Potentially Active Iron,Sulfur, and Sulfate Reducing Bacteria in Skagerrak and Bothnian Bay Sediments

In many marine surface sediments, the reduction of manganese (Mn) and iron (Fe) oxides is obscured by sulfate reduction, which is regarded as the predominant anaerobic microbial respiration process. However, many dissimilatory sulfate and sulfur reducing microorganisms are known to utilize alternative electron acceptors such as metal oxides. In this study, we tested whether sulfate and sulfur reducing bacteria are linked to metal oxide reduction based on biogeochemical modeling of porewater concentration profiles of Mn²⁺ and Fe²⁺ in Bothnian Bay (BB) and Skagerrak (SK) sediments. Steady-state modeling of Fe²⁺ and Mn²⁺ porewater profiles revealed zones of net Fe (0–9 cm BB; ～10 and 20 cm SK) and Mn (0–5 cm BB; 2–8 cm SK) species transformations. 16S rRNA pyrosequencing analysis of the in-situ community showed that Desulfobacteraceae, Desulfuromonadaceae and Desulfobulbaceae were present in the zone of Fe-reduction of both sediments. Rhodobacteraceae were also detected at high relative abundance in both sediments. BB sediments appeared to harbor a greater diversity of potential Fe-reducers compared to SK. Additionally, when the upper 10 cm of sediment from the SK was incubated with lepidocrocite and acetate, Desulfuromonas was the dominant bacteria. Real-time quantitative polymerase chain reaction (qPCR) results showed decreasing dsrA gene copy numbers with depth coincided with decreased Fe-reduction activity. Our results support the idea that sulfur and sulfate reducing bacteria contribute to Fe-reduction in the upper centimeters of both sediments.     

Impacts of the 2014 severe drought on the Microcystis bloom in San Francisco Estuary

The increased frequency and intensity of drought with climate change may cause an increase in the magnitude and toxicity of freshwater cyanobacteria harmful algal blooms (CHABs), including Microcystis blooms, in San Francisco Estuary, California. As the fourth driest year on record in San Francisco Estuary, the 2014 drought provided an opportunity to directly test the impact of severe drought on cyanobacteria blooms in SFE. A field sampling program was conducted between July and December 2014 to sample a suite of physical, chemical, and biological variables at 10 stations in the freshwater and brackish reaches of the estuary. The 2014 Microcystis bloom had the highest biomass and toxin concentration, earliest initiation, and the longest duration, since the blooms began in 1999. Median chlorophyll a concentration increased by 9 and 12 times over previous dry and wet years, respectively. Total microcystin concentration also exceeded that in previous dry and wet years by a factor of 11 and 65, respectively. Cell abundance determined by quantitative PCR indicated the bloom contained multiple potentially toxic cyanobacteria species, toxic Microcystis and relatively high total cyanobacteria abundance. The bloom was associated with extreme nutrient concentrations, including a 20-year high in soluble reactive phosphorus concentration and low to below detection levels of ammonium. Stable isotope analysis suggested the bloom varied with both inorganic and organic nutrient concentration, and used ammonium as the primary nitrogen source. Water temperature was a primary controlling factor for the bloom and was positively correlated with the increase in both total and toxic Microcystis abundance. In addition, the early initiation and persistence of warm water temperature coincided with the increased intensity and duration of the Microcystis bloom from the usual 3 to 4 months to 8 months. Long residence time was also a primary factor controlling the magnitude and persistence of the bloom, and was created by a 66% to 85% reduction in both the water inflow and diversion of water for agriculture during the summer. We concluded that severe drought conditions can lead to a significant increase in the abundance of Microcystis and other cyanobacteria, as well as their associated toxins.     

Shifts in microbial populations in Rusitec fermenters as affected by the type of diet and impact of the method for estimating microbial growth (15N v. microbial DNA)

Rusitec fermenters are in vitro systems widely used to study ruminal fermentation, but little is known about the microbial populations establishing in them. This study was designed to assess the time evolution of microbial populations in fermenters fed medium- (MC; 50% alfalfa hay : concentrate) and high-concentrate diets (HC; 15 : 85 barley straw : concentrate). Samples from solid (SOL) and liquid (LIQ) content of fermenters were taken immediately before feeding on days 3, 8 and 14 of incubation for quantitative polymerase chain reaction and automated ribosomal intergenic spacer analysis analyses. In SOL, total bacterial DNA concentration and relative abundance of Ruminococcus flavefaciens remained unchanged over the incubation period, but protozoal DNA concentration and abundance of Fibrobacter succinogenes, Ruminococcus albus and fungi decreased and abundance of methanogenic archaea increased. In LIQ, total bacterial DNA concentration increased with time, whereas concentration of protozoal DNA and abundance of methanogens and fungi decreased. Diet×time interactions were observed for bacterial and protozoal DNA and relative abundance of F. succinogenes and R. albus in SOL, as well as for protozoal DNA in LIQ. Bacterial diversity in SOL increased with time, but no changes were observed in LIQ. The incubated diet influenced all microbial populations, with the exception of total bacteria and fungi abundance in LIQ. Bacterial diversity was higher in MC-fed than in HC-fed fermenters in SOL, but no differences were detected in LIQ. Values of pH, daily production of volatile fatty acids and CH₄ and isobutyrate proportions remained stable over the incubation period, but other fermentation parameters varied with time. The relationships among microbial populations and fermentation parameters were in well agreement with those previously reported in in vivo studies. Using ¹⁵N as a microbial marker or quantifying total microbial DNA for estimating microbial protein synthesis offered similar results for diets comparison, but both methods presented contrasting results for microbial growth in SOL and LIQ phases. The study showed that fermentation parameters remained fairly stable over the commonly used sampling period (days 8 to 14), but shifts in microbial populations were detected. Moreover, microbial populations differed markedly from those in the inocula, which indicates the difficulty of directly transposing results on microbial populations developed in Rusitec fermenters to in vivo conditions.     

葱蝇热激蛋白DaHSP23基因的克隆及在冬滞育和夏滞育蛹中的表达分析（英文）

【目的】低分子量（12~43 kDa）热激蛋白(sHSPs)具有抗逆应答的功能,滞育是昆虫抵抗不良环境的特殊发育形式,但sHSPs在昆虫滞育发育过程中的作用仍不清楚。本研究克隆和特征化葱蝇Delia antiqua sHSP基因,并研究它在夏滞育和冬滞育发育过程中的表达模式,为阐明sHSPs在滞育发育上的功能奠定基础。【方法】通过RACE-PCR方法克隆了葱蝇HSP23基因,通过相似性比较分析了其特征、结构域及与双翅目代表性同源基因的系统发育关系;采用实时荧光定量PCR研究了该基因在葱蝇冬滞育蛹和夏滞育蛹发育过程中的表达情况,通过表达的差异比较揭示了该基因与滞育发育的关系。【结果】克隆出了葱蝇HSP23基因,命名为DaHSP23（GenBank登录号：HQ392521.1）,其cDNA全长序列为904 bp,编码186个氨基酸,推测蛋白分子量为20.9 kDa,等电点为6.42。该基因的编码蛋白与其他双翅目昆虫的sHSPs有超过66%的氨基酸序列一致性,与已报道的其他双翅目昆虫的滞育相关HSP23基因同源。基因组测序显示该基因无内含子。DaHSP23基因在葱蝇非滞育蛹的发育过程中一直保持在较低的水平,各发育阶段间的表达量不存在显著差异。但在冬滞育和夏滞育蛹中,该基因从滞育起始期开始逐渐显著升高表达,到滞育维持期的中后期达到峰值,在滞育终止期逐渐降到较低的水平。【结论】DaHSP23基因在葱蝇冬滞育和夏滞育发育过程中明显上调表达,但存在差异,它在滞育期的调控可能是种专化的。DaHSP23可能在葱蝇两种类型的滞育上起重要作用。     

Development of a quantitative PCR method to differentiate between viable and nonviable bacteria in environmental water samples

Ethidium monoazide bromide (EMA) treatment of pure culture and environmental waters at low concentrations (1.0–7.5 μg/ml) indicated effective enumeration of viable and viable but nonculturable Escherichia coli in pure cultures, creek waters, and secondary activated sludge effluent samples by quantitative polymerase chain reaction (qPCR) amplification of the uidA and fliC gene targets at turbidity values <10 NTU. However, EMA treatment was not effective in primary clarifier and secondary trickling filter effluents where turbidities were ≥10 NTU. In viable pure cultures, rapidly dividing and senescent cells were most affected by increasing EMA concentrations. Amplification of heat-killed pure bacterial cultures decreased 4 to 6 logs depending on EMA concentration and culture age. The greatest difference was observed in 5-h cultures using 7.5 μg/ml EMA. Turbidity (≥100 NTU) in environmental samples inhibited EMA effectiveness on viability discrimination. Enumeration of E. coli in certain wastewaters using EMA-qPCR was similar to culture suggesting that EMA treatment could be incorporated into qPCR assays for the quantification of viable bacteria increasing assay time no more than 30 min. Our results indicate that EMA can be used in routine qPCR assays, but optimum conditions for exposure must be identified for each sample type due to sample matrix effects such as turbidity.     

Zebrafish β-adrenergic receptor mRNA expression and control of pigmentation

Beta adrenergic receptors (β-ARs) are members of the G-protein-coupled receptor superfamily and mediate various physiological processes in many species. The expression patterns and functions of β-ARs in zebrafish are, however, largely unknown. We have identified zebrafish β-AR orthologs, which we have designated as adrb1, adrb2a, adrb2b, adrb3a and adrb3b. adrb1 was found to be expressed in the heart and brain. Expression of adrb2a predominated in the brain and skin, whereas adrb2b was found to be highly expressed in muscle, pancreas and liver. Both adrb3a and adrb3b were exclusively expressed in blood. Knock-down of these β-ARs by morpholino oligonucleotides revealed a functional importance of adrb2a in pigmentation. Expression of atp5a1 and atp5b, genes that encode subunits of F1F0-ATPase, which is known to be involved in pigmentation, was significantly increased by knock-down of adrb2a. Our data suggest that adrb2a may regulate pigmentation, partly by modulating F1F0-ATPase.     

实时定量PCR芯片——功能基因组研究的有用工具

结合实时定量PCR（real-time quantitative polymerase chain reaction,qPCR）和基因芯片技术,实时定量PCR芯片（qPCR array）为功能基因组研究提供了一种高度灵敏、可靠的方法。在基因功能分类基础上,结合qPCR准确定量的特性,qPCR array可同时定量分析上百个基因的mRNA表达水平,是研究一组特定功能基因表达的理想方法。