一株中药渣源溶磷菌的全基因组分析及功能验证

doi:10.13560/j.cnki.biotech.bull.1985.2025-0949

摘要/Abstract

摘要：

目的明确中药渣源不动杆菌DF1的溶磷机制，为高效溶磷工程菌株构建及生物菌肥开发提供理论基础与技术支持。方法通过培养DF1菌株、提取基因组DNA进行测序组装与多数据库功能注释，克隆溶磷基因pqqB和pqqC并构建重组载体转化大肠杆菌，经筛选鉴定后测定重组菌株溶磷能力。结果 DF1基因组全长3 831 735 bp、GC含量38.9%，共预测含3 482个蛋白质编码基因、91个非编码RNA，且基因组中富集多种与溶磷代谢相关的功能基因。克隆溶磷关键基因pqqB和pqqC，构建重组表达载体并转化大肠杆菌，经抗性筛选与PCR鉴定获得阳性重组菌株。溶磷能力测定结果表明，两株重组菌株均具备显著的无机磷与有机磷溶解能力，且能分泌草酸、酒石酸、苹果酸、乙酸及柠檬酸5种有机酸，其中柠檬酸分泌量分别高达（1 753.10±156.38）mg/L和（1 720.30±206.35）mg/L。结论首次阐明了中药渣源不动杆菌DF1的基因组特征，明确了其溶磷分子机制，并证实pqqB和pqqC基因在溶磷过程中发挥核心作用。

关键词: 不动杆菌, 全基因组测序, 溶磷菌, 生物菌肥

Abstract:

Objective To clarify the phosphate-solubilizing mechanism of Acinetobacter sp. DF1 isolated from traditional Chinese medicine residues, and provide a theoretical basis and technical support for the construction of high-efficiency phosphate-solubilizing engineered strains and the development of biological bacterial fertilizers. Method The DF1 strain was cultured, and its genomic DNA was extracted for sequencing, assembly and functional annotation against multiple databases. The phosphate-solubilizing genes pqqB and pqqC were cloned, recombinant vectors were constructed and transformed into Escherichia coli. After screening and identification, the phosphate-solubilizing capacity of the recombinant strains was determined. Result The results showed that the genome of DF1 was 3 831 735 bp in full length with a GC content of 38.9%, which was predicted to contain 3 482 protein-coding genes and 91 non-coding RNAs, and was enriched in various functional genes related to phosphate-solubilizing metabolism. The key phosphate-solubilizing genes pqqB and pqqC were cloned, and recombinant expression vectors were constructed and transformed into E. coli. Positive recombinant strains were obtained through resistance screening and PCR identification. The results of the phosphate-solubilizing capacity assay demonstrated that both recombinant strains exhibited significant ability to solubilize inorganic and organic phosphorus, and secreted five organic acids including oxalic acid, tartaric acid, malic acid, acetic acid and citric acid, among which the citric acid secretion levels reached as high as (1 753.10 ± 156.38) mg/L and (1 720.30 ± 206.35) mg/L, respectively. Conclusion This study first clarifies the genomic characteristics of Acinetobacter sp. DF1 isolated from traditional Chinese medicine residues, elucidates the molecular mechanism of phosphate solubilization, and confirms that the pqqB and pqqC genes play a core role in the phosphate-solubilizing process.

Key words: Acinetobacter, whole-genome sequencing, phosphorus-solubilizing bacteria, biological bacterial fertilizer

丁洪霞, 高贵文, 黄玉兰, 赵蕊, 黄奕博, 冯和鑫, 龙莎. 一株中药渣源溶磷菌的全基因组分析及功能验证[J]. 生物技术通报, 2026, 42(7): 304-314.

DING Hong-xia, GAO Gui-wen, HUANG Yu-lan, ZHAO Rui, HUANG Yi-bo, FENG He-xin, LONG Sha. Whole-genome Analysis and Functional Verification of a Phosphate-solubilizing Bacterium Isolated from Traditional Chinese Medicine Residues[J]. Biotechnology Bulletin, 2026, 42(7): 304-314.

图/表 14

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ARO accession	ARO name	Drug class	Gene ID
ARO：3000777	adeF	Fluoroquinolone antibiotic； tetracycline antibiotic	gene1142
ARO：3003818	Acinetobacter baumannii parC conferring resistance to fluoroquinolones	Fluoroquinolone antibiotic	gene3355
ARO：3003817	Acinetobacter baumannii gyrA conferring resistance to fluoroquinolones	Fluoroquinolone antibiotic； nybomycin-like antibiotic	gene0858
ARO：3003685	Pseudomonas aeruginosa parE conferring resistance to fluoroquinolones	Fluoroquinolone antibiotic	gene0128
ARO：3000775	adeB	Glycylcycline； tetracycline antibiotic	gene1698
ARO：3004577	Acinetobacter baumannii AmvA	Disinfecting agents and antiseptics； macrolide antibiotic	gene1413
ARO：3000778	adeG	Fluoroquinolone antibiotic； tetracycline antibiotic	gene1143
ARO：3004573	Acinetobacter baumannii AbaF	phosphonic acid antibiotic	gene2185
ARO：3000753	abeM	Disinfecting agents and antiseptics； fluoroquinolone antibiotic	gene3173
ARO：3003809	Acinetobacter baumannii OprD conferring resistance to imipenem	Carbapenem； cephalosporin； cephamycin； monobactam； penam；penem	gene3350
ARO：3004578	Acinetobacter baumannii AbuO	Aminoglycoside antibiotic；carbapenem； cephalosporin； disinfecting agents	gene3308
ARO：3004574	Acinetobacter baumannii AbaQ	Fluoroquinolone antibiotic	gene1608

ARO accession	ARO name	Drug class	Gene ID
ARO：3000777	adeF	Fluoroquinolone antibiotic； tetracycline antibiotic	gene1142
ARO：3003818	Acinetobacter baumannii parC conferring resistance to fluoroquinolones	Fluoroquinolone antibiotic	gene3355
ARO：3003817	Acinetobacter baumannii gyrA conferring resistance to fluoroquinolones	Fluoroquinolone antibiotic； nybomycin-like antibiotic	gene0858
ARO：3003685	Pseudomonas aeruginosa parE conferring resistance to fluoroquinolones	Fluoroquinolone antibiotic	gene0128
ARO：3000775	adeB	Glycylcycline； tetracycline antibiotic	gene1698
ARO：3004577	Acinetobacter baumannii AmvA	Disinfecting agents and antiseptics； macrolide antibiotic	gene1413
ARO：3000778	adeG	Fluoroquinolone antibiotic； tetracycline antibiotic	gene1143
ARO：3004573	Acinetobacter baumannii AbaF	phosphonic acid antibiotic	gene2185
ARO：3000753	abeM	Disinfecting agents and antiseptics； fluoroquinolone antibiotic	gene3173
ARO：3003809	Acinetobacter baumannii OprD conferring resistance to imipenem	Carbapenem； cephalosporin； cephamycin； monobactam； penam；penem	gene3350
ARO：3004578	Acinetobacter baumannii AbuO	Aminoglycoside antibiotic；carbapenem； cephalosporin； disinfecting agents	gene3308
ARO：3004574	Acinetobacter baumannii AbaQ	Fluoroquinolone antibiotic	gene1608

VFDB ID	Vfs	Related genes	Product	Gene ID
VFG037612	Csu fimbriae （VF0461）	csuD	Csu pilus usher protein CsuD	gene1238
VFG037177	Phospholipase C （VF0470）	plc2	Phospholipase C	gene1415
VFG050742	T2SS （VF1335）	gspD	General secretion pathway protein D	gene3294
VFG037358	Acinetobactin （VF0467）	bauA	TonB-dependent siderophore receptor BauA	gene1064
VFG037176	Phospholipase C （VF0470）	plc1	Phospholipase C	gene3508
VFG050670	TFP （VF1334）	pilJ	Methyl-accepting chemotaxis protein	gene0667
VFG050372	TFP （VF1334）	pilQ	Type IV pilus secretin PilQ	gene0335
VFG050303	Ata （VF1333）	ata	Trimeric autotransporter adhesin Ata	gene2598
VFG051098	T6SS （VF1337）	clpV/tssH	Type VI secretion system ATPase TssH	gene2210
VFG037330	Acinetobactin （VF0467）	basD	Acinetobactin biosynthesis protein BasD	gene1066
VFG050959	T6SS （VF1337）	tssM	Type VI secretion system membrane subunit TssM	gene2214
VFG051070	T6SS （VF1337）	tssC	Type VI secretion system contractile sheath large subunit	gene2220
VFG050470	TFP （VF1334）	pilC	Type II secretion system F family protein	gene3237
VFG050428	TFP （VF1334）	fimV	Hypothetical protein	gene3140

VFDB ID	Vfs	Related genes	Product	Gene ID
VFG037612	Csu fimbriae （VF0461）	csuD	Csu pilus usher protein CsuD	gene1238
VFG037177	Phospholipase C （VF0470）	plc2	Phospholipase C	gene1415
VFG050742	T2SS （VF1335）	gspD	General secretion pathway protein D	gene3294
VFG037358	Acinetobactin （VF0467）	bauA	TonB-dependent siderophore receptor BauA	gene1064
VFG037176	Phospholipase C （VF0470）	plc1	Phospholipase C	gene3508
VFG050670	TFP （VF1334）	pilJ	Methyl-accepting chemotaxis protein	gene0667
VFG050372	TFP （VF1334）	pilQ	Type IV pilus secretin PilQ	gene0335
VFG050303	Ata （VF1333）	ata	Trimeric autotransporter adhesin Ata	gene2598
VFG051098	T6SS （VF1337）	clpV/tssH	Type VI secretion system ATPase TssH	gene2210
VFG037330	Acinetobactin （VF0467）	basD	Acinetobactin biosynthesis protein BasD	gene1066
VFG050959	T6SS （VF1337）	tssM	Type VI secretion system membrane subunit TssM	gene2214
VFG051070	T6SS （VF1337）	tssC	Type VI secretion system contractile sheath large subunit	gene2220
VFG050470	TFP （VF1334）	pilC	Type II secretion system F family protein	gene3237
VFG050428	TFP （VF1334）	fimV	Hypothetical protein	gene3140

抗生素名称 Antibiotic name	每片含量 Content per tablet （μg）	抑制圈直径 Diameter of inhibition zone （mm）	敏感性 Sensitivity
丁胺卡那（AK）	30	14.75	中介
复方新诺明（SXT）	23.75/1.25	11.5	耐药
红霉素（E）	15	16	敏感
环丙沙星（CIP）	5	23	敏感
氯霉素（C）	30	20.5	敏感
诺氟沙星（NOR）	10	16.75	敏感
庆大霉素（GM）	10	14.25	中介