北黄海沉积物可培养产蛋白酶细菌分离鉴定

【目的】揭示北黄海沉积物中可培养产胞外蛋白酶细菌及蛋白酶多样性,增加人们对北黄海生态系统中产蛋白酶菌多样性的认识,为海洋产蛋白酶微生物的挖掘提供菌群资源。【方法】分别将5个北黄海沉积物样品梯度稀释涂布至酪蛋白明胶筛选平板,选择性分离产蛋白酶细菌;并通过分析基于16S rRNA基因序列的系统发育关系,揭示这些细菌的分类地位和遗传多样性;分别测定胞外蛋白酶活性并对酶活较高的39株菌进行基于苯甲基磺酰氟(PMSF,丝氨酸蛋白酶抑制剂)、邻菲罗啉(o-phenanthroline,O-P,金属蛋白酶抑制剂)、E-64(半胱氨酸蛋白酶抑制剂)和pepstatin A(天冬氨酸蛋白酶抑制剂)4种抑制剂的酶活抑制实验以及所有菌株对3种底物(酪蛋白、明胶、弹性蛋白)的水解能力;分析这些细菌所产胞外蛋白酶的特性及多样性。【结果】从5个北黄海沉积物样品中分离获得66株产蛋白酶细菌,这些菌株隶属于Bacteroidetes、Proteobacteria、Actinobacteria和Firmicutes 4个门的7个属,其中Pseudoalteromonas(69.9%)、Sulfitobacter(12.1%)和Salegentibacter(10.6%)是优势菌群;沉积物中可培养的产蛋白酶细菌的丰度为104 CFU/g;蛋白酶酶活抑制实验表明所有测定菌株产生的胞外蛋白酶属于丝氨酸蛋白酶和/或金属蛋白酶,仅有少数菌株所产蛋白酶具有半胱氨酸蛋白酶或天冬氨酸蛋白酶活性。【结论】北黄海沉积物中可培养产蛋白酶细菌类群较为丰富,Pseudoalteromonas、Sulfitobacter和Salegentibacter菌株是优势菌群,测定菌株所产胞外蛋白酶主要是丝氨酸蛋白酶和/或金属蛋白酶。     

Spatial structure and ecological efficiency in the summer zooplankton of a glacial lake

In June–July of 1993 and 1995, pelagic zooplankton dynamics of aglacial lake was investigated. In those years weather conditions diffferedsubstantially. This resulted not only in a great difference in the watersurface temperature (in 1993, <17.8± 0.47 °C; in 1995, 20.4± 0.33 °C), but also in an increase in the depth of metalimnionzone in 1995 in comparison with 1993. The ecological efficiency(K_e, the ratio of secondary to primary consumer productions,was 0.28 ± 0.031 in 1993 and 0.21± 0.034 in 1995. The higherK_e was accompanied by lowering of spatial structurization,primary and secondary consumer production. This revised version was published online in July 2006 with corrections to the Cover Date.     

Flagellate grazing on bacteria in a small dystrophic lake

Fluorescent beads were used to determine the grazing on bacteria by heterotrophic and mixotrophic flagellates in a highly humic (water colour 300–600 mg Pt l^–1) small lake. In summer phagotrophic flagellates constituted about three quarters of the numbers of phytoplankton (including heterotrophic or mixotrophic flagellates) in the uppermost epilimnion. Due to their small size their respective contribution to the biomass was about one quarter. The most important phagotrophic species were Ochromonas sp., and Chromulina spp. which ingested 75–203% of their body carbon per day from bacteria.In view of the abundance and biomass of phagotrophic and mixotrophic flagellates and their very high growth potential, they clearly play a significant role in the food chains of this lake. In addition to providing energy, bacteriovory also represents an important supply of inorganic and organic nutrients under nutrient limiting conditions.     

Impact of a flood event on the zooplankton of an estuarine lake

Shallow coastal lakes are prone to large fluctuations in physico-chemical variables such as salinity and turbidity. This is now escalating in response to global change. A flood event in March 2014 resulted in a silt plume spreading through part of Lake St Lucia (South Africa). To determine the impact of this event on zooplankton, the Narrows region of St Lucia was sampled on a monthly basis from March to September 2014. For comparative purposes, data from samples collected prior to the flood event were included in the analyses. Analysis of similarity (ANOSIM) revealed dissimilarities in zooplankton community structure among the sampling occasions. The March 2014–May 2014 period was characterized by the highest abundance of freshwater species. Conversely, the abundance of the resident St Lucia copepods Acartiella natalensis and Oithona brevicornis was lowest during this time, and highest in September 2014. The other dominant copepod Pseudodiaptomus stuhlmanni prevailed in March 2014, but declined markedly in April. As of September 2014, P. stuhlmanni had yet to regain its pre-flood densities. The BIOENV procedure, which relates biological and environmental data, revealed that turbidity, salinity and dissolved oxygen were responsible for the observed changes in zooplankton community structure during the study period. Careful management of turbidity and salinity is stressed, as both factors are major drivers of the biota of St Lucia and similar systems worldwide.     

Growth limitation of planktonic bacteria in a large mesotrophic lake

We studied nutrient-limitation of bacterioplankton growth in Lake Constance, a mesotrophic lake, between February and August in 1992. We amended 1-m filtrates with a single nutrient or nutrient combinations at 5 or 10 m final concentration, and the limiting nutrient or nutrient combination was inferred from the assay in which bacterial growth was most stimulated. The following nutrients were added individually or in combination: glucose, amino acids, peptone, and ammonium as C and N sources, and inorganic phosphate. From January until the beginning of the phytoplankton spring bloom in mid-April, C alone was growth-limiting. During the spring bloom a complex growth-limitation pattern occurred; first P was limiting, then for only 1 week C + N together, and thereafter P + C. During the clear-water phase with very low chlorophyll concentrations, P + C together limited bacterial growth again, interrupted by a period when C + N + P shortage caused a triple limitation. Later in the season, P + C were growth-limiting again. The growth efficiency (bacterial biomass produced/substrates used) on the basis of amino acid and carbohydrate used varied between 17 and 35%. The addition of various C and N sources indicated that the growth efficiency strongly depended on the quality of the substrates and the adaptation of the bacterial assemblages, for example, whether C and N originated from amino acids or glucose and ammonium.     

Zooplankton grazing on bacteria and cyanobacteria in a eutrophic lake

During in situ experiments in a cyanobacteria- and copepod-dominatedeutrophic subtropical lake, all taxa of protozoa and metazoanzooplankton grazed fluorescently labelled bacteria and all metazoanzooplankton taxa grazed large filamentous and colonial cyanobacteria.     

Mercury inputs and outputs at a small lake in northern Minnesota

Storages and cycling of total mercury (Hg_T), methylmercury (MeHg), and Hg⁰ are described for Spring Lake, a small bog lake in the Marcell Experimental Forest in north-central Minnesota. We quantified photoredox transformations, MeHg photolysis, burial to the sediments, and internal and external loadings of Hg_T and MeHg. Atmospheric deposition was the main input of Hg_T; MeHg was supplied by a combination of atmospheric, near-shore wetland, and biotic (methylation) sources. Hg_T outputs were dominated by burial (67%), and Hg⁰ evasion accounted for 26% of Hg_T outputs. The watershed of Spring Lake is small (3.7× lake surface area), and accordingly, bog and upland runoff were minor contributors to both Hg_T and MeHg inputs. Wet deposition was ∼9% of total MeHg input, and other external inputs (runoff, sediment porewater) provided only an additional 7%, indicating that internal production of MeHg was occurring in the lake. Photolysis of MeHg, measured in the field and laboratory, removed ∼3× the lake mass of MeHg (20 mg) annually, and was the dominant sink for MeHg. Residence times of MeHg and Hg_T in the lake were 48 and 61 days, respectively, during the open-water season, compared with only 8 days for the residence time of MeHg on settling particles (seston). Photoreduction of Hg²⁺ to Hg⁰ was greater than the reverse reaction (Hg⁰ photooxidation), and the residence time of Hg⁰ in the photic zone was short (hours). Data from this study show active cycling of all the measured species of mercury (Hg_T, MeHg, and Hg⁰) and the importance of MeHg photolysis and photo-redox processes.     

Evidence for cryptic northern refugia in the last glacial period in Cryptomeria japonica

MethodsGenetic diversity and structure were examined using 20 microsatellite markers in 37 populations of C. japonica. The locations of glacial refugia were assessed using STRUCTURE analysis, and potential habitats under current and past climate conditions were predicted using SDM. The process of genetic divergence was also examined using the approximate Bayesian computation procedure (ABC) in DIY ABC to test the divergence time between the gene pools detected by the STRUCTURE analysis.ConclusionsThe combined evidence from microsatellites and SDM clearly indicates that climatic changes have shaped the genetic structure of C. japonica. The gene pool detected in northern Tohoku district is likely to have been established by cryptic northern refugia on the coast of the Japan Sea to the west of the Archipelago. The gene pool in Yakushima Island can probably be explained simply by long-term isolation from the other gene pools since the LGM. These results are supported by those of SDM and the predicted divergence time determined using ABC analysis.     

The relationship between planktonic algae and bacteria in a small lake

The seasonal changes and vertical distribution of the aerobic and anaerobic bacteria in a small edaphically eutrophic lake which exhibited thermal and chemical stratification are described. There was some correspondence between the phytoplankton and particularly the aerobic bacteria but this was not consistent. Increases in the numbers of anaerobic bacteria coincided with the low dissolved oxygen concentrations in the hypolimnion when algal populations were first senescent and then increasing actively in size.     

Distribution of predatory bacteria that attack chromatiaceae in a sulfurous lake

Predatory bacteria that attack Chromatiaceae (purple sulfur bacteria) recovered from Lake Cisó (a mostly anaerobic holomictic lake) have been studied over two annual cycles. During the mixing period the lake was completely anaerobic; both predator and prey populations were found along the water column, and even at the surface. Throughout the stratification period maximum Chromatiaceae occurred between a depth of 1 and 3 m depth. The maximum numbers of predators and prey (Chromatiaceae) also occurred in this range.A collapse took place in the lake during the second annual cycle in 1986. It brought about changes in the physicochemical parameters of the lake, thus altering the population dynamics. Nevertheless, during both cycles the number of predatory bacteria was maximum immediately below the depth at which the maximum number of prey bacteria occurred. Send offprint requests to: N. Gaju.     

Use of dissolved carbohydrates by planktonic bacteria in a mesotrophic lake

Dissolved carbohydrates comprise one of the largest pools of labile organic matter readily available for bacterial use in pelagic ecosystems. Despite this fact, very little is known about use of dissolved carbohydrates by planktonic bacteria. We studied use of total dissolved carbohydrates (TDCHO) by planktonic bacteria in mesotrophic Lake Constance, Germany, from April until August 1992. We examined the decrease of TDCHO over time together with the increase of bacterial numbers in 1-m filtered lake water incubated at in situ temperature in the dark. TDCHO analyses were done after hydrolysis by sulfuric acid as free monosaccharides and oxidation by periodate to formaldehyde with 3-methyl-2-benzothiazolinone-hydrazon-hypochloride (MBTH). Preliminary tests showed that hydrolysis by sulfuric acid gave higher yields than hydrolysis by HCl. Our results show that TDCHO are readily used by bacteria and compose substantial fractions of the C requirements for their growth. Concentrations of TDCHO varied between 1.7 and 5.5 m (glucose equivalents), and use rates varied between 0.47 and 3.43 g C liter^–1 h^–1. Highest rates of TDCHO use occurred during the phytoplankton spring bloom, during the clear-water phase in June, and during a phytoplankton bloom in August. Ratios of use of TDCHO/bacterial biomass production varied between 0.17 and 3.05. During the spring bloom, TDCHO and total dissolved amino acids (TDAA) were used in equal amounts. During the clear-water phase at chlorophyll a concentrations <3 g liter^–1, however, bacteria only consumed TDCHO and excreted amino acids. The growth efficiency based on the consumption of TDCHO and TDAA varied between 16 and 21% during the phytoplankton spring bloom and was 35% during the clear-water phase. Correspondence to: M. Simon     

Bacterial grazing by phagotrophic phytoflagellates in a deep humic lake in northern Sweden

Bacterial grazing was measured from June to August 1995 in Lake Ortrasket, a deep brown-water lake in northern Sweden. Mixotrophic chrysophytes were the dominating bacterivores at all times, grazing 3-14% of bacterial standing stock daily. The effects of altered nutrient supply and light availability on grazing activity and growth were studied in two mesocosm experiments. Incubation in the dark did not stimulate phagotrophy, which would otherwise be expected if bacteria were mainly being used as an energy source. Furthermore, clearance rates were not reduced after alleviation of nutrient limitation conditions. Rather, phagotrophy may work as a relatively fixed attribute of the mixotrophic community in this lake. When availability of dissolved nutrients is restricted, phagotrophy permits the mixotrophs to outcompete other phytoplankton, but they become less competitive at high nutrient concentrations. The relative share of mixotrophs in relation to total phytoplankton decreased considerably after enrichment with nitrogen + phosphorus.      

Community shifts of actively growing lake bacteria after N-acetyl-glucosamine addition: improving the BrdU-FACS method

In aquatic environments, community dynamics of bacteria, especially actively growing bacteria (AGB), are tightly linked with dissolved organic matter (DOM) quantity and quality. We analyzed the community dynamics of DNA-synthesizing and accordingly AGB by linking an improved bromodeoxyuridine immunocytochemistry approach with fluorescence-activated cell sorting (BrdU-FACS). FACS-sorted cells of even oligotrophic ecosystems in winter were characterized by 16S rRNA gene analysis. In incubation experiments, we examined community shifts of AGB in response to the addition of N-acetyl-glucosamine (NAG), one of the most abundant aminosugars in aquatic systems. Our improved BrdU-FACS analysis revealed that AGB winter communities of oligotrophic Lake Stechlin (northeastern Germany) substantially differ from those of total bacteria and consist of Alpha-, Beta-, Gamma-, Deltaproteobacteria, Actinobacteria, Candidatus OP10 and Chloroflexi. AGB populations with different BrdU-fluorescence intensities and cell sizes represented different phylotypes suggesting that single-cell growth potential varies at the taxon level. NAG incubation experiments demonstrated that a variety of widespread taxa related to Alpha-, Beta-, Gammaproteobacteria, Bacteroidetes, Actinobacteria, Firmicutes, Planctomycetes, Spirochaetes, Verrucomicrobia and Chloroflexi actively grow in the presence of NAG. The BrdU-FACS approach enables detailed phylogenetic studies of AGB and, thus, to identify those phylotypes which are potential key players in aquatic DOM cycling.     

Small-scale spatial beta diversity of bacteria in the mixed upper layer of a lake

Bacterial community composition among individual, experimentally generated ‘lake snow’ particles may be highly variable. Since such aggregates are seasonally abundant in the mixed upper layer of lakes, we hypothesized that particle-attached (PA) bacteria disproportionally contribute to the small-scale spatial beta diversity of pelagic communities. Community composition was analysed in sets of small (10 mL) samples collected from a pre-alpine lake in May, July and October 2018. Bacteria were classified as free-living (FL) or PA depending on their presence in large, 5-μm pre-filtered reference samples. FL exhibited clear seasonal differences in community composition and assembly. They were spatially uniform in May and July, and only a few FL taxa exhibited significant spatial variability. Spatial heterogeneity of FL in October was caused by high alpha and beta diversity of rare taxa, many with a presumably ‘tychoplanktic’ (alternating attached and free-living) lifestyle. The spatial beta diversity of PA was always high, and only about 10% of their seasonal richness was present in any single sample. Thus, most compositional variability of pelagic bacteria at spatial scales of cm to m either directly or indirectly originated from PA. On a functional level, this genotypic heterogeneity might affect the spatial distribution of rare metabolic traits.     

The effects of flood pulse on multiple aquatic organisms in a seasonal shallow lake

Different aquatic organisms have different reactions to environmental variations due to their different ways of reaction traits. Understanding the effect of hydrological disturbance on lake from the perspective of multiple aquatic organisms is important for lake management. Poyang Lake (PYL) experiences severe hydrological disturbance under the effects of flood pulse during the rainy season. In this study, we analyzed the responses of phytoplankton, zooplankton, and macroinvertebrates to hydrological disturbance during the rainy season in PYL. Flood pulse determined the spatial variation in turbidity (Turb), transparency (Trans), dissolved oxygen (DO), total phosphorus (TP), and soluble reactive phosphorus. Physical factors (Turb, Trans, conductivity) explained phytoplankton (9.6%) and zooplankton (15%) community variation, and the combination of physical and nutrient factors explained macroinvertebrate (3.8%) community variation. Cyclotella sp., Attheya zachariasi, and Melosira ambigua were the keystone taxa of phytoplankton, and these taxa were driven by Turb and Trans. Brachionus angularia, Polyarthra vulgaris, Filinia longiseta, and Diffugia globulosa were the keystone taxa of zooplankton and entire aquatic organisms, which contributed a lot to maintain the biological community stability possibly through the food web. These keystone taxa were stimulated by high Trans, DO, dissolved organic carbon (DOC) and nitrate (NO₃^?). We found habitat conditions (Turb, Trans and DO) were the prominent factors influencing the aquatic organism structure in strong hydrological disturbance environment. For the keystone taxa, both habitat conditions (i.e., DO) and exogenous nutrient inputs (i.e., DOC and NO₃^?) caused significant effect. This study provides new insights into the holistic response of multiple aquatic communities to flood pulse as well as the role of keystone species in maintaining community stability, which could guide the conservation and management of seasonal lake ecosystems.

     

Trophic interactions among native and introduced fishes in a northern Patagonian oligotrophic lake

In summer the littoral zone of Lake Rivadavia contained almost all the juveniles of the native fish species present in the lake together with juveniles of the introduced salmonid Oncorhynchus mykiss . In particular, a high degree of foraging interaction was found among juveniles of the native species Aplochiton zebra and Percichthys trucha and juveniles O. mykiss. Differences in the juvenile assemblage composition, both in the presence and in the relative proportion of the species were associated with areas with different densities of vegetation. The adults of nearly all species were generalized benthic invertebrate predators. Only adults of A. zebra were absent in the littoral. Juvenile and adult O. mykiss were the only fish that preyed on planktonic organisms. Oncorhynchus mykiss was the only species that performed an ontogenetic diet shift between juveniles and adults, changing main prey categories from benthic to planktonic species.     

Trophic relationships between bacteria and protozoa in the hypolimnion of a meromictic mesostrophic lake

The development of ciliated protozoan biomass in the hypolimnion of Piburger See, a small subalpine lake, was demonstrated to depend mainly on two factors. Firstly, the availability of oxygen or nitrate as electron acceptors determines the depth profiles of ciliates. Large quantities of ciliates and even maximum numbers were found at depths where no oxygen could be detected. If nitrate also disappeared during the summer stagnation period, the biomass of protozoa was strongly reduced. Nitrite peaks generally corresponded with ciliate peaks. An extension of Finlay's findings (dissimilatory nitrate reductase within the inner mitochondrial membrane) to other ciliate groups is hypothesized.Secondly, the biomass development of hypolimnetic ciliates was strongly correlated with the bacterial biomass registered approx. 2 weeks before (r² = 0.891, n = 14). The biomass of bacteria, on the other hand, was dependent upon the sedimentation rate of organic carbon (r² = 0.850, n = 15), if a time lag of approx. 2 weeks was taken into account. Therefore a total time lag of approx. 4 weeks was assumed to take place between sedimentation of organic substance and the corresponding increase in ciliate biomass (r² = 0.853, n = 14). Bacteria were shown to be an important intermediate link in the food chain of the hypolimnion. They appear to represent the principal energy source for pelagic ciliates. Sedimentation of organic carbon, nitrogen and phosphorus is the driving force for the establishment of the hypolimnetic microbial community.     

Survival of sulfate-reducing bacteria in oxic surface sediment of a seawater lake

Abstract Microhabitats and survival of sulfate-reducing bacteria (SRB) in an oxic surface sediment of a seawater lake were examined. The size of fractionation of the sediment suspension showed that most of SRB were associated with sediment particles larger than 10 μm. The D values (time in h required to destroy 90% of the initial viable population) for SRB in the whole sediment suspension and for SRB i n the < μ m and the < 5 μ m fractions were, respectively, 23.7, 10 and 4 when the SRB were exposed to air. Survival of the FeS-associated Desulfovibrio desulfuricans ( D value, 9.3) was higher than that of the free-living ones ( D value, 1.8). These results show that particle-associated SRB are more protected against oxygen than free-living ones in oxic sediments.     

Vertical distribution of major sulfate-reducing bacteria in a shallow eutrophic meromictic lake

The vertical distribution of sulfate-reducing bacteria was investigated in a shallow, eutrophic, meromictic lake, Lake Harutori, located in a residential area of Kushiro, Japan. A steep chemocline, characterized by gradients of oxygen, sulfide and salinity, was found at a depth of 3.5–4.0 m. The sulfide concentration at the bottom of the lake was high (up to a concentration of 10.7 mM). Clone libraries were constructed using the aprA gene, which encodes adenosine-5′-phosphosulfate reductase subunit A, in order to monitor sulfate-reducing bacteria. In the aprA clone libraries, the most abundant sequences were those from the Desulfosarcina–Desulfococcus (DSS) group. A primer set for a DSS group-specific 16S rRNA gene was used to construct another clone library, analysis of which revealed that the uncultured group of sulfate-reducing bacteria, SEEP SRB-1, accounted for nearly half of the obtained sequences. Quantification of the major bacterial groups by catalyzed reporter deposition-fluorescence in situ hybridization demonstrated that the DSS group accounted for 3.2–4.8% of the total bacterial community below the chemocline. The results suggested that the DSS group was one of the major groups of sulfate-reducing bacteria and that these presumably metabolically versatile bacteria might play an important role in sulfur cycling in Lake Harutori.