| Greenhouse vegetables play an important role in farmers’ income increasing,economic development and even agricultural modernization in Shanxi Province.Root-knot nematodes(Meloidogyne spp.)are the most frequently observed and damaging pests in vegetable production.They not only reduce crop yield but also affect product quality and finally cause considerable economic losses.Currently,chemical nematicides are the most efficient and reliable means of controlling nematodes.However,their active compounds are non-specific and notoriously toxic to human health and the environment,and most nematicides are being progressively banned or highly restricted for protecting vegetable production.Research on agents that work against root-knot nematodes and do not have a detrimental impact on the environment is being increasingly important.In this study,111 bacterial strains were isolated from the rhizosphere soil,and six strains were screened for having a strong inhibitory effect on the second-stage juveniles(J2)of M.incognita;these six strains were identified using morphological,physiological and biochemical characteristics,16 S r RNA gene sequences and Bayesian phylogenetic relationship analysis.Two strains,A-27 and A-30,were chose to conduct the inhibit effects on J2 and eggs,pot control efficacy,stability,fermentation optimization and field control efficacy.The results are as follows:1.In this study,111 strains of bacteria were isolated from rhizosphere soil using dilution coating plate method.Among these,51 strains fermentation broth exhibited high ability to kill J2.However,six strains exhibited distinct effect and recorded more than 80% J2 mortality after 24 h.A-9,A-22,A-25,A-27,A-29 and A-30 had the mortality percentage of 84.54%,83.17%,97.41%,100.00%,92.64% and 81.62%,respectively.Based on morphological,physiological and biochemical characteristics,16 S r RNA gene sequences and Bayesian phylogenetic relationship analysis,A-9 was identified as Stenotrophomonas maltophilia,A-22 and A-25 both were Pseudomonas brassicacearum,A-27 and A-30 both were Bacillus velezensis,A-29 was Ochrobactrum pseudogrignonense.Strains A-27 and A-30 were chosen for the subsequent experiments.2.The M.incognita J2 mortality was tested with three different concentrations of fermentation broth,fermentation filtrate and bacterial suspension of strains A-27 and A-30.The results showed that J2 mortality treated with fermentation broth and fermentation filtrate were higher than bacterial suspension,the overall inhibitory effect of fermentation broth was the best.In the pot experiment,tomato plant treated with fermentation broth of A-27 and A-30 exhibited a drastic reduction in root gall index by 84.61% and 76.92%,improvement in whole fresh weight by 17.20% and 8.00%,and improvement in whole height by 38.03% and 12.41%,and increase in roots dry weight by 32.46% and 14.53%,respectively.3.There was a maximum absorption at a wavelength of 400 nm for the fermentation broth of strains A-27 and A-30.The stability study revealed that strain A-27 grew well in light or dark,between 50 to 100℃,pH 4 to8 and under ultraviolet irradiation.Strain A-30 was insensitive to light or dark,pH value and ultraviolet,but sensitive to temperature and it grew unstably between 50 to 100℃.4.The parameters for fermentation were optimized using the one-variable-at-a-time approach and orthogonal test design.The results showed that the optimal fermentation medium for strain A-27 consisted of 15 g sucrose,50 g peptone,10 g magnesium sulfate,pH 7.0 and added distilled water to make a total volume of 1000 m L;optimum culture conditions were 28℃ fermentation temperature,100 m L in 250 m L liquid volume and 7%(7 m L)inoculation volume.In addition,the optimal fermentation medium for A-30 strain consisted of15 g sucrose,50 g peptone,10 g dipotassium hydrogen phosphate,pH 7.0 and added distilled water to make a total volume of 1000 m L;optimum culture conditions were 30℃ fermentation temperature,125 m L in 250 m L liquid volume and 3%(3.75 ml)inoculation volume.5.In greenhouse experiments,celery plants from M.incognita J2-infested soils treated with the fermentation broth of strains A-27 and A-30 exhibited a drastic reduction in root gall index and J2 density,while a significant improvement in top height and fresh weight of celery plants.The biocontrol efficacy of strains A-27 and A-30 reached up to 64.29% and 60.71%,respectively.The diversity of rhizosphere soil microorganisms was measured by high-throughput sequencing.The results revealed that strains A-27 and A-30 fermentation broth changed the community structure of soil microorganisms and improved the abundance of bacteria and fungi under the rhizosphere of celery.Proteobacteria,Actinobacteria,Chloroflexi,Firmicutes and Acidobacteria were the dominant bacteria groups,while Ascomycota,Basidiomycota and Mortierellomycota were the dominant fungi groups.The functional prediction analysis revealed that the functional structure of bacteria did not change obviously,whereas the functional structure of fungi changed significantly,and the relative abundance of functions related to animal and plant pathogens significantly decreased. |
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