黄海西北近岸沉积物中细菌群落空间分布特征

采用16S rDNA文库和变性梯度凝胶电泳(DGGE)技术,对黄海西北部3个近岸站位沉积物中细菌群落多样性及空间分布特征进行了调查和解析。对表层沉积物16S rDNA序列统计表明,各站位细菌群落多样性很高,γ-和δ-变形菌纲分别占克隆序列总数的20%~32%,是沉积物中的绝对优势类群。DGGE图谱分析表明,同一站位中不同深度的细菌群落结构相似性较高,而不同站位间群落结构相差较远。研究表明在黄海西北近岸沉积物中细菌群落多样性较高,优势类群明显,在较小尺度范围内群落结构的垂直变化不明显。     

Microbial diversity in polluted harbor sediments I: Bacterial community assessment based on four clone libraries of 16S rDNA

Bacteria, as the most abundant sediment organism, play a major role in the fate of pollutants. Therefore, many pollutant-related bacteria have been studied in harbor sediments, yet the entire bacterial profiles have not been reported. The bacterial diversity and community structures from sediments in Victoria Harbor (Hong Kong), including two polluted (VH and VHW) and two adjacent (open oceanic, TLC; estuary discharge affected, PC) sites, were characterized by analyses of four 16S rDNA clone libraries. Upon comparisons of RFLP patterns from 254 clones in the libraries, 178 unique phylotypes were retrieved. LIBSHUFF and Rarefaction analyses indicated that the sediment bacterial communities at the four sites showed high 16S rDNA richness and were significantly different from each other. Phylogenetic analysis of full-length 16S rDNA revealed 19 bacterial phyla in Victoria Harbor sediments. γ- and δ-proteobacteria, holophaga/acidobacteria, and planctomycetales were recorded in all the libraries. In addition, γ- and δ-proteobacteria were dominant at all sites (33.33–11.67%). Besides these two phyla, ε-proteobacteria, firmicutes, aminobacterium, holophaga/acidobacteria and bacteroidetes were judged to be major components of a given library since they constituted 10% or more of the total OTUs of the given library. The cyanobacteria, verrucomicrobia, β-proteobacteria, aminobacterium, chlorofiexi, and candidate division OP1, OP8 were detected in minor proportions in various libraries. A portion of the clones were only distantly related to sequences in the GenBank, suggesting bacteria in Victoria Harbor sediments were unique and diversified.     

Distribution of ammonia-oxidizing Betaproteobacteria community in surface sediment off the Changjiang River Estuary in summer

The spatial distribution of ammonia-oxidizing Betaproteobacteria ( β AOB) was investigated by FISH (fluorescence in situ hybridization) and DGGE (denaturing gradient gel electrophoresis) techniques in the sediment off the Changjiang River Estuary.Sediment samples were collected from eight stations in June before the formation of hypoxia zone in 2006.The abundance of β AOB ranged from 1.87 × 10 5 to 3.53 × 10 5 cells/g of sediment.β AOB abundance did not present a negative correlation with salinity,whereas salinity was implicated as the primary factor affecting nitrification rates.The DGGE profiles of PCR-amplified amo A gene fragments revealed that the β AOB community structure of sample S2 separated from other samples at the level of 40% similarity.The variations in composition of β AOB were significantly correlated with the salinity,temperature,absorption ability of sediments and TOC. The statistical analysis indicates that the β AOB abundance was a main factor to influence nitrification rates with an influence ratio of 87.7% at the level of 40% biodiversity similarity.Considering the good correlation between β AOB abundance and nitrification estimates,the abundance and diversity of β AOB community could be expected as an indirect index of nitrification activity at the study sea area in summer.     

条斑紫菜外生细菌的遗传多样性

为研究藻际微生物多样性与紫菜健康养殖的关系,从江苏南通(养殖)、山东青岛(野生)、浙江温州(养殖)三地的条斑紫菜叶状体和江苏东台条斑紫菜贝壳丝状体及其周围的海水样品中分离了条斑紫菜外生细菌。根据菌落特征和菌体形态将获得的63株细菌分为21种表型。16SrDNA-V3片段DGGE分析结合序列测序比对结果表明:这些菌株与假交替单孢菌(Pseudoalteromonas)、嗜冷单胞菌(Psychrobacter)和芽孢杆菌(Bacillus)等10属相应菌株具有较高的同源性。通过对条斑紫菜外生细菌区系多样性分析得出:条斑紫菜藻体外生细菌组成与生活环境相关,但不同于周围海水细菌群落;不同来源(地点)、不同生理状况(健康与病烂)和不同生长阶段(叶状体和丝状体)的条斑紫菜外生菌种群组成差异较大。许多外生细菌对常见测试菌、海水养殖动物致病菌和人致病菌具有抑菌活性,其抑菌特点与紫菜生理性状相关。从健康条斑紫菜样品分离的外生菌中假交替单孢菌占优势,病烂紫菜未分离到假交替单孢菌,提示假交替单孢菌可能与紫菜健康生长有密切关系。     

Phylogenetic analysis of bacterial community in deep-sea sediment from the western Pacific “warm pool”

A depth profile of bacterial community structure in one deep-sea sediment core of the western Pacific ＂warm pool＂ （WP） was investigated and compared with that in a sediment sample from the eastern Pacific （EP） by phylogenetic analysis of 16S rDNA fragments. Five bacterial 16S rDNA clone libraries were constructed, and 133 clones with different restriction fragment length polymorphism （RFLP） patterns were sequenced. A phylogenetic analysis of these sequences revealed that the bacterial diversity in a sample from the WP was more abundant than that in the EP sample. The bacterial population in the sediment core of WP was composed of eight major lineages of the domain bacteria. Among them the γ-Proteobacteria was the predominant and most diverse group in each section of WP sediment core, followed by the α-Proteobacteria. The genus Colwellia belonging to γ-Proteobacteria was predominant in this sample. The shift of bacterial communities among different sections of the WP sediment core was δ-, ε-Proteobacteria, and Cytopahga-Flexibacteria-Bacteroides （CFB） group. The ratios between them in the bacterial communities all showed inversely proportional to the depth of sediment. The sequences related to sulphate reducing bacteria （SRB） were detected in every section. The bacterial community structure in this sediment core might be related to the environmental characteristics of the surface seawater of the western Pacific WP.     

Nematode species distribution patterns at the Håkon Mosby Mud Volcano (Norwegian Sea)

The Håkon Mosby Mud Volcano (HMMV) is a methane seep that is densely populated by the bacteria Beggiatoa spp. as well as by tubeworms of the Family Siboglinidae. Analyses of samples from seven stations located in five different habitats (bare sediments, bacterial mats, siboglinid fields, sediments with small patches of siboglinids and areas of ‘normal’ sea floor) revealed differences in the density and species‐level diversity of nematode communities. Total densities increased from stations inside the mud volcano and on its rim towards the ‘normal’ areas outside the volcano. Nematode species diversity was similar in samples from the siboglinid fields and the bare HMMV sediments. However, the sediment with patches of siboglinids collected outside the volcano was characterised by a diverse nematode community comprising 47 species, whereas the control sediments without siboglinids yielded only 27 species. The nematode assemblage at the bacterial mat site included only two identified species, Halomonhystera disjuncta and Sabatieria ornata, with H. disjuncta being strongly dominant. Terschellingia distlamphida, S. ornata and Aponema ninae dominated nematode communities at the siboglinid fields and in bare HMMV sediments. Species dominating at stations inside the caldera were found in comparable low densities in the sediment with siboglinid patches collected outside the volcano, but were missing in the background samples, where species of Endeolophos, Acantholaimus and Desmoscolex were dominant. Species diversity generally decreased with sediment depth. A subsurface‐dwelling nematode community was observed in the siboglinid fields and the bare sediments. Background sediments showed a sharp decline with 18–20 species at 0–2 cm versus three species at 3–5 cm sediment depth. The trophic composition of the nematode fauna at the HMMV showed a prevalence of deposit feeders at almost all stations, whereas background sediments exhibited a uniform distribution of non‐selective and selective deposit feeders and epistrate feeders. The high percentage of deposit feeders inside the mud volcano could be related to the grain‐size distribution and the significantly higher bacterial biomass compared to the control sediments.     

Phylogenetic diversity of bacteria associated with the marine sponge Agelas robusta from South China Sea

It is well known that marine sponges harbor large numbers of bacteria, some of which have been proved to be sponge-specific. The diversity of bacteria in marine sponges distributed along the coast of South China Sea has been previously studied but that of bacteria in sponges inhabiting the open sea has been rarely investigated. In this report, the diversity of bacteria associated with the marine sponge Agelas robusta from a remote coral reef in the South China Sea was documented employing 16S rDNA library construction, amplified ribosomal DNA restriction analysis (ARDRA) and phylogenetic analysis. A total of 32 phylotypes were finally categorized in nine phyla including Cyanobacteria, Proteobacteria, Chloroflexi, Firmicutes, Actinobacteria, Acidobacteria, Planctomycetes, Bacteroidetes and Gemmatimonadetes. The dominant phylum was Proteobacteria whereas the dominant genus was Synechococcus in Cyanobacteria. Some spongespecific bacteria were also found in the bacteria population, but the proportion (5 OTUs/32 OTUs) was much lower than other sponges. This study reveals the phylogenetic diversity of bacteria in A. robusta and confirms the presence of some sponge-specific bacteria in the South China Sea. Understanding the diversity of sponge-associated bacteria in China assists to exploit the bacteria resources for biotechnology.     

渤海沉积物中细菌群落结构和基因丰度的空间分布特征

This study investigated differences in the community structure and environmental responses of the bacterial community in sediments of the Bohai Sea.Illumina high-throughput sequencing technology and real-time PCR were used to assay the bacterial 16S rRNA genes in the surface sediments of 13 sampling stations in the Bohai Sea.The results showed that sediments at the majority of the 13 sampling stations were contaminated by heavy metal mercury.The main phyla of bacteria recorded included Proteobacteria(52.92%),Bacteroidetes(11.76%),Planctomycetes(7.39%),Acidobacteria(6.53%)and Chloroflexi(4.97%).The genus with the highest relative abundance was Desulfobulbus(4.99%),which was the dominant genus at most sampling stations,followed by Lutimonas and Halioglobus.The main factors influencing bacterial community structure were total organic carbon,followed by depth and total phosphorus.The content of lead,cadmium,chromium,copper and zinc had a consistent effect on community structure.Arsenic showed a negative correlation with bacterial community structure in most samples,while the impact of mercury on community structure was not significant.The bacterial community in sediment samples from the Bohai Sea was rich in diversity and displayed an increase in diversity from high to low latitudes.The data indicated that the Bohai Sea had abundant microbial resources and was rich in bacteria with the potential to metabolize many types of pollutants.     

Molecular genetic diversity of bacteria in the bottom section of arctic sea ice from the Canada Basin

PCR-DGGE approach was used to analyze bacterial diversity in the bottom section of seven arctic sea ice samples colleted from the Canada Basin. Thirty-two 16S rDNA sequences were obtained from prominent DGGE bands. The closest relatives of these sequences are found to be those of cultivated or uncultured bacteria from antarctic or arctic sea ice. Phylogenetic analysis clustered these sequences or phylotypes within α- proteobacteria, γ-proteobacteria and CFB (cytophaga-flexibacter-bacteroides) group. Sequences belonging to γ-proteobacteria were dominant and members of the CFB group were highly abundant. It was suggested that the CFB group was the representative of the bottom section of sea ice samples.     

海州湾赤潮高发区浮游细菌与赤潮及环境因子的关系

海州湾是南黄海赤潮高发区之一。为研究浮游细菌与赤潮及环境因子的关系,于2020年11月在海州湾赤潮期间进行了现场调查,采集了8个站位的浮游细菌、浮游植物样品,获取了温度、盐度、叶绿素a以及氮、磷、硅营养盐浓度等环境因子信息,分析了赤潮原因种及浮游细菌群落结构情况。结果显示,此次赤潮原因种为三叶原甲藻(Prorocentrum triestinum)、赤潮异弯藻(Heterosigma akashiwo)及塔胞藻(Pyramimonas sp.),发生赤潮时水温、盐度较低,盐度与赤潮异弯藻的分布有一定的联系。使用16SrRNA高通量测序方法分析调查海域的浮游细菌情况,结果显示赤潮发生站位的浮游细菌多样性略低于其他非赤潮发生站位,优势细菌与其他站位有明显不同,以假单胞菌目(Pseudomonadales)、红杆菌目(Rhodobacterales)和黄杆菌目(Flavobacteriales)为主。赤潮发生站位的浮游细菌群落结构也存在一定的差异,不同站位可能处于不同的赤潮阶段,对浮游细菌群落具有不同的影响。相关性分析显示,黄杆菌目与叶绿素a显著正相关,表明黄杆菌目与赤潮发生有一定的关系。     

Deep-sea bacterial communities in sediments and guts of deposit-feeding holothurians in Portuguese canyons (NE Atlantic)

Deposit-feeding holothurians often dominate the megafauna in bathyal deep-sea settings, in terms of both abundance and biomass. Molpadia musculus is particularly abundant at about 3400 m depth in the Nazaré Canyon on the NE Atlantic Continental Margin. However, these high abundances are unusual for burrowing species at this depth. The objective of this research was to understand the reasons of the massive occurrence of these molpadiid holothurians in the Nazaré Canyon. To address this question we investigated possible trophic interactions with bacteria at sites where the organic content of the sediment was different (Setúbal and Cascais Canyons, NE Atlantic Continental Margin). The molecular fingerprinting technique of Denaturing Gradient Gel Electrophoresis (DGGE) with band sequencing, combined with non-metric multi-dimensional scaling and statistical analyses, was used to compare the bacterial community diversity in canyon sediments and holothurian gut contents. Our results suggest that M. musculus does not need to develop a specialised gut bacterial community to aid digestion where the sediment is rich in organic matter (Nazaré Canyon); in contrast, such a community may be developed where the sediment is poorer in organic matter (Cascais Canyon).     

施肥对盐沼沉积物微生物生物量和细菌群落组成的影响

The effects of nitrogen(N) addition on microbial biomass, bacterial abundance, and community composition in sediment colonized by Suaeda heteroptera were examined by chloroform fumigation extraction method, real-time quantitative polymerase chain reaction, and denaturing gradient gel electrophoresis(DGGE) in a salt marsh located in Shuangtai Estuary, China. The sediment samples were collected from plots treated with different amounts of a single N fertilizer(urea supplied at 0.1, 0.2, 0.4 and 0.8 g/kg(nitrogen content in sediment) and different forms of N fertilizers(urea,(NH_4)_2SO_4, and NH_4NO_3, each supplied at 0.2 g/kg(calculated by nitrogen).The fertilizers were applied 1–4 times during the plant-growing season in May, July, August, and September of 2013. Untreated plots were included as a control. The results showed that both the amount and form of N positively influenced microbial biomass carbon, microbial biomass nitrogen, and bacterial abundance. The DGGE profiles revealed that the bacterial community composition was also affected by the amount and form of N.Thus, our findings indicate that short-term N amendment increases microbial biomass and bacterial abundance,and alters the structure of bacterial community.     

南极菲尔德斯半岛土壤微生物在富集传代培养中对菲的响应

为揭示富集传代培养过程中南极多环芳烃降解菌群的演替规律,以菲为唯一碳源和能源对南极菲尔德斯半岛不同地理位置的6份土壤样品进行了富集和连续传代培养,并采用高通量测序技术分析了富集传代培养过程中菲降解菌群的群落结构及生物多样性.Alpha多样性分析结果表明,在富集培养阶段南极菲降解菌的物种丰富度显著高于传代培养阶段(P＜0...     

Microbial community structure and diversity in deep-sea hydrothermal vent sediments along the Eastern Lau Spreading Centre

The aim of this study is to investigate microbial structures and diversities in five active hydrothermal fields’ sediments along the Eastern Lau Spreading Centre (ELSC) in the Lau Basin (southwest Pacific). Microbial communities were surveyed by denatured gradient gel electrophoresis (DGGE) and clone library analysis of 16S rRNA genes. The differences in microbial community structures among sediment samples from the five deep-sea hydrothermal sites were revealed by DGGE profiles. Cluster analysis of DGGE profiles separated the five hydrothermal samples into two groups. Four different 16S rRNA gene clone libraries, representing two selected hydrothermal samples (19-4TVG8 and 19-4TVG11), were constructed. Twenty-three and 32 phylotypes were identified from 166 and 160 bacterial clones respectively, including Proteobacteria, Bacteroidetes, Firmicutes, Nitrospirae and Planctomycetes. The phylum Proteobacteria is dominant in both bacterial libraries with a predominance of Gamma-Proteobacteria. A total of 31 and 25 phylotypes were obtained from 160 and 130 archaeal clones respectively, including Miscellaneous Crenarchaeotic Group, Marine Group Ⅰ and Ⅲ, Marine Benthic Group E, Terrestrial Hot Spring Crenarchaeota and Deep-sea Hydrothermal Vent Euryarchaeota. These results show a variety of clones related to those involved in sulfur cycling, suggesting that the cycling and utilization of sulfur compounds may extensively occur in the Lau Basin deep-sea hydrothermal ecosystem.     

抗生素对浒苔形态及共附生细菌多样性的影响

本文研究了3种抗生素联合处理黄海绿潮浒苔(Ulva prolifera)形态及共附生细菌多样性的影响。将浒苔放散的配子加入到含有3种广谱抗生素(青霉素G、硫酸新霉素、多黏菌素B)的培养液中进行培养, 与对照组相比, 发现浒苔幼苗生长速度缓慢, 且细胞变小, 藻体无法形成正常形态。进一步采用16S rDNA高通量测序分析抗生素处理藻体前后的共附生细菌多样性, 发现抗生素处理后浒苔共附生细菌丰富度和多样性降低, 其中Maribacter丰度降低。将藻体与形态发生化合物thallusin(Maribacter spp. 提取物)共培养, 发现thallusin可促进藻体形态恢复和生长, 推测抗生素可通过抑制Maribacter等细菌从而影响浒苔形态和生长, 这将为浒苔共附生细菌功能的深入研究奠定一定基础。     

东太平洋深海沉积物中DNA的提取及细菌多样性初步分析

以东太平洋海隆附近深海柱状沉积物为材料,通过化学裂解和酶消化相结合的方法提取了沉积物微生物的总基因组DNA,并进行了纯化。结果表明所得到的DNA分子片段大小在21kb左右,纯化后的DNA可直接用于PCR等分子生物学操作。细菌16SrDNAV3可变区的PCR—DGGE图谱展示出15条以上条带,表明深海沉积物中细菌多样性较高,群落结构比较复杂。对其中9条主要条带进行回收、测序和系统发育分析,结果表明所获得的序列分属放线菌门（Actinobacteria）,绿弯菌门（Chloroflexi）,γ-变形细菌亚门（Gamma—proteobacteria）,α-变形细菌亚门（Alpha—proteobacteria）和嗜酸菌门（Acidobacteria）5个大类群。     

东海陆架表层沉积物细菌群落结构及地理分布研究

本研究通过构建16S r DNA克隆文库,对东海陆架五个站点的沉积物样品进行群落结构及地理分布研究。结果表明:东海陆架沉积物细菌分属于13个类群,包括变形菌门(Proteobacteria)、厚壁菌门(Firmicutes)、酸杆菌门(Acidobacteria)、绿弯菌门(Chloroflexi)、放线菌门(Actinobacteria)、硝化螺旋菌门(Nitrospira)、WS3、拟杆菌门(Bacteroidetes)、螺旋体门(Spirochaetes)、OP8、浮霉菌门(Planctomycetacia)、疣微菌门(Verrucomicrobiae)和芽单胞菌门(Gemmatimonadetes),其中变形菌门(γ-变形菌纲、δ-变形菌纲)和绿弯菌门为优势菌群,Latescibacteria(WS3)、疣微菌门和芽单胞菌门分别是站点DH6、DH21和DH9的特有类群。五个站点细菌多样性从高到低排序依次为DH9DH21DH17DH6DH16,其群落结构和丰度与各站点不同沉积环境类型有关。     

冬季刺参养殖环境与肠道内细菌菌群的研究

运用传统细菌分离培养与分子生物学技术相结合的方法,对2008年11月至2009年1月冬季刺参(Apostichopus japonicus Liao)养殖池塘环境(养殖水、底泥、附着基)及刺参肠道内的细菌菌群进行了分析。应用平板稀释涂布培养计数法测得刺参养殖池塘水体、底泥、附着基和肠道细菌数量分别为0.75×102～1.4×104cfu/mL、8.7×104～8.1×105cfu/g、3.8×105～2.8×106cfu/g、7.1×105～1.5×107cfu/g;根据形态学差异从培养所得的细菌中筛选得到22株菌,用限制性内切酶Rsa I和Msp I对所分离菌株进行ARDRA(Amplified rDNA Restriction Analysis)分析,这22株菌被分为8种不同的分类单元(Operational Taxonomic Unit,OTU),其中OTU2与OTU3所包含的菌株分别占分离菌株种数的30%和20%;此外,作者对不同环境培养所得的优势度最高的细菌进行分子鉴定分析。结果表明:水环境中优势菌为施氏假单胞菌(Pseudomonas stutzeri)、门多萨假单胞菌(Pseudomonas mendocina)及巨大芽孢杆菌(Bacillus megaterium),沉积物中优势菌为巨大芽孢杆菌(B.megaterium)、施氏假单胞菌(P.stutzeri)、苏云金芽孢杆菌(Bacillus thuringiensis),附着基中优势菌为巨大芽孢杆菌(B.megaterium)、苏云金芽孢杆菌(B.thuringiensis),灿烂弧菌(Vibrio splendidus),肠道中优势菌为巨大芽孢杆菌(B.megaterium)、苏云金芽孢杆菌(B.thuringiensis),灿烂弧菌(V.splendidus)、施氏假单胞菌(P.stutzeri)。通过对冬季刺参池塘细菌菌群多样性分析和优势菌鉴定,为筛选低温益生菌和防治刺参疾病提供了有益参考。     

近海沉积物与水体中天然大分子聚合物降解菌的原位富集与多样性分析

本研究选取5种海洋动植物大分子高聚物或其天然组织,包括虾壳、鱼鳞、海带叶片、几丁质和壳聚糖,分别在海水表层和沉积物环境中进行富集,定期取样,通过高通量测序分析菌群多样性。结果发现,不同有机物原位富集的细菌多样性存在较大的差异,而且同种底物在海水表层与沉积物中的降解菌菌群差异较大。从物种多样性看,在海水表层环境中富集的鱼鳞样品种群最丰富,而沉积物环境中富集的海带叶片样品菌群多样性最低,除其优势菌群为热袍菌门外,其他所有富集物中优势菌均为变形菌门。其中脱硫杆菌科、黄杆菌科、脱硫弧菌科和弧菌科占有较大比例,脱硫杆菌科在所有样品中优势较大,黄杆菌科在海水表层环境样品中为优势菌群,弧菌科在沉积物样品几丁质和壳聚糖样品中占比较高。本研究通过对同种海洋环境中不同富集样品之间以及同种底物不同环境中富集菌群之间的比较,分析结果得到了原位条件下参与大分子聚合物降解的菌群种类,但有待于在更多不同的海域富集物中进行验证。     

深海沉积物中微量DNA的提取及应用

采用化学裂解和酶解相结合的方法，进行了深海沉积物中微量DNA的提取，并采用DNA吸附树脂进行纯化。结果表明，该方法能有效地去除沉积物中的腐殖酸等抑制剂，每克湿重沉积物样品可得到DNA约16μg，回收率可达95％，所得到的DNA分子片段均在23kb左右，纯化后的DNA可直接应用于各种分子生物学操作。利用细菌16SrDNA通用引物对所提取的深海沉积物DNA进行了PCR—RRJ及系统发育分析，各主要细菌类群均能检出，证实该方法可以应用于深海等极端环境中微生物的多样性调查、系统发育分析以及特殊功能基因的筛选，同时还可应用于环境样品中生物量的半定量估计。