Biocontrol of Ralstonia solanacearum by Treatment with Lytic Bacteriophages

Abstract

Ralstonia solanacearum is a Gram-negative bacterium and the causative agent of bacterial wilt in many important crops. We treated R. solanacearum with three lytic phages: RSA1, RSB1, and RSL1. Infection with RSA1 and RSB1, either alone or in combination with the other phages, resulted in a rapid decrease in the host bacterial cell density. Cells that were resistant to infection by these phages became evident approximately 30 h after phage addition to the culture. On the other hand, cells infected solely with RSL1 in a batch culture were maintained at a lower cell density (1/3 of control) over a long period. Pretreatment of tomato seedlings with RSL1 drastically limited penetration, growth, and movement of root-inoculated bacterial cells. All RSL1-treated tomato plants showed no symptoms of wilting during the experimental period, whereas all untreated plants had wilted by 18 days postinfection. RSL1 was shown to be relatively stable in soil, especially at higher temperatures (37 to 50°C). Active RSL1 particles were recovered from the roots of treated plants and from soil 4 months postinfection. Based on these observations, we propose an alternative biocontrol method using a unique phage, such as RSL1, instead of a phage cocktail with highly virulent phages. Using this method, RSL1 killed some but not all bacterial cells. The coexistence of bacterial cells and the phage resulted in effective prevention of wilting.

Bacteriophages of Ralstonia solanacearum: Their Diversity and Utilization as Biocontrol Agents in Agriculture

“…Either RSA1 or RSB1, which kills cells quickly, could not bring about similar plant-protecting effects. Using these observations, Fujiwara et al (2011) proposed an alternative phage biocontrol method using a unique phage, such as RSL1, instead of a phage cocktail containing highly lytic phages. With this method, bacterial cells are not killed completely, but a sustainable state of phagebacteria coexistence (with a low level of bacterial population) is maintained.…”

“…A phage cocktail has been recommended to prevent the problem of resistance, which contains several phages with different host specificities, different replication mechanisms, and/or different infection cycles (Gill and Abedon, 2003;. Fujiwara et al (2011) used three lytic phages, RSA1, RSB1, and RSL, for biocontrol of tomato bacterial wilt caused by R. solanacearum. Although RSA1 and RSB1 infection resulted in quick lysis of host cells, multi-resistant cells arose approximately 30 h post infection.…”

Characterization of Bacteriophages Cp1 and Cp2, the Strain-Typing Agents for Xanthomonas axonopodis pv. citri

“…The nucleotide sequences of these bands were determined. By comparing the nucleotide sequences with each other and with the Cp1 genomic sequence, the cos sequence was determined according to the method of Fujiwara et al (24).…”

Smart biologics for crop protection in agricultural systems

“…Boulé et al (2011) In greenhouse trials, pretreatment of tomato seedlings with RSL1 prevented bacterial wilt in all plants Isolated and partly characterized in vitro for control of R. solanecearum. Fujiwara et al (2011) trigger the activation of a particular mode of action would provide a powerful tool to investigate evolutionary principles and applications of the specificity of biocontrol agents. However, it is imperative that scientists developing opportunistic pathogens continue to rigorously weigh the advantages and hazards posed by these new biocontrol agents and help regulatory authorities make decisions.…”