Longitudinal Changes in the Bacterial Community Composition of the Danube River: a Whole-River Approach

The Danube River is the second longest river in Europe, and its bacterial community composition has never been studied before over its entire length. In this study, bacterial community composition was determined by denaturing gradient gel electrophoresis (DGGE) analysis of PCR-amplified portions of the bacterial 16S rRNA gene from a total of 98 stations on the Danube River (73 stations) and its major tributaries (25 stations), covering a distance of 2,581 km. Shifts in the bacterial community composition were related to changes in environmental conditions found by comparison with physicochemical parameters (e.g., temperature and concentration of nutrients) and the concentration of chlorophyll a (Chl a). In total, 43 distinct DGGE bands were detected. Sequencing of selected bands revealed that the phylotypes were associated with typical freshwater bacteria. Apparent bacterial richness in the Danube varied between 18 and 32 bands and correlated positively with the concentration of P-PO₄ (r = 0.56) and negatively with Chl a (r = −0.52). An artificial neural network-based model explained 90% of the variation of apparent bacterial richness using the concentrations of N-NO₂ and P-PO₄ and the distance to the Black Sea as input parameters. Between the cities of Budapest and Belgrade, apparent bacterial richness was significantly lower than that of other regions of the river, and Chl a showed a pronounced peak. Generally, the bacterial community composition developed gradually; however, an abrupt and clear shift was detected in the section of the phytoplankton bloom. Large impoundments did not have a discernible effect on the bacterial community of the water column. In conclusion, the riverine bacterial community was largely influenced by intrinsic factors.     

Particle-attached and free-living bacterial communities in a contrasting marine environment: Victoria Harbor, Hong Kong

Diversity of particle-attached and free-living marine bacteria in Victoria Harbor, Hong Kong, and its adjacent coastal and estuarial environments was investigated using DNA fingerprinting and clone library analysis. Denaturing gradient gel electrophoresis (DGGE) analysis of 16S rRNA genes showed that bacterial communities in three stations of Victoria Harbor were similar, but differed from those in adjacent coastal and estuarine stations. Particle-attached and free-living bacterial community composition differed in the Victoria Harbor area. DNA sequencing of 28 bands from DGGE gel showed Alphaproteobacteria was the most abundant group, followed by the Bacteroidetes, and other Proteobacteria. Bacterial species richness (number of DGGE bands) differed among stations and populations (particle-attached and free-living; bottom and surface). BIOENV analysis indicated that the concentrations of suspended solids were the major contributing parameter for the spatial variation of total bacterial community structure. Samples from representative stations were selected for clone library (548 clones) construction and their phylogenetic distributions were similar to those of sequences from DGGE. Approximately 80% of clones were affiliated to Proteobacteria, Bacteroidetes and Cyanobacteria. The possible influences of dynamic pollution and hydrological conditions in the Victoria Harbor area on the particle-attached and free-living bacterial community structures were discussed.     

Spatial heterogeneity of bacterial community structure in the sediments of the Pearl River estuary

Denaturing gradient gel electrophoresis (DGGE) and multivariate statistical analytical methods were applied to investigate the spatial variation of bacterial community structure in the Pearl River estuary sediment and to address the relationship between microbial community composition and bottom water chemistry in ten different stations. Preliminary results of sequencing analysis of the excised DGGE bands suggested that α-Proteobacteria, γ-Proteobacteria, Acidobacteria and Actinobacteria were the dominant bacterial groups in the Pearl River estuary sediment. Results of multidimensional scaling analysis of these field data suggested that the composition of bacterial communities varied with sampling sites. Finally, canonical correspondence analysis of the data of environmental variables and bacterial community suggested that bacterial community structure was significantly influenced by the change of environmental variables (total phosphorus, nitrite, ammonium, dissolved oxygen, pH and salinity).     

Complexity of Bacterial Communities in a River-Floodplain System (Danube, Austria)

Natural floodplains play an essential role in the processing and decomposition of organic matter and in the self-purification ability of rivers, largely due to the activity of bacteria. Knowledge about the composition of bacterial communities and its impact on organic-matter cycling is crucial for the understanding of ecological processes in river-floodplain systems. Particle-associated and free-living bacterial assemblages from the Danube River and various floodplain pools with different hydrological characteristics were investigated using terminal restriction fragment length polymorphism analysis. The particle-associated bacterial community exhibited a higher number of operational taxonomic units (OTUs) and was more heterogeneous in time and space than the free-living community. The temporal dynamics of the community structure were generally higher in isolated floodplain pools. The community structures of the river and the various floodplain pools, as well as those of the particle-associated and free-living bacteria, differed significantly. The compositional dynamics of the planktonic bacterial communities were related to changes in the algal biomass, temperature, and concentrations of organic and inorganic nutrients. The OTU richness of the free-living community was correlated with the concentration and origin of organic matter and the concentration of inorganic nutrients, while no correlation with the OTU richness of the particle-associated assemblage was found. Our results demonstrate the importance of the river-floodplain interactions and the influence of damming and regulation on the bacterial-community composition.     

Complexity of bacterial communities in a river-floodplain system (Danube, Austria)

Natural floodplains play an essential role in the processing and decomposition of organic matter and in the self-purification ability of rivers, largely due to the activity of bacteria. Knowledge about the composition of bacterial communities and its impact on organic-matter cycling is crucial for the understanding of ecological processes in river-floodplain systems. Particle-associated and free-living bacterial assemblages from the Danube River and various floodplain pools with different hydrological characteristics were investigated using terminal restriction fragment length polymorphism analysis. The particle-associated bacterial community exhibited a higher number of operational taxonomic units (OTUs) and was more heterogeneous in time and space than the free-living community. The temporal dynamics of the community structure were generally higher in isolated floodplain pools. The community structures of the river and the various floodplain pools, as well as those of the particle-associated and free-living bacteria, differed significantly. The compositional dynamics of the planktonic bacterial communities were related to changes in the algal biomass, temperature, and concentrations of organic and inorganic nutrients. The OTU richness of the free-living community was correlated with the concentration and origin of organic matter and the concentration of inorganic nutrients, while no correlation with the OTU richness of the particle-associated assemblage was found. Our results demonstrate the importance of the river-floodplain interactions and the influence of damming and regulation on the bacterial-community composition.     

Response of bacterioplankton community structures to hydrological conditions and anthropogenic pollution in contrasting subtropical environments

Bacterioplankton community structures under contrasting subtropical marine environments (Hong Kong waters) were analyzed using 16S rRNA gene denaturing gradient gel electrophoresis (DGGE) and subsequent sequencing of predominant bands for samples collected bimonthly from 2004 to 2006 at five stations. Generally, bacterial abundance was significantly higher in the summer than in the winter. The general seasonal variations of the bacterial community structure, as indicated by cluster analysis of the DGGE pattern, were best correlated with temperature at most stations, except for the station close to a sewage discharge outfall, which was best explained by pollution-indicating parameters (e.g. biochemical oxygen demand). Anthropogenic pollutions appear to have affected the presence and the intensity of DGGE bands at the stations receiving discharge of primarily treated sewage. The relative abundance of major bacterial species, calculated by the relative intensity of DGGE bands after PCR amplification, also indicated the effects of hydrological or seasonal variations and sewage discharges. For the first time, a systematic molecular fingerprinting analysis of the bacterioplankton community composition was carried out along the environmental and pollution gradient in a subtropical marine environment, and it suggests that hydrological conditions and anthropogenic pollutions altered the total bacterial community as well as the dominant bacterial groups.     

Development of the bacterial compartment along the Danube River: a continuum despite local influences

Microbial food webs dominate heterotrophic food webs in large rivers with bacterial metabolism being a key component of carbon processing. Thus, analysis of bacterial population dynamics is critical to understanding patterns and mechanisms of material cycling and energy fluxes in large rivers. Within the frame of the Joint Danube Survey (JDS) 2007, the longitudinal development of the natural bacterial community in the Danube in terms of bacterial numbers, morphotype composition, and heterotrophic production of the suspended and particle-attached fractions was followed at a fine spatial resolution of approximately 30 km for the first time in such a large river along a 2,600-km stretch. Twenty-one major tributaries and branches were also included. This allowed us to investigate whether bacterial standing stock and production undergo continuous, linear changes or whether discontinuities and local processes like the merging of tributaries or the potential impact of sewage input drive the bacterial population in the Danube. The presented investigation revealed surprising continuous patterns of changes of bacterial parameters along the Danube River. Despite the presence of impoundments or hydropower plants, large municipalities, and the discharge of large tributaries, most bacterial parameters (standing stock, morphotype succession, and attached bacterial production) developed gradually, indicating that mainly broad-scale drivers and not local conditions shape and control the bacterial community in the midstream of this large river. As most important broad-scale drivers, nutrients (inorganic and organic) and changes in particle concentrations were identified. These data are also in remarkable accordance with the patterns of changes of the genetic bacterial community composition, observed during the first JDS (2001) 6 years before. In contrast, bacterial activity did not follow a continuous trend and was mainly controlled by the input of sewage from large cities in the middle section, leading to a bloom of phytoplankton. The observed patterns and the comparison between the Danube, its tributaries and other large rivers worldwide indicate that the bacterial community in rivers has a powerful indicator function for estimating the ecological status of large river ecosystems once enough information has been collected at various temporal and spatial scales.     

Succession of bacterial community structure along the Changjiang River determined by denaturing gradient gel electrophoresis and clone library analysis

Bacterial community structure along the Changjiang River (which is more than 2,500 km long) was studied by using denaturing gradient gel electrophoresis (DGGE) and clone library analysis of PCR-amplified 16S ribosomal DNA (rDNA) with universal bacterial primer sets. DGGE profiles and principal-component analysis (PCA) demonstrated that the bacterial community gradually changed from upstream to downstream in both 1998 and 1999. Bacterial diversity, as determined by the Shannon index (H'), gradually decreased from upstream to downstream. The PCA plots revealed that the differences in the bacterial communities among riverine stations were not appreciable compared with the differences in two adjacent lakes, Lake Dongting and Lake Poyang. The relative stability of the bacterial communities at the riverine stations was probably due to the buffering action of the large amount of water flowing down the river. Clone library analysis of 16S rDNA revealed that the dominant bacterial groups changed from beta-proteobacteria and the Cytophaga-Flexibacter-Bacteroides group upstream to high-G+C-content gram-positive bacteria downstream and also that the bacterial community structure differed among the stations in the river and the lakes. The results obtained in this study should provide a reference for future changes caused by construction of the Three Gorges Dam.     

Phylogenetic diversity, composition and distribution of bacterioplankton community in the Dongjiang River, China

Bacterioplankton community compositions in the Dongjiang River were characterized using denaturing gradient gel electrophoresis (DGGE) and 16S rRNA gene clone library construction. Water samples in nine different sites were taken along the mainstem and three tributaries. In total, 24 bands from DGGE gels and 406 clones from the libraries were selected and sequenced, subsequently analyzed for the bacterial diversity and composition of those microbial communities. Bacterial 16S rRNA gene sequences from freshwater bacteria exhibited board phylogenetic diversity, including sequences representing the Alphaproteobacteria, Betaproteobacteria, Gammaproteobacteria, Actinobacteria, Bacteriodetes, Verrucomicrobia, and candidate division TM7. Members of Betaproteobacteria group were the most dominant in all sampling sites, followed by Gammaproteobacteria, Alphaproteobacteria, and Actinobacteria. DGGE profiles and the ∫-LIBSHUFF analysis revealed similar patterns of bacterial diversity among most sampling sites, while spatial distribution variances existed in all sites along the river basin. Statistical analysis showed that bacterial species distribution strongly correlated with environmental variables, such as nitrate and ammonia, suggesting that nitrogen nutrients may shape the microbial community structure and composition in the Dongjiang River. This study had important implications for the comparison with other rivers elsewhere and contributed to the growing data set on the factors that structure bacterial communities in freshwater ecosystems.     

Microbial biogeography along an estuarine salinity gradient: combined influences of bacterial growth and residence time

Shifts in bacterioplankton community composition along the salinity gradient of the Parker River estuary and Plum Island Sound, in northeastern Massachusetts, were related to residence time and bacterial community doubling time in spring, summer, and fall seasons. Bacterial community composition was characterized with denaturing gradient gel electrophoresis (DGGE) of PCR-amplified 16S ribosomal DNA. Average community doubling time was calculated from bacterial production ([(14)C]leucine incorporation) and bacterial abundance (direct counts). Freshwater and marine populations advected into the estuary represented a large fraction of the bacterioplankton community in all seasons. However, a unique estuarine community formed at intermediate salinities in summer and fall, when average doubling time was much shorter than water residence time, but not in spring, when doubling time was similar to residence time. Sequencing of DNA in DGGE bands demonstrated that most bands represented single phylotypes and that matching bands from different samples represented identical phylotypes. Most river and coastal ocean bacterioplankton were members of common freshwater and marine phylogenetic clusters within the phyla Proteobacteria, Bacteroidetes, and ACTINOBACTERIA: Estuarine bacterioplankton also belonged to these phyla but were related to clones and isolates from several different environments, including marine water columns, freshwater sediments, and soil.     

Diversity and abundance of freshwater Actinobacteria along environmental gradients in the brackish northern Baltic Sea

Actinobacteria are highly abundant in pelagic freshwater habitats and also occur in estuarine environments such as the Baltic Sea. Because of gradients in salinity and other environmental variables estuaries offer natural systems for examining factors that determine Actinobacteria distribution. We studied abundance and community structure of Bacteria and Actinobacteria along two transects in the northern Baltic Sea. Quantitative (CARD-FISH) and qualitative (DGGE and clone libraries) analyses of community composition were compared with environmental parameters. Actinobacteria accounted for 22–27% of all bacteria and the abundance changed with temperature. Analysis of 549 actinobacterial 16S rRNA sequences from four clone libraries revealed a dominance of the freshwater clusters ac I and ac IV, and two new subclusters ( ac I-B scB-5 and ac IV-E) were assigned. Whereas ac I was present at all stations, occurrence of ac II and ac IV differed between stations and was related to dissolved organic carbon (DOC) and chlorophyll a (Chl a ) respectively. The prevalence of the ac I-A and ac I-B subclusters changed in relation to total phosphorus (Tot-P) and Chl a respectively. Community structure of Bacteria and Actinobacteria differed between the river station and all other stations, responding to differences in DOC, Chl a and bacterial production. In contrast, the composition of active Actinobacteria (analysis based on reversely transcribed RNA) changed in relation to salinity and Tot-P. Our study suggests an important ecological role of Actinobacteria in the brackish northern Baltic Sea. It highlights the need to address dynamics at the cluster or subcluster phylogenetic levels to gain insights into the factors regulating distribution and composition of Actinobacteria in aquatic environments.     

Microbial Biogeography along an Estuarine Salinity Gradient: Combined Influences of Bacterial Growth and Residence Time

Shifts in bacterioplankton community composition along the salinity gradient of the Parker River estuary and Plum Island Sound, in northeastern Massachusetts, were related to residence time and bacterial community doubling time in spring, summer, and fall seasons. Bacterial community composition was characterized with denaturing gradient gel electrophoresis (DGGE) of PCR-amplified 16S ribosomal DNA. Average community doubling time was calculated from bacterial production ([¹⁴C]leucine incorporation) and bacterial abundance (direct counts). Freshwater and marine populations advected into the estuary represented a large fraction of the bacterioplankton community in all seasons. However, a unique estuarine community formed at intermediate salinities in summer and fall, when average doubling time was much shorter than water residence time, but not in spring, when doubling time was similar to residence time. Sequencing of DNA in DGGE bands demonstrated that most bands represented single phylotypes and that matching bands from different samples represented identical phylotypes. Most river and coastal ocean bacterioplankton were members of common freshwater and marine phylogenetic clusters within the phyla Proteobacteria, Bacteroidetes, and Actinobacteria. Estuarine bacterioplankton also belonged to these phyla but were related to clones and isolates from several different environments, including marine water columns, freshwater sediments, and soil.     

Succession of Bacterial Community Structure along the Changjiang River Determined by Denaturing Gradient Gel Electrophoresis and Clone Library Analysis

Bacterial community structure along the Changjiang River (which is more than 2,500 km long) was studied by using denaturing gradient gel electrophoresis (DGGE) and clone library analysis of PCR-amplified 16S ribosomal DNA (rDNA) with universal bacterial primer sets. DGGE profiles and principal-component analysis (PCA) demonstrated that the bacterial community gradually changed from upstream to downstream in both 1998 and 1999. Bacterial diversity, as determined by the Shannon index (H′), gradually decreased from upstream to downstream. The PCA plots revealed that the differences in the bacterial communities among riverine stations were not appreciable compared with the differences in two adjacent lakes, Lake Dongting and Lake Poyang. The relative stability of the bacterial communities at the riverine stations was probably due to the buffering action of the large amount of water flowing down the river. Clone library analysis of 16S rDNA revealed that the dominant bacterial groups changed from β-proteobacteria and the Cytophaga-Flexibacter-Bacteroides group upstream to high-G+C-content gram-positive bacteria downstream and also that the bacterial community structure differed among the stations in the river and the lakes. The results obtained in this study should provide a reference for future changes caused by construction of the Three Gorges Dam.     

珠江口水体浮游细菌种群多样性空间分布特征

为认识珠江口水体浮游细菌的多样性分布规律,运用聚合酶链式反应-变性梯度凝胶电泳(PCR-DGGE)和多维尺度分析(MDS)的方法,研究了春季珠江口十个站位表底水层浮游细菌种群16SrRNA基因多样性特征。研究结果表明,珠江口浮游细菌种群具有丰富的基因多样性;不同站位细菌群落结构和优势种群变化显著:大多数站位表底层细菌群落结构比较相似,河口外站位(A12,A14和C5),表底层细菌群落结构差异性较大:多样性分析表明A14,B6和C5站位底层细菌多样性大于表层。遗传发育分析表明,序列归属于变形细菌(Proteobacteria),酸杆菌(Acidobacteria),蓝细菌(Cyanobacteria),厚壁细菌(Firmicutes)和放线菌(Actinobacteria)。变形细菌(Proteobacteria)种类最多,从河口内淡水区到河口外海水区都有大量分布,是珠江口水域占优势的菌群;同时也检测到种类丰富的放线菌(Actinobacteria)的存在,也是珠江口水域的优势菌群。     

Effects of site and plant species on rhizosphere community structure as revealed by molecular analysis of microbial guilds

The bacterial and fungal rhizosphere communities of strawberry (Fragaria ananassa Duch.) and oilseed rape (Brassica napus L.) were analysed using molecular fingerprints. We aimed to determine to what extent the structure of different microbial groups in the rhizosphere is influenced by plant species and sampling site. Total community DNA was extracted from bulk and rhizosphere soil taken from three sites in Germany in two consecutive years. Bacterial, fungal and group-specific (Alphaproteobacteria, Betaproteobacteria and Actinobacteria) primers were used to PCR-amplify 16S rRNA and 18S rRNA gene fragments from community DNA prior to denaturing gradient gel electrophoresis (DGGE) analysis. Bacterial fingerprints of soil DNA revealed a high number of equally abundant faint bands, while rhizosphere fingerprints displayed a higher proportion of dominant bands and reduced richness, suggesting selection of bacterial populations in this environment. Plant specificity was detected in the rhizosphere by bacterial and group-specific DGGE profiles. Different bulk soil community fingerprints were revealed for each sampling site. The plant species was a determinant factor in shaping similar actinobacterial communities in the strawberry rhizosphere from different sites in both years. Higher heterogeneity of DGGE profiles within soil and rhizosphere replicates was observed for the fungi. Plant-specific composition of fungal communities in the rhizosphere could also be detected, but not in all cases. Cloning and sequencing of 16S rRNA gene fragments obtained from dominant DGGE bands detected in the bacterial profiles of the Rostock site revealed that Streptomyces sp. and Rhizobium sp. were among the dominant ribotypes in the strawberry rhizosphere, while sequences from Arthrobacter sp. corresponded to dominant bands from oilseed rape bacterial fingerprints.     

Viruses and flagellates sustain apparent richness and reduce biomass accumulation of bacterioplankton in coastal marine waters

To gain a better understanding of the interactions among bacteria, viruses and flagellates in coastal marine ecosystems, we investigated the effect of viral lysis and protistan bacterivory on bacterial abundance, production and diversity [determined by 16S rRNA gene polymerase chain reaction (PCR) and denaturing gradient gel electrophoresis (DGGE)] in three coastal marine sites with different nutrient supplies in Hong Kong. Six experiments were set up using filtration and dilution methods to develop virus, flagellate and virus+flagellate treatments for natural bacterial populations. All three predation treatments had significant repressing effects on bacterial abundance. Bacterial production was significantly repressed by flagellates and both predators (flagellates and viruses). Bacterial apparent species richness (indicated as the number of DGGE bands) was always significantly higher in the presence of viruses, flagellates and both predators than in the predator-free control. Cluster analysis of the DGGE patterns showed that the effects of viruses and flagellates on bacterial community structure were relatively stochastic while the co-effects of predators caused consistent trends (DGGE always showed the most similar patterns when compared with those of in situ environments) and substantially increased the apparent richness. Overall, we found strong evidence that viral lysis and protist bacterivory act additively to reduce bacterial production and to sustain diversity. This first systematic attempt to study the interactive effects of viruses and flagellates on the diversity and production of bacterial communities in coastal waters suggests that a tight control of bacterioplankton dominants results in relatively stable bacterioplankton communities.     

利用细胞计数手段和DGGE技术分析松花江干流部分地区的细菌种群多样性

松花江是我国东北地区的重要河流之一,为加强对其水环境微生物状况的了解,对松花江干流部分地区的微生物数量和多样性进行了分析。应用传统平板培养法和流式细胞技术测定水样中的细菌数;直接提取样品中的总DNA,以巢式PCR(Polymerase Chain Reaction)扩增细菌16SrDNA片段,应用聚丙烯酰胺凝胶电泳(Denaturing Gradient Gel Electrophoresis,DGGE)技术对扩增片段进行分离,研究水样和底泥样品细菌的种群多样性。实验结果显示,pH值为影响水环境中微生物总细胞数量的主要因素。水样中细菌群落多样性主要根据上下游分区,分区点在哈尔滨段附近,而底泥中细菌群落多样性的影响因素呈多样化,没有显示出较为明确的分区特征。     

Bacterial Community Succession in Natural River Biofilm Assemblages

Temporal bacterial community changes in river biofilms were studied using 16S rRNA gene-based polymerase chain reaction–denaturing gradient gel electrophoresis (DGGE) followed by sequence analysis. Naturally occurring biofilms were sampled in 2001 during an undisturbed 7-month low-water period in the River Garonne (SW France). During the sampling period epilithic biomass exhibited a particular pattern: two 3-month periods of accumulation that resulted in two peaks in summer and fall, each at about 25 g ash-free dry mass per square meter. Bacterial community DGGE profiles differed between the summer and fall biomass peaks and shared only 30% common operational taxonomic units (OTUs), suggesting the influence of seasonal factors on these communities. During the second biomass accrual phase, bacterial richness and the appearance of new OTUs fitted a conceptual model of bacterial biofilm succession. During succession, five OTUs (corresponding to Dechloromonas sp., Nitrospira sp., and three different Spirosoma spp.) exhibited particular patterns and were present only during clearly defined successional stages, suggesting differences in life-history strategies for epilithic bacteria. Co-inertia analysis of DGGE banding patterns and physical–chemical data showed a significant relationship between community structure and environmental conditions suggesting that bacterial communities were mainly influenced by seasonal changes (temperature, light) and hydrodynamic stability. Within the periods of stability, analysis of environmental variables and community patterns showed the dominant influence of time and maturation on bacterial community structure. Thus, succession in these naturally occurring epilithic biofilm assemblages appears to occur through a combination of allogenic (seasonal) and autogenic changes.     

太湖浮游细菌与春末浮游藻类群落结构演替的相关分析

为研究浮游细菌与浮游藻类群落演替的相关性,2005年4月至6月在太湖5个观测点采集浮游细菌及浮游藻类样本。分别采用聚合酶链式反应-变性梯度凝胶电泳（PCR—DGGE）和显微观察的方法分析浮游细菌及浮游藻类群落组成。结果表明,春末夏初,浮游细菌与藻类均呈现较高的多样性,浮游细菌DGGE图谱具有43种不同条带,浮游藻类的常见种有29种。浮游细菌群落聚类分析显示,丝藻（Ulothrix sp．）和微囊藻（Microcystis spp．）占优势时,浮游细菌群落基因组成存在明显差异。以藻类种群Shannon—Wiener多样性指数（Hp）,浮游藻类总细胞数（N）以及Microcystis spp．（M）百分含量为变量,典型对应分析（CCA）结果显示浮游细菌与浮游藻类群落结构变化的相关系数为30．9％,表明春末夏初太湖浮游细菌与浮游藻类群落演替具有较高的相关性。     

Seasonal and successional influences on bacterial community composition exceed that of protozoan grazing in river biofilms

The effects of protozoa (heterotrophic flagellates and ciliates) on the morphology and community composition of bacterial biofilms were tested under natural background conditions by applying size fractionation in a river bypass system. Confocal laser scanning microscopy (CLSM) was used to monitor the morphological structure of the biofilm, and fingerprinting methods (single-stranded conformation polymorphism [SSCP] and denaturing gradient gel electrophoresis [DGGE]) were utilized to assess changes in bacterial community composition. Season and internal population dynamics had a greater influence on the bacterial biofilm than the presence of protozoa. Within this general framework, bacterial area coverage and microcolony abundance were nevertheless enhanced by the presence of ciliates (but not by the presence of flagellates). We also found that the richness of bacterial operational taxonomic units was much higher in planktonic founder communities than in the ones establishing the biofilm. Within the first 2 h of colonization of an empty substrate by bacteria, the presence of flagellates additionally altered their biofilm community composition. As the biofilms matured, the number of bacterial operational taxonomic units increased when flagellates were present in high abundances. The additional presence of ciliates tended to at first reduce (days 2 to 7) and later increase (days 14 to 29) bacterial operational taxonomic unit richness. Altogether, the response of the bacterial community to protozoan grazing pressure was small compared to that reported in planktonic studies, but our findings contradict the assumption of a general grazing resistance of bacterial biofilms toward protozoa.