发酵料中促平菇生长菌株的分离与鉴定

Isolation and Identification of Oyster Mushroom Growth-Promoting Bacteria from Fermented Materials

平菇发酵培养料含有吲哚乙酸(IAA),IAA促进平菇生长发育。为了找到产生IAA能力强的微生物菌株,首先,采集不同时期的平菇发酵料样品,采取稀释涂布平板法分离细菌菌株。其次,挑选120株菌利用显色反应进行产IAA能力的定性分析及定量测定,并对高产IAA菌株进行平菇促生试验研究。最后,利用GUTC法提取产IAA能力强的菌株的基因组DNA并进行16S rRNA基因扩增及测序分析,基于构建的16S rRNA基因进化树进行系统发育分析,确定菌株的分类学地位。结果表明,120株菌中有116株菌产生显色反应,即证明这116株菌有产IAA的能力。相对于对照组显色程度,有88株菌的IAA显色程度较低,28株菌的IAA显色程度相对较高。挑取这28株菌进行定量测定,其中,AFX-13-3、XWW-4-3、LTX-11-3、AMB-8-2、AMB-10-2、AMB-14-3、ST-1-2、ST-3-2、ST-4-2等9株菌所产IAA的质量浓度达到20 mg/L以上。AFX-13-3产IAA的能力最强,其IAA平均质量浓度达到85.18 mg/L;其次为AMB-10-2、ST-3-2、AMB-8-2、XWW-4-3,其IAA平均质量浓度分别为39.35、34.12、30.94、30.71 mg/L;ST-1-2、ST-4-2、LTX-11-3、AMB-14-3产IAA的能力相对较弱,其IAA平均质量浓度分别为24.41、24.09、22.26、21.94 mg/L。平菇促生试验结果表明,AFX-13-3、XWW-4-3、LTX-11-3均有较好的促生作用。通过16S rRNA基因测序和系统发育分析,确定AFX-13-3属于巨大芽孢杆菌属,XWW-4-3、LTX-11-3属于谷氨酸杆菌属。

The fermentation culture of oyster mushroom contains indoleacetic acid(IAA),which promotes the growth and development of oyster mushroom.In order to find the microbial strains with strong IAA production ability,firstly,samples of oyster mushroom fermentation material at different time were collected,and bacterial strains were separated by dilution coating plate method.Secondly,120 strains were selected for qualitative analysis and quantitative determination of IAA-producing capacity by chromogenic reaction,and high-yield IAA strains were used to test the oyster mushroom growth-promoting effect.Finally,the genomic DNA of the IAA-producing strains was extracted by GUTC,16S rRNA gene amplification and sequencing analysis were carried out,and phylogenetic analysis was performed based on the constructed 16S rRNA gene phylogenetic tree to determine the taxonomic status of the strains.The results showed that in 120 bacterial strains isolated,116 strains produced color reaction,which were proved to have the ability to produce IAA.Compared with the control group,IAA color development degree of 88 strains was lower,28 strains were relatively higher.28 strains with relatively high IAA color rendering degree after qualitative determination were selected for quantitative determination,among them,AFX-13-3,XWW-4-3,LTX-11-3,AMB-8-2,AMB-10-2,AMB-14-3,ST-1-2,ST-3-2,ST-4-2 could produce IAA more than 20 mg/L.Among the nine strains,AFX-13-3 had the strongest ability to produce IAA,with average mass concentration of 85.18 mg/L,followed by AMB-10-2,ST-3-2,AMB-8-2 and XWW-4-3,with the average mass concentrations of IAA of 39.35,34.12,30.94 and 30.71 mg/L,respectively.The ability of the remaining four strains ST-1-2,ST-4-2,LTX-11-3 and AMB-14-3 to produce IAA was relatively weaker,and the average concentrations of IAA were 24.41,24.09,22.26and 21.94 mg/L,respectively.After the growth promotion test of oyster mushroom,it was found that AFX-13-3,XWW-4-3 and LTX-11-3 had good growth promotion effect on oyster mushroom.Through 16S rRNA gene sequencing and phylogenetic analysis,it was determined that AFX-13-3 belonged to Priestia,and XWW-4-3 and LTX-11-3 belonged to Glutamicibacter.