhizosphere

Biocontrol bacteria strains Y4 and Y8 alleviate tobacco bacterial wilt disease by altering their rhizosphere soil bacteria community

https://doi.org/10.1016/j.rhisph.2021.100390Get rights and content

Abstract

Tobacco (Nicotiana tabacum L.) bacterial wilt caused by Ralstonia solanacearum was widespread in recent years, resulting in a great damage in tobacco production. Biological control is an effective measurement for the bacterial wilt, however, related work has not been researched deeply. By screening the biocontrol microorganisms in Wenshan tobacco growing fields in Yunnan province, we discovered a Bacillus amyloliquefaciens strain Y4 and a Pseudomonas sp. strain Y8 that could significantly suppress the growth of R. solanacearum in the culture. The occurrence of wilt disease by field application of Y4 and/or Y8 was 3–4 folds lower than that of the untreatment at different tobacco growth stages. The microbial composition analysis showed that the relative abundance of the Enterobacteriaceae, Sphingomonadaceae, Caulobacteraceae, Sphingobacteriaceae, Flavobacteriacea, Streptomycetaceae, and Burkholderoaceae were significantly increased in the rhizosphere soils of infected tobacco. Moreover, field application of Y4 or Y8 significantly changed the rhizosphere soil microbiota. The relative abundance of the Lactobacillus was increased, however, that of R. solanacearum, Pseudoxanthomonas mexicana, Sphingomonas koreensis, Variovorax paradoxus, and Bradyrhizobium elkanii were decreased. Our results indicate that biocontrol microorganisms reduce plant disease occurrence by changing their microbiota in rhizosphere soil.

Introduction

Bacterial wilt disease is caused by the causal agent Ralstonia solanacearum, a soil born bacterial pathogen. R. solanacearum infects over 200 plant species, including tobacco (Nicotiana tobaccum), tomato (Lycopersicon esculentum), and Pepper (Capsicum annuum) (Denny, 2007). The pathogen primarily infects the plants through wounding or nature opening pores. Once the pathogen successfully colonizes a plant, it develops in the xylem and fills the xylem, leading to the clog of plant xylem, which eventually blocks the water and nutrient transportation (Hikichi et al., 2017). In many cases, the infected plants wilt or die. The tobacco wilt disease has been occurring frequently in the tobacco growing area in south of China during the past decade, resulting in great yield losses (Jiang et al., 2017). The wilt disease could occur at any tobacco plant growth stages. Among all the economic crops in China, tobacco wilt disease is the most sever and prevalent disease due to continuous cropping culture without rotation.

Current measurements to control wilt disease are mostly through the resistant cultivars, the chemical pesticides, and agricultural management (Wang et al., 2020). However, these management techniques are inefficient, do not provide durable resistance for the plants. Furthermore, the chemical controls are being challenged as they are not environment friendly, which typically is either poisonous or has pesticide residue (Quadroni and Bettinetti, 2019). In fact, pesticide residue has profoundly affected the tobacco quality. Biological control has been considered as the potential alternative approaches which does not have the above-mentioned drawbacks and has been widely accepted by farmers. The known biocontrol agents include the plant-derived bioactive compounds that can either suppress or kill pathogens (Li et al., 2018). Some biocontrol agents are antibiotics or the antagonistic microorganisms. Plant pathogens can be inhibited by many means, which in soil can be suppressed by other competitive microorganisms. The competitors could compete for the nutrition or secrete antibiotics to limit pathogen growth (Li et al., 2019). The induced defense of plant immune system could also suppress pathogen colonization (Olanrewaju et al., 2017). Bacillus species has been used as a biocontrol agent against many plant pathogens through various supressing mechanisms, and their derived products account for approximately half of the commercially available biopesticides (Wei et al., 2011; Fira et al., 2018). However, the control of bacterial wilt is difficult and ineffective due to the high genetic variability, persistence in the environment, and broad host range of R. solanacearum (Yuliar et al., 2015). Thus, the alternative antimicobial agents are needed to explored.

The rhizobacterial community is very important for both plant growth and health (Wang et al., 2017). It plays critical roles in regulating soil fertility, cycling of nutrients, and protecting plants from diseases (Wang et al., 2017). The rhizosphere soil contains thousands of beneficial plant rhizobacteria. On the other side, it also harbors pathogenic microorganisms, which cause plant disease by breaking the protective microbial shield and overcoming the plant's innate defense mechanisms (Mendes et al., 2011). Pathogen infection could alter the structure of the rhizosphere microbial community and influence the constituent of soil microorganisms (Mazzola, 2004). For example, R. solanacearum invasion significantly affected the diversity and composition of the bacterial community in the tomato rhizosphere, leading to reductions in bacterial diversity and the abundance of nonpathogenic bacteria (Wei et al., 2018). Thus, altering the soil microbiota has been considered to be a promising strategy to control plant diseases in the fields (Fu et al., 2017). However, it is still unclear for the soil microbial community structure in the bacterial wilt infected soils, especially in the antimicobial agents treated rhizosphere.

In this study, we evaluated the rhizosphere microbial community differences between bacterial wilt infected soils and healthy soils. In addition, we found several bacteria strains that could significantly suppress the wilt disease development. Among them, a Bacillus amyloliquefaciens and a Pseudomonas sp. strain could promote the tobacco plant disease resistance by modulating the rhizosphere soil microbial community. This information provides more biocontrol resources for the control of bacterial wilt.

Section snippets

Pathogen and reagents

R. solanacearum strain RS-22 used in this study and the antagonistic bacteria Bacillus amyloliquefaciens strain Y4 and Pseudomonas sp. strain Y8 were isolated in our laboratory from the infected tobacco of bacterial wilt from the fields in Yunnan Province, China (Li et al., 2021). The beef extract peptone medium (Trypton 10 g/L, beef extract 3 g/L, NaCl 10 g/L and agar 15 g/L) was used to culture the bacteria.

Biocontrol activity against tobacco bacteria wilt in greenhouse

The antagonistic bacteria Y4 and Y8 were cultured in liquid beef extract peptone

Rhizosphere bacterial community of healthy and R. solanacearum-infected tobacco

To explore the variation of bacterial community in tobacco rhizosphere, we collected the rhizosphere soil samples from the healthy and R. solanacearum-infected tobacco plants in the fields of Wenshan county (hereafter abbreviated as CK and TBW, respectively), and the 16S rRNA amplicon sequencing technology was used to sequence the soil microbiome subsequently. A total of 496, 893 raw reads were obtained, and 468, 494 high quality clean reads were remained after quality filtering, which contains

Discussion

Tobacco wilt disease is one of the notorious diseases in tobacco plants. Although many agricultural practices and pesticides have been applied, the wilt disease is currently out of control in the tobacco fields of Yunnan province. In addition, the diverse isolates of R. solanacearum in different tobacco growing regions further increase the difficulties in controlling the wilt disease (Huet G, 2014). We have isolated the R. solanacearum-22 from Wenshan fields in Yunnan Province (Li et al., 2021

Funding information

This study was supported by the Key Project of Yunnan Tobacco Company (2018530000241017, 2017YN06).

Author contributions

J.L. and C.Y. performed most of the experiments; Q.Z. and H.W. performed the data analysis; Q.Z. and C.Y. wrote the article.

Declaration of competing interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

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