Impact of Peripheral Hydrogen Bond on Electronic Properties of the Primary Acceptor Chlorophyll in the Reaction Center of Photosystem I.

Photosystem I (PS I) is a photosynthetic pigment-protein complex that absorbs light and uses the absorbed energy to initiate electron transfer. Electron transfer has been shown to occur concurrently along two (A- and B-) branches of reaction center (RC) cofactors. The electron transfer chain originates from a special pair of chlorophyll a molecules (P700), followed by two chlorophylls and one phylloquinone in each branch (denoted as A-1, A0, A1, respectively), converging in a single iron-sulfur complex Fx. While there is a consensus that the ultimate electron donor-acceptor pair is P700+A0-, the involvement of A-1 in electron transfer, as well as the mechanism of the very first step in the charge separation sequence, has been under debate. To resolve this question, multiple groups have targeted electron transfer cofactors by site-directed mutations. In this work, the peripheral hydrogen bonds to keto groups of A0 chlorophylls have been disrupted by mutagenesis. Four mutants were generated: PsaA-Y692F; PsaB-Y667F; PsaB-Y667A; and a double mutant PsaA-Y692F/PsaB-Y667F. Contrary to expectations, but in agreement with density functional theory modeling, the removal of the hydrogen bond by Tyr → Phe substitution was found to have a negligible effect on redox potentials and optical absorption spectra of respective chlorophylls. In contrast, Tyr → Ala substitution was shown to have a fatal effect on the PS I function. It is thus inferred that PsaA-Y692 and PsaB-Y667 residues have primarily structural significance, and their ability to coordinate respective chlorophylls in electron transfer via hydrogen bond plays a minor role.

Bacterial community composition in the Northern Gulf of Mexico intertidal sediment bioturbated by the ghost shrimp Lepidophthalmus louisianensis.

Bioturbation plays an important role in structuring microbial communities in coastal sediments. This study investigates the bacterial community composition in sediment associated with the ghost shrimp Lepidophthalmus louisianensis at two locations in the Northern Gulf of Mexico (Bay St. Louis, MS, and Choctawhatchee Bay, FL). Bacteria were analysed for shrimp burrows and for three different depths of bioturbated intertidal sediment, using second-generation sequencing of the 16S rRNA gene. Burrow walls held a unique bacterial community, which was significantly different from those in the surrounding sediment communities. Communities in burrow walls and surrounding sediment communities also differed between the two geographic locations. The burrow wall communities from both locations were more similar to each other than to sediment communities from same location. Alpha- and Gammaproteobacteria were more abundant in burrows and surface sediment than in the subsurface, whereas Deltaproteobacteria were more abundant in burrows and subsurface sediment, suggesting sediment mixing by the bioturbator. However, abundance of individual ASVs was geographic location-specific for all samples. Therefore, it is suggested that the geographic location plays an important role in regional microbial communities distinctiveness. Bioturbation appears to be an important environmental driver in structuring the community around burrows. Sampling was conducted during times of the year and water salinity, tidal regime and temperature were variable, nevertheless the structure microbial communities appeared to remain realatively stable suggesting that these environmental variable played only a minor role.

Formulation of a Simulated Wastewater Influent Composition for Use in the Research of Technologies for Managing Wastewaters Generated during Manned Long-Term Space Exploration and Other Similar Situations-Literature-Based Composition Development.

The prospect of humans inhabiting planetary bodies is gaining interest among research and development communities, with the moon being considered as a transitory base camp and Mars the next planet humans will inhabit. NASA's Mission to Mars program is set to have humans inhabiting Mars within on-planet space camps by the Year 2030, which has tremendously increased research and development for space exploration-including research oriented toward human life support in long-term planetary lodging camps. The sustenance of human life on Mars will not be trivial due to the unavailability of an appropriate atmosphere and usable water. This situation requires a self-sustaining human life support system that can provide the basic needs such are breathable air, potable water, food, and energy. The feasibility of sending a payload with resources adequate to support long-term human inhabitation is not reasonable, which means every resource within a Mars space camp is valuable, including human-produced wastes. A biorefinery system that treats wastewater and can also produce valuable products such as oxygen, food, and energy offers a form of circular utilization of valuable resources. To conduct research for such systems requires a wastewater influent that is representative of the wastewater to be generated by the space crew within this isolated, confined environment, which is different from what is generated on Earth due to limited variability in diet, human activity, and lifestyle in this confined area. Collection of actual wastewater influent from an isolated environment supporting humans is challenging. Additionally, to ensure a safe working environment in the laboratory and avoid the imposed threat of handling actual human feces, the proposed synthetic, non-human feces containing wastewater influent formulation offers an easy-to-produce and safer-to-handle option. This paper reviews several synthetic wastewater compositions that have been formulated for space exploration purposes. None of the formulations were found to be realistic nor adequate for a space-camp-type scenario. Thus, the formulation of a synthetic wastewater for simulating a wastewater influent from a human space-based camp is proposed in this paper. In addition, the physical, chemical, and biodegradation characteristics of the final formulation designed are presented to illustrate the value of the proposed influent formulation.

High-Quality Draft Genome Sequence of Fischerella thermalis JSC-11, a Siderophilic Cyanobacterium with Bioremediation Potential.

Here, we report the draft genome sequence of the siderophilic cyanobacterium Fischerella thermalis JSC-11, which was isolated from an iron-depositing hot spring. JSC-11 has bioremediation potential because it is capable of both extracellular absorption and intracellular mineralization of colloidal iron. This genomic information will facilitate the exploration of JSC-11 for bioremediation.

High-Quality Draft Genome Sequence of the Siderophilic and Thermophilic Leptolyngbyaceae Cyanobacterium JSC-12.

The siderophilic, thermophilic Leptolyngbyaceae cyanobacterium JSC-12 was isolated from a microbial mat in an iron-depositing hot spring. Here, we report the high-quality draft genome sequence of JSC-12, which may help elucidate the mechanisms of resistance to extreme iron concentrations in siderophilic cyanobacteria and lead to new remediation biotechnologies.

Draft Genome Sequence of the Putative Marine Pathogen Thalassobius sp. I31.1.

Thalassobius sp. I31.1 is a putative pathogen involved in epizootic shell disease in the American lobster (Homarus americanus). We report here the draft genome sequence for Thalassobius sp. I31.1 and provide insight into its metabolism and links to environmental pollutant degradation.

Draft Genome Sequence of the Putative Marine Pathogen Aquimarina sp. Strain I32.4.

Aquimarina sp. strain I32.4 (formerly Aquimarina sp. 'homaria') is a putative pathogen involved in epizootic shell disease in the American lobster (Homarus americanus). We report here the draft genome sequence for Aquimarina sp. strain I32.4 and describe virulence factors that may provide insight into its mechanism of pathogenicity.

Real-time PCR assay for Aquimarina macrocephali subsp. homaria and its distribution in shell disease lesions of Homarus americanus, Milne-Edwards, 1837, and environmental samples.

Epizootic shell disease (ESD) is causing major losses to the lobster fishery in southern New England. Potential pathogens have been identified in lesion communities, but there are currently no efficient means of detecting and quantifying their presence. A qPCR assay was developed for a key potential pathogen, Aquimarina macrocephali subsp. homaria found to be ubiquitous in ESD lesions but not the unaffected integument. Application of the assay to various samples demonstrated that A. macrocephali subsp. homaria is ubiquitous and abundant in lobster lesions, commonly associated with healthy surfaces of crabs and is scarce in water and sediment samples from southern New England suggesting the affinity of this microorganism to the Arthropod integument. The qPCR assay developed here can be applied in future in vivo and in vitro studies to better understand the ecology and role of A. macrocephali subsp.homaria. in shell disease.

Evaluation of thioesterases from Acinetobacter baylyi for production of free fatty acids.

Acinetobacter baylyi is one of few Gram-negative bacteria capable of accumulating storage lipids in the form of triacylglycerides and wax esters, which makes it an attractive candidate for production of lipophilic products, including biofuel precursors. Thioesterases play a significant dual role in the triacylglyceride and wax ester biosynthesis by either providing or removing acyl-CoA from this pathway. Therefore, 4 different thioesterase genes were cloned from Acinetobacter baylyi ADP1 and expressed in Escherichia coli to investigate their contribution to free fatty acids (FFAs) accumulation. Overexpression of the genes tesA' (a leaderless form of the gene tesA) and tesC resulted in increased accumulation of FFAs when compared with the host E. coli strain. Overexpression of tesA' showed a 1.87-fold increase in production of long-chain fatty acids (C16 to C18) over the host strain. Unlike TesC and the other investigated thioesterases, the TesA' thioesterase also produced shorter chain FFAs (e.g., myristic acid) and unsaturated FFAs (e.g., cis-vaccenic acid (18:1Δ11)). A comparison of the remaining 3 A. baylyi ADP1 thioesterases (encoded by the tesB, tesC, and tesD genes) revealed that only the strain containing the tesC gene produced statistically higher levels of FFAs over the control, suggesting that it possesses the acyl-ACP thioesterase activity. Both E. coli strains containing the tesB and tesD genes produced levels of FFAs similar to those of the plasmid-free control E. coli strain, which indicates that TesB and TesD lack the acyl-ACP thioesterase activity.

The Diversity of Sulfide Oxidation and Sulfate Reduction Genes Expressed by the Bacterial Communities of the Cariaco Basin, Venezuela.

Qualitative expression of dissimilative sulfite reductase (dsrA), a key gene in sulfate reduction, and sulfide:quinone oxidoreductase (sqr), a key gene in sulfide oxidation was investigated. Neither of the two could be amplified from mRNA retrieved with Niskin bottles but were amplified from mRNA retrieved by the Deep SID. The sqr and sqr-like genes retrieved from the Cariaco Basin were related to the sqr genes from a Bradyrhizobium sp., Methylomicrobium alcaliphilum, Sulfurovum sp. NBC37-1, Sulfurimonas autotrophica, Thiorhodospira sibirica and Chlorobium tepidum. The dsrA gene sequences obtained from the redoxcline of the Cariaco Basin belonged to chemoorganotrophic and chemoautotrophic sulfate and sulfur reducers belonging to the class Deltaproteobacteria (phylum Proteobacteria) and the order Clostridiales (phylum Firmicutes).

Possible shell disease in 100 million-year-old crabs.

Modern organisms exhibit evidence of many diseases, but recognizing such evidence in fossils remains difficult, thus hampering the study of the evolution of disease. We report on 2 molts of the goniodromitid crabs Distefania incerta and Goniodromites laevis from the mid-Cretaceous (late Albian) of Spain, with both species exhibiting damage to the dorsal carapace in otherwise well-preserved specimens. The subcircular to quadratical holes, found in <0.2% of the specimens, resemble damage caused by bacterial infections on the cuticle of modern decapods in terms of size and shape. Abiotic damage, predation, and encrustation followed by damage to the shell provide less satisfactory explanations, although these agents cannot be completely excluded from a role in shell disease etiology. We hypothesize that the observed fossil lesions are caused primarily by bacterial disease that started prior to molting, with or without other agents of initiation. If correct, this is the only known example of such bacterial infections in decapod crustaceans from the fossil record thus far, pushing back the evolutionary history of this type of shell disease by ~100 million years.

Glycerol Dehydratases: Biochemical Structures, Catalytic Mechanisms, and Industrial Applications in 1,3-Propanediol Production by Naturally Occurring and Genetically Engineered Bacterial Strains.

To date, two types of glycerol dehydratases have been reported: coenzyme B12-dependent and coenzyme B12-independent glycerol dehydratases. The three-dimensional structure of the former is a dimer of αßγ heterotrimer, while that of the latter is a homodimer. Their mechanisms of reaction are typically enzymatic radical catalysis. Functional radical in both the glycerol dehydratases is the adenosyl radical. However, the adenosyl radical in the former originates from coenzyme B12 by homolytic cleavage, and that in the latter from S-adenosyl-methionine. Until some years ago, Clostridium butyricum VPI 1718 was the only microorganism known to possess B12-independent glycerol dehydratase, but since then, several other bacteria with this unique capability have been identified. This article focuses on the glycerol dehydratases and on 1,3-propanediol production from glycerol by naturally occurring and genetically engineered bacterial strains containing glycerol dehydratase.

The dynamics of the bacterial diversity in the redox transition and anoxic zones of the Cariaco Basin assessed by parallel tag sequencing.

Massively parallel tag sequencing was applied to describe the bacterial diversity in the redox transition and anoxic zones of the Cariaco Basin. In total, 14 samples from the Cariaco Basin were collected over a period of eight years from two stations. A total of 244 357 unique bacterial V6 amplicons were sequenced. The total number of operational taxonomic units (OTUs) found in this study was 4692, with a range of 511-1491 OTUs per sample. Approximately 95% of the OTUs found in the redox transition zone and anoxic layers of Cariaco are represented by less than 50 amplicons suggesting that only about 5% of the bacterial OTUs are responsible for the bulk of the microbial processes in the basin redox transition and anoxic zones. The same dominant OTUs were observed across all eight years of sampling although periodic fluctuations in their proportion were apparent. No distinctive differences were observed between the bacterial communities from the redox transition and anoxic layers of the Cariaco Basin water column. The largest proportion of amplicons belongs to Gammaproteobacteria represented mostly by sulfide oxidizers, followed by Marine Group A (originally described as SAR406; Gordon and Giovannoni 1996), a group of uncultured bacteria hypothesized to be involved in metal reduction, and sulfate-reducing Deltaproteobacteria. Gammaproteobacteria, Deltaproteobacteria and Marine Group A make up 67-90% of all V6 amplicons sequenced in this study. This strongly suggests that the basin's microbial communities are actively involved in the sulfur-related metabolism and coupling of the sulfur and carbon cycles. According to detrended canonical correspondence analysis, ecological factors such as chemoautotrophy, nitrate and oxidized and reduced sulfur compounds influence the structuring and distribution of the Cariaco microbial communities.

Cuticles of European and American lobsters harbor diverse bacterial species and differ in disease susceptibility.

Diseases of lobster shells have a significant impact on fishing industries but the risk of disease transmission between different lobster species has yet to be properly investigated. This study compared bacterial biofilm communities from American (Homarus americanus) and European lobsters (H. gammarus), to assess both healthy cuticle and diseased cuticle during lesion formation. Culture-independent molecular techniques revealed diversity in the bacterial communities of cuticle biofilms both within and between the two lobster species, and identified three bacterial genera associated with shell lesions plus two putative beneficial bacterial species (detected exclusively in healthy cuticle or healing damaged cuticle). In an experimental aquarium shared between American and European lobsters, heterospecific transmission of potentially pathogenic bacteria appeared to be very limited; however, the claws of European lobsters were more likely to develop lesions when reared in the presence of American lobsters. Aquarium biofilms were also examined but revealed no candidate pathogens for environmental transmission. Aquimarina sp. 'homaria' (a potential pathogen associated with a severe epizootic form of shell disease) was detected at a much higher prevalence among American than European lobsters, but its presence correlated more with exacerbation of existing lesions rather than with lesion initiation.

Identification of bacteria in enrichment cultures of sulfate reducers in the Cariaco Basin water column employing Denaturing Gradient Gel Electrophoresis of 16S ribosomal RNA gene fragments.

BACKGROUND: The Cariaco Basin is characterized by pronounced and predictable vertical layering of microbial communities dominated by reduced sulfur species at and below the redox transition zone. Marine water samples were collected in May, 2005 and 2006, at the sampling stations A (10°30' N, 64°40' W), B (10°40' N, 64°45' W) and D (10°43'N, 64°32'W) from different depths, including surface, redox interface, and anoxic zones. In order to enrich for sulfate reducing bacteria (SRB), water samples were inoculated into anaerobic media amended with lactate or acetate as carbon source. To analyze the composition of enrichment cultures, we performed DNA extraction, PCR-DGGE, and sequencing of selected bands. RESULTS: DGGE results indicate that many bacterial genera were present that are associated with the sulfur cycle, including Desulfovibrio spp., as well as heterotrophs belonging to Vibrio, Enterobacter, Shewanella, Fusobacterium, Marinifilum, Mariniliabilia, and Spirochaeta. These bacterial populations are related to sulfur coupling and carbon cycles in an environment of variable redox conditions and oxygen availability. CONCLUSIONS: In our studies, we found an association of SRB-like Desulfovibrio with Vibrio species and other genera that have a previously defined relevant role in sulfur transformation and coupling of carbon and sulfur cycles in an environment where there are variable redox conditions and oxygen availability. This study provides new information about microbial species that were culturable on media for SRB at anaerobic conditions at several locations and water depths in the Cariaco Basin.

Bacterial communities associated with lesions of two forms of shell disease in the American lobster (Homarus americanus, Milne Edwards) from Atlantic Canada.

Shell disease is a major threat to the American lobster (Homarus americanus, Milne Edwards) fishery. Here we describe the composition of microbial communities associated with lesions of 2 forms of shell disease in Atlantic Canada, (i) a trauma shell disease (TSD) characterized by massive lesions and (ii) an enzootic shell disease (EnSD) characterized by irregularly shaped lesions with a distinct orange to yellow color. The microbiology of the lesions was described by polymerase chain reaction and denaturing gradient gel electrophoresis of 16S rDNA amplified from scrapings of the shell lesions and was compared with communities of unaffected carapaces and previously described forms of shell diseases. Both TSD and EnSD lesions were dominated by members of Alphaproteobacteria, Gammaproteobacteria, and Flavobacteria, all commonly detected in other forms of shell disease; however, unique members of Epsilonproteobacteria were also present. Two Vibrio spp. and 2 Pseudoalteromonas spp. were dominant in lesions of TSD and a Tenacibaculum sp. and Tenacibaculum ovolyticum were dominant in lesions of EnSD. The TSD and EnSD in this study contained similar taxa as other shell disease forms; however, their microbiology is mostly different and neither resembles that of epizootic shell disease.

Culture-independent analysis of bacterial communities in hemolymph of American lobsters with epizootic shell disease.

Epizootic shell disease (ESD) of the American lobster Homarus americanus H. Milne Edwards, 1837 is a disease of the carapace that presents grossly as large, melanized, irregularly shaped lesions, making the lobsters virtually unmarketable because of their grotesque appearance. We analyzed the bacterial communities present in the hemolymph of lobsters with and without ESD using nested-PCR of the 16S rRNA genes followed by denaturing gradient gel electrophoresis. All lobsters tested (n = 42) had bacterial communities in their hemolymph, and the community profiles were highly similar regardless of the sampling location or disease state. A number of bacteria were detected in a high proportion of samples and from numerous locations, including a Sediminibacterium sp. closely related to a symbiont of Tetraponera ants (38/42) and a Ralstonia sp. (27/42). Other bacteria commonly encountered included various Bacteroidetes, Pelomonas aquatica, and a Novosphingobium sp. One bacterium, a different Sediminibacterium sp., was detected in 20% of diseased animals (n = 29), but not in the lobsters without signs of ESD (n = 13). The bacteria in hemolymph were not the same as those known to be present in lesion communities except for the detection of a Thalassobius sp. in 1 individual. This work demonstrates that hemolymph bacteremia and the particular bacterial species present do not correlate with the incidence of ESD, providing further evidence that microbiologically, ESD is a strictly cuticular disease. Furthermore, the high incidence of the same species of bacteria in hemolymph of lobsters may indicate that they have a positive role in lobster fitness, rather than in disease, and further investigation of the role of bacteria in lobster hemolymph is required.

Bacterial community composition in a large marine anoxic basin: a Cariaco Basin time-series survey.

Redox transition zones play a crucial role in biogeochemical cycles of several major elements. Because microorganisms mediate many reactions of these cycles, they actively participate in establishing geochemical gradients. In turn, the geochemical gradients structure microbial communities. We studied the interrelationship between the bacterial community structure and the geochemical gradient in the Cariaco Basin, the largest truly marine anoxic basin. This study's dataset includes bacterial community composition in 113 water column samples as well as the data for environmental variables (gradients of oxygen, hydrogen sulfide, sulfite, thiosulfate, ammonia, nitrate, nitrite, dissolved manganese and iron, dark CO2 fixation, and bacterial abundance) collected between 1997 and 2006. Several prominent bacterial groups are present throughout the entire water column. These include members of Gamma-, Delta-, and Epsilonproteobacteria, as well as members of the Marine Group A, the candidate divisions OP11 and Car731c. Canonical correspondence analysis indicated that microbial communities segregate along vectors representing oxygenated conditions, nitrite, nitrate and anoxic environments represented by chemoautotrophy, ammonia, sulfite, and hydrogen sulfide.

Lesion bacterial communities in American lobsters with diet-induced shell disease.

In southern New England, USA, shell disease affects the profitability of the American lobster Homarus americanus fishery. In laboratory trials using juvenile lobsters, exclusive feeding of herring Clupea harengus induces shell disease typified initially by small melanized spots that progress into distinct lesions. Amongst a cohabitated, but segregated, cohort of 11 juvenile lobsters fed exclusively herring, bacterial communities colonizing spots and lesions were investigated by denaturing gradient gel electrophoresis of 16S rDNA amplified using 1 group-specific and 2 universal primer sets. The Bacteroidetes and Proteobacteria predominated in both spots and lesions and included members of the orders Flavobacteriales (Bacteriodetes), Rhodobacterales, Rhodospirillales and Rhizobiales (Alphaproteobacteria), Xanthomonadales (Gammaproteobacteria) and unclassified Gammaproteobacteria. Bacterial communities in spot lesions displayed more diversity than communities with larger (older) lesions, indicating that the lesion communities stabilize over time. At least 8 bacterial types persisted as lesions developed from spots. Aquimarina 'homaria', a species commonly cultured from lesions present on wild lobsters with epizootic shell disease, was found ubiquitously in spots and lesions, as was the 'Candidatus Kopriimonas aquarianus', implicating putative roles of these species in diet-induced shell disease of captive lobsters.

Denaturing gradient gel electrophoresis analyses of the vertical distribution and diversity of Vibrio spp. populations in the Cariaco Basin.

The Cariaco system is the second largest permanently anoxic marine water body in the world. Its water column is characterized by a pronounced vertical layering of microbial communities. The goal of our study was to investigate the vertical distribution and diversity of Vibrio spp. present in the Cariaco Basin waters using denaturing gradient gel electrophoresis (DGGE) of PCR-amplified 16S rDNA fragments. Representatives of the Vibrio genus were detected by nested and direct PCR in seawater at 10 depths. Sequence analyses of 55 DGGE bands revealed that only 11 different operational taxonomic units (OTU) are identified as Vibrio species. Between one and five OTUs were detected at each depth and the most common OTUs were OTU 1 and OTU 2, which phylogenetically clustered with Vibrio chagasii and Vibrio fortis, respectively. OTUs 3 and 4 were only found in the anoxic zone and were identified as Vibrio orientalis and Vibrio neptunius, respectively. Several Vibrio species detected are potentially pathogenic to human, prawns and corals such as Vibrio parahaemolyticus, Vibrio fischeri and Vibrio shilonii. In the Cariaco Basin, different Vibrio species were found to be specific to specific depths strata, suggesting that this genus is a natural component of the microbial communities in this marine redox environment.