广西北部湾可培养黏细菌资源初探及其抗菌活性研究

doi:10.11978/2022142

热带海洋学报 ›› 2023, Vol. 42 ›› Issue (3): 158-168.doi: 10.11978/2022142CSTR: 32234.14.2022142

广西北部湾可培养黏细菌资源初探及其抗菌活性研究

卢天梅¹(), 官佳松¹^,², 覃诗敬¹, 刘永宏¹^,³, 苏志维¹()

1.广西中医药大学海洋药物研究院, 广西南宁 530200
2.广西大学农学院, 广西南宁 530004
3.热带海洋生物资源与生态重点实验室(中国科学院南海海洋研究所), 广东省海洋药物重点实验室, 广东广州 510301

收稿日期:2022-06-23 修回日期:2022-08-24 出版日期:2023-05-10 发布日期:2022-09-01
作者简介:
卢天梅(1994—), 女, 广西壮族自治区桂平市人, 硕士研究生, 从事海洋药用微生物资源挖掘与利用研究。email: 2646013152@qq.com
基金资助:
广西中医药大学海洋药物研究院团队科研专项经费(2018ZD005-A08); 国家自然科学基金(32060098); 广西八桂学者专项经费(05019055); 广西中医药大学研究生教育创新计划项目(YCSZ2022024); 大学生创新项目(S202110600122); 广西科技计划项目(桂科AB19259010)

Preliminary study on the culturable myxobacteria resources from the Beibu Gulf, Guangxi and their antibacterial activity

LU Tianmei¹(), GUAN jiasong¹^,², QIN Shijing¹, LIU Yonghong¹^,³, SU Zhiwei¹()

1. Institute of Marine Drugs, Guangxi University of Chinese Medicine, Nanning 530200, China
2. College of Agriculture, Guangxi University, Nanning 530004, China
3. Key Laboratory of Tropical Marine Bio-resources and Ecology (South China Sea Institute of Oceanology, Chinese Academy of Sciences), Guangdong Key Laboratory of Marine Materia Medica, Guangzhou 510301, China

Received:2022-06-23 Revised:2022-08-24 Online:2023-05-10 Published:2022-09-01
Supported by:
Scientific Research Foundation of Institute of Marine Drugs, GUCM(2018ZD005-A08); National Natural Science Foundation of China(32060098); Special Fund for Bagui Scholars to Yonghong Liu(05019055); Innovation Project of Guangxi Graduate Education of GXUCM(YCSZ2022024); University Student Innovation Project(S202110600122); Natural Science Foundation of Guangxi Zhuang Autonomous Region(AB19259010)

摘要/Abstract

摘要：

采用大肠杆菌诱导法和滤纸诱导法对广西北部湾不同海洋生境采集的样品进行黏细菌的分离纯化, 基于菌落形态特征结合16S rRNA基因序列分析对黏细菌纯培养物进行初步鉴定。从21份海洋生境样品中共分离纯化得到38株黏细菌, 隶属于4属10种, 其中黏球菌属和珊瑚球菌属分别占菌株总数的63.16%和26.32%, 属于优势菌属。采用VY/2培养基进行固体培养发酵, 利用丙酮提取浓缩获得提取物, 进一步利用滤纸片琼脂扩散法对菌株粗提物进行抗菌活性初步筛选。结果发现24株黏细菌提取物对9种指示菌具有不同程度的抑制作用, 抗菌阳性率达63.16%, 其中Myxococcus macrosporus (1H01)的提取物对肺炎克雷伯菌的抑制活性显著, 优于阳性药(氨苄青霉素钠, 1mg·mL^-1)。研究结果表明广西北部湾特殊海洋生境黏细菌多样性丰富, 分离出的黏细菌具有良好的抗菌活性, 为新型抗生素应用研究奠定了物质基础。

关键词: 北部湾, 黏细菌, 分离鉴定, 抗菌活性

Abstract:

The objective of this study is to isolate myxobacteria from different types of marine biotopes in the Guangxi Beibu Gulf, on the base of living-Escherichia coli and filter paper methods. 38 strains were isolated from 21 samples, and were preliminary identified based on colony morphological characteristics and 16S rRNA gene sequence analysis. The purified strains belong to 4 genera and 10 species, in which Myxococcus and Corallococcus were dominant genera, accounting for 63.16% and 26.32% of the total strains, respectively. Then the purified strains were subjected to the small-scale fermentations through the VY/2 medium, and the extracts were obtained by the extraction and concentration of acetone. The crude extracts were further screened for antimicrobial activity by filter paper agar diffusion. The results showed that the metabolites of 24 myxobacterial strains had different degrees of inhibition on 9 indicator bacteria, and the positive rate was 63.16%. Among them, the metabolites of Myxococcus macrosporus (1H01) showed significant inhibitory activity against Klebsiella pneumoniae, which was superior to the positive drugs (ampicillin sodium, 1 mg·mL^-1). The results explain that the diversity of myxobateria is rich in different types marine sediment from the Guangxi Beibu Gulf, and the isolated myxobacteria has a prominent antibacterial activity, laying a foundation for the application of new antibiotics.

Key words: Beibu Gulf, myxobacteria, isolation and identification, antibacterial activity

卢天梅, 官佳松, 覃诗敬, 刘永宏, 苏志维. 广西北部湾可培养黏细菌资源初探及其抗菌活性研究[J]. 热带海洋学报, 2023, 42(3): 158-168.

LU Tianmei, GUAN jiasong, QIN Shijing, LIU Yonghong, SU Zhiwei. Preliminary study on the culturable myxobacteria resources from the Beibu Gulf, Guangxi and their antibacterial activity[J]. Journal of Tropical Oceanography, 2023, 42(3): 158-168.

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样品编号	样品名称	采样区域	经纬度
1H	木榄根际沉积物	珍珠湾	108°13′55″E, 21°36′27″N
2H	木榄腐木	珍珠湾	108°13′55″E, 21°36′27″N
3H	木榄胚轴	珍珠湾	108°13′55″E, 21°36′27″N
4H	木榄树皮	珍珠湾	108°13′55″E, 21°36′27″N
5H	木榄根际沉积物	珍珠湾	108°13′55″E, 21°36′27″N
6H	白骨壤根际沉积物	珍珠湾	108°13′55″E, 21°36′27″N
1D	海蚀坑沉积物	涠洲岛	109°05′52″E, 21°00′04″N
2D	滩岩老鼠簕根际沉积物	涠洲岛	109°05′58″E, 21°00′42″N
3D	礁石地衣	涠洲岛	109°05′56″E, 21°00′39″N
4D	滩岩仙人掌根际沉积物	涠洲岛	109°05′55″E, 21°00′15″N
1Y	盐田淤泥	银海区	109°16′39″E, 21°26′35″N
2Y	盐田淤泥	银海区	109°16′22″E, 21°26′35″N
3Y	盐田淤泥	银海区	109°16′23″E, 21°26′35″N
1C	海草床沉积物	山心村	108°12′06″E, 21°34′14″N
2C	互花米草根际沉积物	山心村	108°11′10″E, 21°35′11″N
3C	互花米草根际沉积物	山心村	108°10′57″E, 21°34′33″N
4C	贝壳喜盐草根际沉积物	山心村	108°11′29″E, 21°34′59″N
5C	贝壳喜盐草根际沉积物	山心村	108°11′21″E, 21°34′53″N
6C	贝壳喜盐草根际沉积物	山心村	108°11′20″E, 21°34′51″N
7C	贝壳喜盐草根际沉积物	山心村	108°11′36″E, 21°34′58″N
8C	贝壳喜盐草根际沉积物	山心村	108°11′24″E, 21°34′51″N

菌属	菌株数	子实体形态	菌落形态
Myxococcus	24	球状、卵球状, 单生, 多为橙色、白色或红粉色, 表面光滑柔软	圆形, 菌膜薄且呈同心圆扩展
Corallococcus	10	波浪状、肠状或珊瑚状, 单生或簇生, 多为淡黄色、粉色或乳白色, 表面不光滑、较硬	圆形, 菌膜薄且呈波浪纹扩展
Archangium	3	颗粒状, 簇生, 多为橘红色或棕色, 表面不光滑、较黏稠难挑	圆形, 菌膜薄、黏稠且呈放射状扩展
Melittangium	1	球状或扁平, 似蘑菇的菌伞, 分支生长, 黄褐色,表面不光滑	圆形, 菌膜薄、黏稠且呈放射状扩展

种名	菌株编号	相似度/%	耐盐能力/%
Corallococcus aberystwythensis	3C01	99.61	1
Corallococcus exercitus	3D01	98.97	1
Corallococcus exiguus	6C01	98.85	1
Melittangium boletus	2D03	99.49	<1
Archangium gephyra	2D01	100.00	1
	1Y01	99.74	2
	4D01	99.15	1
Corallococcus interemptor	1C01	100.00	1
	1D01	100.00	1
	2D02	100.00	1
	6C02	99.87	1
	8C01	99.87	1.5
	1C02	99.74	1
	4H01	99.35	1
Myxococcus fulvus	2C01	99.87	1
	1H02	99.74	<1
	5C01	99.36	1
	6H01	99.36	1
	1C04	98.85	1
	1D02	98.59	1
Myxococcus macrosporus	2Y01	100.00	1.5
	5H01	99.87	1.5
	1H01	99.74	1
	4H02	99.36	<1
	3Y01	99.10	2
	3H01	98.97	2
	1C03	98.87	1
Myxococcus stipitatus	4C01	99.87	1
	7C01	99.74	<1
	1C06	99.74	<1
	3D02	99.62	1
	1H03	99.50	1
	5H02	99.49	1
	1C05	99.49	1
Myxococcus virescens	8C02	99.49	2
	1Y02	99.36	2
	2H01	99.36	2
	1C07	99.10	1.5

采样生境	样品份数	菌种数	黏细菌菌株数				总计
采样生境	样品份数	菌种数	黏球菌属Myxococcus	珊瑚球菌属Corallococcus	原囊菌属Archangium	蜂窝囊菌属Melittangium	总计
红树林	6	5	9	1	-	-	10
海草床	8	7	10	6	-	-	16
海岛	4	6	2	3	2	1	8
盐田	3	3	3	-	1	-	4
总计	21	-	24	10	3	1	38

菌种	编号	抑菌圈直径/mm
		G⁺						G^-
		A	B	C	D	E	F	G	H	I
C. aberystwythensis	3C01	-	-	-	-	12.03±0.32	12.58±0.44	-	-	-
C. exiguus	6C01	-	-	-	-	-	9.28±0.14	-	-	-
A. gephyra	1Y01	-	-	-	-	12.07±0.04	-	-	-	-
C. interemptor	1C01	-	-	-	-	11.95±0.01	-	-	-	-
	1D01	-	-	-	-	13.43±0.40	8.64±0.09	12.25±0.29	-	-
	2D02	-	-	9.53±0.25	-	11.83±0.25	11.10±0.16	-	-	-
	6C02	-	-	-	-	-	9.97±0.05	-	-	-
	1C02	-	-	-	10.57±0.05	15.97±0.06	-	-	-	-
M. fulvus	2C01	-	-	-	10.33±0.32	14.93±0.11	11.54±0.44	-	-	-
	5C01	-	7.67±0.42	-	-	-	12.79±0.18	-	-	-
	1C04	10.82±0.04	7.34±0.02	12.03±0.65	12.23±0.24	15.79±0.05	14.05±0.20	10.91±0.17	11.14±0.07	13.78±0.06
	1D02	-	-	8.40±0.30	-	-	13.76±0.08	-	-	-
M. macrosporus	1H01	-	-	-	-	-	-	18.16±0.24	-	9.65±0.03
	4H02	-	-	-	-	-	-	-	-	10.39±0.51
	1C03	-	-	-	-	14.19±0.05	-	-	-	-
M. stipitatus	7C01	-	-	-	14.77±0.15	-	9.89±0.62	-	-	-
	1C06	9.76±0.47	9.14±0.03	-	13.70±0.02	12.24±0.04	10.22±0.05	9.75±0.08	8.01±0.04	-
	3D02	14.33±0.14	13.87±0.12	11.90±0.10	14.57±0.60	19.74±0.29	16.61±0.04	13.74±0.08	8.43±0.08	-
	1H03	11.22±0.05	12.15±0.18	11.86±0.09	10.75±0.02	17.36±0.07	8.65±0.06	12.94±0.09	-	-
	1C05	12.90±0.36	-	-	-	-	15.99±0.27	12.38±0.22	-	-
M. virescens	8C02	-	-	-	-	-	14.00±0.26	-	-	-
	1Y02	13.90±0.36	8.72±0.08	10.50±0.30	13.10±0.44	11.31±0.27	15.66±0.47	-	-	-
	2H01	10.03±0.09	7.54±0.13	10.15±0.13	11.59±0.03	12.25±0.03	10.09±0.12	-	-	-
	1C07	-	-	-	-	15.59±0.03	-	-	-	-
氨苄青霉素钠	-	19.46±0.30	10.40±0.15	14.33±0.38	20.17±0.23	22.10±0.19	17.11±0.56	17.02±0.03	15.72±0.02	18.39±0.12
空白对照	-	-	-	-	-	-	-	-	-	-

广西北部湾可培养黏细菌资源初探及其抗菌活性研究

Preliminary study on the culturable myxobacteria resources from the Beibu Gulf, Guangxi and their antibacterial activity

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