嗜中性微好氧铁氧化菌研究进展

在弱酸至近中性微氧条件下,嗜中性微好氧铁氧化菌能够通过依赖氧气的呼吸机制将二价亚铁氧化成三价铁,并获得生长所需能量。这一生物铁氧化过程的主要产物之一是无定形羟基氧化铁——一种异化铁还原作用(铁呼吸)的理想底物,故可加速铁元素在氧化还原分界层的地质循环。有关嗜中性微好氧铁氧化菌的记载可追溯到19世纪30年代,但对其生理、生态与系统发育学的研究自20世纪90年代中期才取得显著进展,主要得益于专性铁氧化菌新种、属的成功培养与分离。已知微好氧铁氧化菌广泛分布于弱酸及近中性富铁地下水、湿地和深海等环境,其参与调控的铁氧化过程对铁及其他元素(如碳、氮、磷、锰和砷等)的生物地球化学循环具有重要意义。这类古老微生物在金属成矿、地壳演变、全球气候变化及其它生源要素地球化学过程中的作用研究已逐渐受到关注,正成为地质与环境微生物学领域的研究热点。主要总结国外近15a对嗜中性微好氧铁氧化菌的研究进展,包括其代谢机理、种类和分布、生态学研究方法和技术、以及细菌铁氧化作用的实际应用和环境意义等,并对今后研究方向提出展望。     

Digestive proteinases of red shrimp Pleoticus muelleri (Decapoda, Penaeoidea): partial characterization and relationship with molting

The present study describes the activity and some characteristics of proteinases in the hepatopancreas of red shrimp Pleoticus muelleri during the different stages of the molting cycle. Proteolytic activity was highest between pH 7.5 and 8. The hepatopancreatic protein content in the premolt stage was higher than in the other stages of the molting cycle (P<0.05). No significant differences were found in total proteolytic activity in the hepatopancreas when comparing molting stages. The proteolytic activity of the P. muelleri hepatopancreas enzyme preparations is the main responsibility of serine proteinases. TLCK, a trypsin inhibitor, reduced azocasein hydrolysis between 26% (intermolt) and 37% (premolt). TPCK, a chymotrypsin inhibitor, did not decrease hydrolytic activity, except for in postmolt. Low trypsin and chymotrypsin activities were found during intermolt, and increased in postmolt. The electrophoretogram of the enzyme extracts shows 12 bands of activity during intermolt (from 16.6 to 53.1 kDa). Some fractions were not detected in the postmolt and premolt stages. Three low molecular weight trypsin forms (17.4, 19.1 and 20 kDa) were found in all molting stages. One band of chymotrypsin (21.9 kDa) was observed in all molting stages. High molecular mass active bands (66-205 kDa) could not be characterized with inhibitors. Comparison of the protease-specific activity of the hepatopancreas of some species indicated a relationship between digestive enzyme activity and feeding habits of the shrimp. Omnivorous shrimp, such as Penaeus vannamei (syn: Litopenaeus vannamei) and Penaeus monodon, showed higher protease activity than the carnivorous shrimp, Penaeus californiensis (syn: Farfantepenaeus californiensis) and P. muelleri. In fact, the enzymatic activity in the hepatopancreas of P. muelleri showed variations in relation to feeding habit and molting cycle.     

Planktonic marine iron oxidizers drive iron mineralization under low‐oxygen conditions

Observations of modern microbes have led to several hypotheses on how microbes precipitated the extensive iron formations in the geologic record, but we have yet to resolve the exact microbial contributions. An initial hypothesis was that cyanobacteria produced oxygen which oxidized iron abiotically; however, in modern environments such as microbial mats, where Fe(II) and O₂ coexist, we commonly find microaerophilic chemolithotrophic iron‐oxidizing bacteria producing Fe(III) oxyhydroxides. This suggests that such iron oxidizers could have inhabited niches in ancient coastal oceans where Fe(II) and O₂ coexisted, and therefore contributed to banded iron formations (BIFs) and other ferruginous deposits. However, there is currently little evidence for planktonic marine iron oxidizers in modern analogs. Here, we demonstrate successful cultivation of planktonic microaerophilic iron‐oxidizing Zetaproteobacteria from the Chesapeake Bay during seasonal stratification. Iron oxidizers were associated with low oxygen concentrations and active iron redox cycling in the oxic–anoxic transition zone (<3 μm O₂, <0.2 μm H₂S). While cyanobacteria were also detected in this transition zone, oxygen concentrations were too low to support significant rates of abiotic iron oxidation. Cyanobacteria may be providing oxygen for microaerophilic iron oxidation through a symbiotic relationship; at high Fe(II) levels, cyanobacteria would gain protection against Fe(II) toxicity. A Zetaproteobacteria isolate from this site oxidized iron at rates sufficient to account for deposition of geologic iron formations. In sum, our results suggest that once oxygenic photosynthesis evolved, microaerophilic chemolithotrophic iron oxidizers were likely important drivers of iron mineralization in ancient oceans.     

Acquisition of epibiotic bacteria along the life cycle of the hydrothermal shrimp Rimicaris exoculata

The caridean shrimp Rimicaris exoculata dominates the fauna at several Mid-Atlantic Ridge hydrothermal vent sites. This shrimp has an enlarged gill chamber, harboring a dense ectosymbiotic community of chemoautotrophic bacteria associated with mineral oxide deposits. Until now, their acquisition is not fully understood. At three hydrothermal vent sites, we analyzed the epibionts diversity at different moult stages and also in the first stages of the shrimp life (eggs, hatched eggs (with larvae) and juveniles). Hatched eggs associated with young larvae were collected for the first time directly from gravid females at the Logachev vent site during the Serpentine cruise. An approach using 16S rRNA clone libraries, scanning and transmission electron microscopy, and fluorescent in situ hybridization was used. Molecular results and microscope observations indicated a switch in the composition of the bacterial community between early R. exoculata life cycle stage (egg libraries dominated by the Gammaproteobacteria) and later stages (juvenile/adult libraries dominated by the Epsilonproteobacteria). We hypothesized that the epibiotic phylotype composition could vary according to the life stage of the shrimp. Our results confirmed the occurrence of a symbiosis with Gammaproteobacteria and Epsilonproteobacteria, but more complex than previously assumed. We revealed the presence of active type-I methanotrophic bacteria colonizing the cephalothorax of shrimps from the Rainbow site. They were also present on the eggs from the Logachev site. This could be the first ‘epibiotic'' association between methanotrophic bacteria and hydrothermal vent crustacean. We discuss possible transmission pathways for epibionts linked to the shrimp life cycle.     

Changes in integument histology and protein expression related to the molting cycle of the black tiger shrimp, Penaeus monodon

We investigated changes in the histology and protein expression in the epidermis and sub-epidermis of the black tiger shrimp (Penaeus monondon) during the molting cycle. The epidermis consists of a cell layer located beneath the cuticle, while the sub-epidermis is mainly composed of sub-epidermal cells and tegumental glands. During the molting cycle, the epidermal cells increase in cell height and number, and the sub-epidermis increases in its storage of carbohydrate, protein, mucus, and other unidentified substances at the time of the active period of cuticular regeneration. At the early premolt (stage D0), the epidermal cells are tidily organized, but short. Storage of carbohydrate and protein in the sub-epidermis is not observed. During the rest of the premolt (D1-4 stages) and the early postmolt A stage, epidermal cell height and sub-epidermal deposition are increased, and reached a maximum during the D4 to A stages. The period of late postmolt stages B-C3 is the time for a decrease in epidermal cell height and sub-epidermal depositions. Lastly at intermolt stage C4, the epidermal cells become short, and untidily organized. Sub-epidermal deposition is not observed. Protein expression in the epidermis and sub-epidermis was observed by SDS-PAGE. This revealed that the profile of a protein band with a molecular mass of 57 kDa corresponded with the profile observed by histochemistry. All results point to the conclusion that both the epidermis and sub-epidermis play major roles in cuticular regeneration. It may also reflect the level of metabolic activity of the integument during the molting cycle. In addition, for the first time, this work provides direct evidence of the epidermal and sub-epidermal changes that occur during the molting cycle of the black tiger shrimp.     

Neutrophilic lithotrophic iron-oxidizing prokaryotes and their role in the biogeochemical processes of the iron cycle

Biology of lithotrophic neutrophilic iron-oxidizing prokaryotes and their role in the processes of the biogeochemical cycle of iron are discussed. This group of microorganisms is phylogenetically, taxonomically, and physiologically heterogeneous, comprising three metabolically different groups: aerobes, nitratedependent anaerobes, and phototrophs; the latter two groups have been revealed relatively recently. Their taxonomy and metabolism are described. Materials on the structure and functioning of the electron transport chain in the course of Fe(II) oxidation by members of various physiological groups are discussed. Occurrence of iron oxidizers in freshwater and marine ecosystems, thermal springs, areas of hydrothermal activity, and underwater volcanic areas are considered. Molecular genetic techniques were used to determine the structure of iron-oxidizing microbial communities in various natural ecosystems. Analysis of stable isotope fractionation of ^56/54Fe in pure cultures and model experiments revealed a predominance of biological oxidation over abiotic ones in shallow aquatic habitats and mineral springs, which was especially pronounced under microaerobic conditions at the redox zone boundary. Discovery of anaerobic bacterial Fe(II) oxidation resulted in development of new hypotheses concerning the possible role of microorganisms and the mechanisms of formation of the major iron ore deposits during Precambrian era until the early Proterozoic epoch. Paleobiological data are presented on the microfossils and specific biomarkers retrieved from ancient ore samples and confirming involvement of anaerobic biogenic processes in their formation.     

Mate Guarding in the Androdioecious Clam Shrimp Eulimnadia texana: Male Assessment of Hermaphrodite Receptivity

Precopulatory mate guarding primarily occurs when males encounter receptive females at a low enough rate that such females become a valuable resource once encountered. Such circumstances are common in aquatic crustaceans wherein females are only receptive for a short period directly after molting. In these species, males commonly mate guard by physically attaching themselves to their prospective mates for hours to days at a time. To be effective in mate guarding, males must be able to assess the time to receptivity in their mates, which is commonly via chemical cues associated with molting. Clam shrimp in the genus Eulimnadia exhibit mate guarding, but with an important variation: these species are mixtures of males and hermaphrodites (androdioecy) rather than males and females. Nonetheless, the mate guarding behaviors of these shrimp are much the same as in other aquatic crustaceans. In this study, three projects were undertaken to determine the ability of Eulimnadia texana males to assess hermaphroditic receptivity. Males were found to be unable to assess receptivity without physically contacting hermaphrodites. However, after physical contact, males spent a significantly greater amount of time guarding receptive relative to non‐receptive hermaphrodites. Additionally, male interest in mate guarding was highest during the period between the dropping of one clutch of eggs and the extrusion of the following clutch. Because this period is also associated with hermaphroditic molting, it is consistent with the notion that males cue into chemicals associated with molting to determine hermaphroditic receptivity. These findings are consistent with previous studies of mating behavior in this species, and we discuss their importance to future tests of optimal mate guarding planned for these shrimp.     

Effect of short-term starvation on hepatopancreas and plasma energy reserves of the Pacific white shrimp (Litopenaeus vannamei)

Crustaceans are forced to fast during molting. Several physiological, metabolic and behavioral changes have been associated with starvation. Although some of these changes have been well studied, knowledge of the dynamics of fuel reserves during the molting process is limited. To understand the effects of short-term hunger stress on energy reserves, intermolt shrimp Litopenaeus vannamei were starved up to five days. This period corresponds to the normal time that juvenile shrimp starve during molting, since they can not eat. Glucose, glycogen, total soluble protein, total lipids, sterols, and acylglycerides were measured in plasma and hepatopancreas. The same metabolic substrates were measured in organisms that were fed after 96 h of starvation. It is widely accepted that protein is the main energy reserve used by shrimp to deal with starvation. However, under short-term starvation a rapid decrease of plasma and hepatopancreas glucose and an important decrease in hepatopancreatic glycogen were detected. Additionally, acylglycerides content in hepatopancreas decreased significantly at later times, while protein in plasma and hepatopancreas remained fairly constant during the experiment. This study may help understand some aspects of the nutrition physiology of the Pacific white shrimp related to its biology.     

Oxygen Dependency of Neutrophilic Fe(II) Oxidation by Leptothrix Differs from Abiotic Reaction

Neutrophilic Fe(II) oxidizing microorganisms are found in many natural environments. It has been hypothesized that, at low oxygen concentrations, microbial iron oxidation is favored over abiotic oxidation. Here, we compare the kinetics of abiotic Fe(II) oxidation to oxidation in the presence of the bacterium Leptothrix cholodnii Appels isolated from a wetland sediment. Rates of Fe(II) oxidation were determined in batch experiments at 20°C, pH 7 and oxygen concentrations between 3 and 120 μmol/l. The reaction progress in experiments with and without cells exhibited two distinct phases. During the initial phase, the oxygen dependency of microbial Fe(II) oxidation followed a Michaelis-Menten rate expression (K_M = 24.5 ± 10 μmol O₂/l, v_max = 1.8 ± 0.2 μmol Fe(II)/(l min) for 10⁸ cells/ml). In contrast, abiotic rates increased linearly with increasing oxygen concentrations. At similar oxygen concentrations, initial Fe(II) oxidation rates were faster in the experiments with bacteria. During the second phase, the accumulated iron oxides catalyzed further oxidative iron precipitation in both abiotic and microbial reaction systems. That is, abiotic oxidation also dominated the reaction progress in the presence of bacteria. In fact, in some experiments with bacteria, iron oxidation during the second phase proceeded slower than in the absence of bacteria, possibly due to an inhibitory effect of extracellular polymeric substances on the growth of Fe(III) oxides. Thus, our results suggest that the competitive advantage of microbial iron oxidation in low oxygen environments may be limited by the autocatalytic nature of abiotic Fe(III) oxide precipitation, unless the accumulation of Fe(III) oxides is prevented, for example, through a close coupling of Fe(II) oxidation and Fe(III) reduction.     

Influence of molting and starvation on the synthesis of proteolytic enzymes in the midgut gland of the white shrimp Penaeus vannamei

We investigated the effect of starvation as a stimulant of the digestive system on digestive proteinase activities in the white shrimp Penaeus vannamei. The starved organisms were sampled periodically according to the molting stage and compared with a continuously fed group. Molting stage was included as an independent variable. Most analyzed variables, except for trypsin, were more affected by starvation than by molting, indicating that starvation is a stimulant that masks the effect of molting and showing that food or alimentary stress is more conspicuous than physiological ones. We found that starvation is a stimulant that surpasses the effect of molting, and because it affects the activity of digestive proteinases, studies of starving organisms in combination with tools of molecular biology, can be a helpful working model in the understanding of mechanisms of regulation of digestive enzyme activity. In the starved organisms, trypsin and chymotrypsin activities were similar, suggesting dependence of one to the other. Changes in proteolytic activities and the number of protein bands in electrophoresis showed evidence of synthesis regulation in the midgut gland of white shrimp.     

Role of Ferrocyanides in the Prebiotic Synthesis of α-Amino Acids

We investigated the synthesis of α-amino acids under possible prebiotic terrestrial conditions in the presence of dissolved iron (II) in a simulated prebiotic ocean. An aerosol-liquid cycle with a prebiotic atmosphere is shown to produce amino acids via Strecker synthesis with relatively high yields. However, in the presence of iron, the HCN was captured in the form of a ferrocyanide, partially inhibiting the formation of amino acids. We showed how HCN captured as Prussian Blue (or another complex compound) may, in turn, have served as the HCN source when exposed to UV radiation, allowing for the sustained production of amino acids in conjunction with the production of oxyhydroxides that precipitate as by-products. We conclude that ferrocyanides and related compounds may have played a significant role as intermediate products in the prebiotic formation of amino acids and oxyhydroxides, such as those that are found in iron-containing soils and that the aerosol cycle of the primitive ocean may have enhanced the yield of the amino acid production.     

Sulfur Species as Redox Partners and Electron Shuttles for Ferrihydrite Reduction by Sulfurospirillum deleyianum

Iron(III) (oxyhydr)oxides can represent the dominant microbial electron acceptors under anoxic conditions in many aquatic environments, which makes understanding the mechanisms and processes regulating their dissolution and transformation particularly important. In a previous laboratory-based study, it has been shown that 0.05 mM thiosulfate can reduce 6 mM ferrihydrite indirectly via enzymatic reduction of thiosulfate to sulfide by the sulfur-reducing bacterium Sulfurospirillum deleyianum, followed by abiotic reduction of ferrihydrite coupled to reoxidation of sulfide. Thiosulfate, elemental sulfur, and polysulfides were proposed as reoxidized sulfur species functioning as electron shuttles. However, the exact electron transfer pathway remained unknown. Here, we present a detailed analysis of the sulfur species involved. Apart from thiosulfate, substoichiometric amounts of sulfite, tetrathionate, sulfide, or polysulfides also initiated ferrihydrite reduction. The portion of thiosulfate produced during abiotic ferrihydrite-dependent reoxidation of sulfide was about 10% of the total sulfur at maximum. The main abiotic oxidation product was elemental sulfur attached to the iron mineral surface, which indicates that direct contact between microorganisms and ferrihydrite is necessary to maintain the iron reduction process. Polysulfides were not detected in the liquid phase. Minor amounts were found associated either with microorganisms or the mineral phase. The abiotic oxidation of sulfide in the reaction with ferrihydrite was identified as rate determining. Cysteine, added as a sulfur source and a reducing agent, also led to abiotic ferrihydrite reduction and therefore should be eliminated when sulfur redox reactions are investigated. Overall, we could demonstrate the large impact of intermediate sulfur species on biogeochemical iron transformations.     

Biogeochemical controls on microbial diversity in seafloor sulphidic sediments

The ultimate fate of hydrothermal sulphides on the seafloor depends on the nature and rate of abiotic and microbially catalysed reactions where sulphide minerals are exposed to oxic seawater. This study combines organic and inorganic geochemical with microbiological measurements across a suboxic transition zone of highly altered sulphidic sediments from the Trans‐Atlantic Geotransverse hydrothermal field to characterize the reaction products and microbial communities present. There is distinct biogeochemical zonation apparent within the sediment sequence from oxic surface layers through a suboxic transition zone into the sulphide material. The microbial communities in the sediment differ significantly between the biogeochemical horizons sampled, with the identified microbes inferred to be associated with Fe and S redox cycling. In particular, Marinobacter species, organisms associated with circumneutral Fe oxidation, are dominant in a sulphide lens present in the lower core. The dominance of Marinobacter‐related sequences within the relict sulphide lens implies that these organisms play an important role in the alteration of sulphides at the seafloor once active venting has ceased.     

Notes on the life history of the clam shrimp,Eulimnadia texana

Several life history measures (growth rate, egg production, molt frequency, age at maturity and lifespan) were measured on several clam shrimp hermaphrodites (Eulimnadia texana Packard) grown in a laboratory setting under optimal growth conditions. Growth rates were high early in life, and then dropped dramatically when egg production began (day 5–6). Early egg production was low, and increased until approximately day 7, after which production leveled off for several days. Reproductive senescence was noted after day 17, with clutch sizes continuously dropping until death. Average molts per day was approximately 1.1, and molting seemed to be more closely associated with egg production than with growth. Growth and egg production were negatively correlated, indicating a possible trade-off between these two traits. No other trade-offs were detected. These shrimp show typical early-colonist life history traits, displaying high initial growth, early reproduction at a high rate, and then early senescence and death. This revised version was published online in August 2006 with corrections to the Cover Date.     

Variation in foraging investment during the intermolt interval and before egg-laying in the spiderNephila clavipes (Araneae: Araneidae)

Repeated observations were made of marked, freeliving females in three disjunct populations of the orb-weaving spider Nephila clavipes,recording orb size and structure and barrier web complexity. These longitudinal studies revealed regular patterns of variation in orb size associated with the molting cycle and cycle of egg maturation. Juveniles maintained in an insectary under a controlled feeding regime exhibited similar variation in orb size. All spiders ceased orb renewal prior to molting and the majority replaced the viscid orb with a nonviscid platform prior to molting. Differences among the populations in molting web structure are discussed in light of differences in the physical and biological environments. Cessation of orb renewal corresponded temporally with reported physiological changes associated with the molt cycle and possibly reflect changes in resource partitioning between growth and foraging during the molt cycle.     

Metagenomic resolution of microbial functions in deep-sea hydrothermal plumes across the Eastern Lau Spreading Center

Microbial processes within deep-sea hydrothermal plumes affect ocean biogeochemistry on global scales. In rising hydrothermal plumes, a combination of microbial metabolism and particle formation processes initiate the transformation of reduced chemicals like hydrogen sulfide, hydrogen, methane, iron, manganese and ammonia that are abundant in hydrothermal vent fluids. Despite the biogeochemical importance of this rising portion of plumes, it is understudied in comparison to neutrally buoyant plumes. Here we use metagenomics and bioenergetic modeling to describe the abundance and genetic potential of microorganisms in relation to available electron donors in five different hydrothermal plumes and three associated background deep-sea waters from the Eastern Lau Spreading Center located in the Western Pacific Ocean. Three hundred and thirty one distinct genomic ‘bins'' were identified, comprising an estimated 951 genomes of archaea, bacteria, eukarya and viruses. A significant proportion of these genomes is from novel microorganisms and thus reveals insights into the energy metabolism of heretofore unknown microbial groups. Community-wide analyses of genes encoding enzymes that oxidize inorganic energy sources showed that sulfur oxidation was the most abundant and diverse chemolithotrophic microbial metabolism in the community. Genes for sulfur oxidation were commonly present in genomic bins that also contained genes for oxidation of hydrogen and methane, suggesting metabolic versatility in these microbial groups. The relative diversity and abundance of genes encoding hydrogen oxidation was moderate, whereas that of genes for methane and ammonia oxidation was low in comparison to sulfur oxidation. Bioenergetic-thermodynamic modeling supports the metagenomic analyses, showing that oxidation of elemental sulfur with oxygen is the most dominant catabolic reaction in the hydrothermal plumes. We conclude that the energy metabolism of microbial communities inhabiting rising hydrothermal plumes is dictated by the underlying plume chemistry, with a dominant role for sulfur-based chemolithoautotrophy.     

Pathways of carbon and energy metabolism of the epibiotic community associated with the deep-sea hydrothermal vent shrimp Rimicaris exoculata

Background

The shrimp Rimicaris exoculata dominates the faunal biomass at many deep-sea hydrothermal vent sites at the Mid-Atlantic Ridge. In its enlarged gill chamber it harbors a specialized epibiotic bacterial community for which a nutritional role has been proposed.

Methodology/Principal Findings

We analyzed specimens from the Snake Pit hydrothermal vent field on the Mid-Atlantic Ridge by complementing a 16S rRNA gene survey with the analysis of genes involved in carbon, sulfur and hydrogen metabolism. In addition to Epsilon- and Gammaproteobacteria, the epibiotic community unexpectedly also consists of Deltaproteobacteria of a single phylotype, closely related to the genus Desulfocapsa. The association of these phylogenetic groups with the shrimp was confirmed by fluorescence in situ hybridization. Based on functional gene analyses, we hypothesize that the Gamma- and Epsilonproteobacteria are capable of autotrophic growth by oxidizing reduced sulfur compounds, and that the Deltaproteobacteria are also involved in sulfur metabolism. In addition, the detection of proteobacterial hydrogenases indicates the potential for hydrogen oxidation in these communities. Interestingly, the frequency of these phylotypes in 16S rRNA gene clone libraries from the mouthparts differ from that of the inner lining of the gill chamber, indicating potential functional compartmentalization.

Conclusions

Our data show the specific association of autotrophic bacteria with Rimicaris exoculata from the Snake Pit hydrothermal vent field, and suggest that autotrophic carbon fixation is contributing to the productivity of the epibiotic community with the reductive tricarboxylic acid cycle as one important carbon fixation pathway. This has not been considered in previous studies of carbon fixation and stable carbon isotope composition of the shrimp and its epibionts. Furthermore, the co-occurrence of sulfur-oxidizing and sulfur-reducing epibionts raises the possibility that both may be involved in the syntrophic exchange of sulfur compounds, which could increase the overall efficiency of this epibiotic community.