Modelling suggests that the UV radiation environment of the early Earth, with DNA weighted irradiances of about three orders of magnitude greater than those at present, was hostile to life forms at the surface, unless they lived in specific protected habitats. However, we present empirical evidence that challenges this commonly held view. We describe a well-developed microbial mat that formed on the surface of volcanic littoral sediments in an evaporitic environment in a 3.5–3.3 Ga-old formation from the Barberton greenstone belt. Using a multiscale, multidisciplinary approach designed to strongly test the biogenicity of potential microbial structures, we show that the mat was constructed under flowing water by 0.25 μm filaments that produced copious quantities of extracellular polymeric substances, representing probably anoxygenic photosynthesizers. Associated with the mat is a small colony of rods–vibroids that probably represent sulphur-reducing bacteria. An embedded suite of evaporite minerals and desiccation cracks in the surface of the mat demonstrates that it was periodically exposed to the air in an evaporitic environment. We conclude that DNA-damaging UV radiation fluxes at the surface of the Earth at this period must either have been low (absorbed by CO2, H2O, a thin organic haze from photo-dissociated CH4, or SO2 from volcanic outgassing; scattered by volcanic, and periodically, meteorite dust, as well as by the upper layers of the microbial mat) and/or that the micro-organisms exhibited efficient gene repair/survival strategies.
Early Mid Archaean; Barberton; microfossils; littoral zone; UV environment
An unusually thick (∼1 cm) slime developed on a slump of finely disseminated pyrite ore within an extreme acid mine drainage site at Iron Mountain, near Redding, Calif. The slime was studied over the period of 1 year. The subaerial form of the slime distinguished it from more typical submerged streamers. Phylogenetic analysis of 16S rRNA genes revealed a diversity of sequences that were mostly novel. Nearest relatives to the majority of sequences came from iron-oxidizing acidophiles, and it appears that iron oxidation is the predominant metabolic characteristic of the organisms in the slime. The most abundant of the 16S rRNA genes detected were from organisms related to Leptospirillum species. The dominant sequence (71% of clones) may represent a new genus. Sequences within the Archaea of the Thermoplasmales lineage were detected. Most of these were only distantly related to known microorganisms. Also, sequences affiliating with Acidimicrobium were detected. Some of these were closely related to “Ferromicrobium acidophilus,” and others were affiliated with a lineage only represented by environmental clones. Unexpectedly, sequences that affiliated within the delta subdivision of the Proteobacteria were detected. The predominant metabolic feature of bacteria of this subdivision is anaerobic sulfate or metal reduction. Thus, microenvironments of low redox potential possibly exist in the predominantly oxidizing environments of the slime. These results expand our knowledge of the biodiversity of acid mine drainage environments and extend our understanding of the ecology of extremely acidic systems.
In West Africa, the Northern Sahelian zone and the coastal areas are densely populated but the Middle Belt in between is in general sparsely settled. Predictions of climate change foresee more frequent drought in the north and more frequent flooding in the coastal areas, while conditions in the Middle Belt will remain moderate. Consequently, the Middle Belt might become a major area for immigration but there may be constraining factors as well, particularly with respect to water availability. As a case study, the paper looks into the capacity of the Middle Belt zone of Benin, known as the Oueme River Basin (ORB), to reduce diarrhea prevalence. In Benin it links to the Millennium Development Goals on child mortality and environmental sustainability that are currently farthest from realization. However, diarrhea prevalence is only in part due to lack of availability of drinking water from a safe source. Social factors such as hygienic practices and poor sanitation are also at play. Furthermore, we consider these factors to possess the properties of a local public good that suffers from under provision and requires collective action, as individual actions to prevent illness are bound to fail as long as others free ride.
Combining data from the Demographic Health Survey with various spatial data sets for Benin, we apply mixed effect logit regression to arrive at a spatially explicit assessment of geographical and social determinants of diarrhea prevalence. Starting from an analysis of these factors separately at national level, we identify relevant proxies at household level, estimate a function with geo-referenced independent variables and apply it to evaluate the costs and impacts of improving access to good water in the basin.
First, the study confirms the well established stylized fact on the causes of diarrhea that a household with access to clean water and with good hygienic practices will, irrespective of other conditions, not suffer diarrhea very often. Second, our endogeneity tests show that joint estimation performs better than an instrumental variable regression. Third, our model is stable with respect to its functional form, as competing specifications could not achieve better performance in overall likelihood or significance of parameters. Fourth, it finds that the richer and better educated segments of the population suffer much less from the disease and apparently can secure safe water for their households, irrespective of where they live. Fifth, regarding geographical causes, it indicates that diarrhea prevalence varies with groundwater availability and quality across Benin. Finally, our assessment of costs and benefits reveals that improving physical access to safe water is not expensive but can only marginally improve the overall health situation of the basin, unless the necessary complementary measures are taken in the social sphere.
The ORB provides adequate water resources to accommodate future settlers but it lacks appropriate infrastructure to deliver safe water to households. Moreover, hygienic practices are often deficient. Therefore, a multifaceted approach is needed that acknowledges the public good aspects of health situation and consequently combines collective action with investments into water sources with improved management of public wells and further educational efforts to change hygienic practices.
Earth's climate during the Archaean remains highly uncertain, as the relevant geologic evidence is sparse and occasionally contradictory. Oxygen isotopes in cherts suggest that between 3.5 and 3.2 Gyr ago (Ga) the Archaean climate was hot (55–85 °C); however, the fact that these cherts have experienced only a modest amount of weathering suggests that the climate was temperate, as today. The presence of diamictites in the Pongola Supergroup and the Witwatersrand Basin of South Africa suggests that by 2.9 Ga the climate was glacial. The Late Archaean was relatively warm; then glaciation (possibly of global extent) reappeared in the Early Palaeoproterozoic, around 2.3–2.4 Ga.
Fitting these climatic constraints with a model requires high concentrations of atmospheric CO2 or CH4, or both. Solar luminosity was 20–25% lower than today, so elevated greenhouse gas concentrations were needed just to keep the mean surface temperature above freezing. A rise in O2 at approximately 2.4 Ga, and a concomitant decrease in CH4, provides a natural explanation for the Palaeoproterozoic glaciations. The Mid-Archaean glaciations may have been caused by a drawdown in H2 and CH4 caused by the origin of bacterial sulphate reduction. More work is needed to test this latter hypothesis.
palaeoclimate; Precambrian glaciations; methane; greenhouse effect; faint young Sun problem; sulphur isotopes
The respirable particulate matter (RPM; PM10) and total suspended particulate matter (TSP) concentrations in ambient air in Tuticorin, India, were preliminarily estimated. Statistical analyses on so-generated database were performed to infer frequency distributions and to identify dominant meteorological factor affecting the pollution levels. Both the RPM and TSP levels were well below the permissible limits set by the US Environmental Protection Agency. As expected, lognormal distribution always fit the data during the study period. However, fit with the normal was also acceptable except for very few seasons. The RPM concentrations ranged between 20.9 and 198.2 μg/m3, while the TSP concentrations varied from 51.5 to 333.3 μg/m3 during the study period. There was a better correlation between PM10–100 and TSP concentrations than that of PM10 (RPM) and TSP concentrations, but the correlation of RPM fraction was also acceptable. It was found that wind speed was the most important meteorological factor affecting the concentrations of the pollutants of present interest. Significant seasonal variations in the pollutant concentrations of present interest were found at 5% significance level except for TSP concentrations in the year 2006.
ANOVA; Coal-fired power station; Correlation; Frequency distribution; RPM; Seasonal variation; TSP
Objective: To examine whether a program to increase the wearing of seat belts in a South African urban area would be worthwhile in societal terms.
Design: A cost benefit analysis of a one year enhanced seat belt enforcement program in eThekwini (Durban) Municipality.
Methods: Data were drawn from two main sources—a 1998 study of the cost of road crashes in South Africa and, given the absence of other data, a meta-analysis of the effectiveness of various types of interventions to reduce road crash casualties in the United States—and were analyzed using cost benefit analysis.
Results: A program designed to enforce greater wearing of seat belts, estimated to cost 2 million rand in one year, could be reasonably expected to increase seat belt usage rates by 16 percentage points and reduce fatalities and injuries by 9.5%. This would result in saved social costs of 13.6 million rand in the following year or a net present value of 11.6 million rand. There would also be favorable consequences for municipal finances.
Conclusions: Investment in a program to increase seat belt wearing rates is highly profitable in societal terms.
Before the advent of oxygenic photosynthesis, the biosphere was driven by anaerobic metabolisms. We catalogue and quantify the source strengths of the most probable electron donors and electron acceptors that would have been available to fuel early-Earth ecosystems. The most active ecosystems were probably driven by the cycling of H2 and Fe2+ through primary production conducted by anoxygenic phototrophs. Interesting and dynamic ecosystems would have also been driven by the microbial cycling of sulphur and nitrogen species, but their activity levels were probably not so great. Despite the diversity of potential early ecosystems, rates of primary production in the early-Earth anaerobic biosphere were probably well below those rates observed in the marine environment. We shift our attention to the Earth environment at 3.8 Gyr ago, where the earliest marine sediments are preserved. We calculate, consistent with the carbon isotope record and other considerations of the carbon cycle, that marine rates of primary production at this time were probably an order of magnitude (or more) less than today. We conclude that the flux of reduced species to the Earth surface at this time may have been sufficient to drive anaerobic ecosystems of sufficient activity to be consistent with the carbon isotope record. Conversely, an ecosystem based on oxygenic photosynthesis was also possible with complete removal of the oxygen by reaction with reduced species from the mantle.
Archaean; evolution; hydrogen; anoxygenic photosynthesis; iron; metabolism
Earth is the one known example of an inhabited planet and to current knowledge the likeliest site of the one known origin of life. Here we discuss the origin of Earth’s atmosphere and ocean and some of the environmental conditions of the early Earth as they may relate to the origin of life. A key punctuating event in the narrative is the Moon-forming impact, partly because it made Earth for a short time absolutely uninhabitable, and partly because it sets the boundary conditions for Earth’s subsequent evolution. If life began on Earth, as opposed to having migrated here, it would have done so after the Moon-forming impact. What took place before the Moon formed determined the bulk properties of the Earth and probably determined the overall compositions and sizes of its atmospheres and oceans. What took place afterward animated these materials. One interesting consequence of the Moon-forming impact is that the mantle is devolatized, so that the volatiles subsequently fell out in a kind of condensation sequence. This ensures that the volatiles were concentrated toward the surface so that, for example, the oceans were likely salty from the start. We also point out that an atmosphere generated by impact degassing would tend to have a composition reflective of the impacting bodies (rather than the mantle), and these are almost without exception strongly reducing and volatile-rich. A consequence is that, although CO- or methane-rich atmospheres are not necessarily stable as steady states, they are quite likely to have existed as long-lived transients, many times. With CO comes abundant chemical energy in a metastable package, and with methane comes hydrogen cyanide and ammonia as important albeit less abundant gases.
Life probably arose on Earth after the moon-forming impact. It and subsequent impacts probably created transient reducing methane- or CO-rich atmospheres that provided abundant chemical energy.
The internalization of signaling receptors such as the insulin receptor is a complex, multi-step process. The aim of the present work was to determine the various steps in internalization of the insulin receptor and to establish which receptor domains are implicated in each of these by the use of receptors possessing in vitro mutations. We find that kinase activation and autophosphorylation of all three regulatory tyrosines 1146, 1150, and 1151, but not tyrosines 1316 and 1322 in the COOH-terminal domain, are required for the ligand-specific stage of the internalization process; i.e., the surface redistribution of the receptor from microvilli where initial binding occurs to the nonvillous domain of the cell. Early intracellular steps in insulin signal transduction involving the activation of phosphatidylinositol 3'-kinase are not required for this redistribution. The second step of internalization consists in the anchoring of the receptors in clathrin- coated pits. In contrast to the first ligand specific step, this step is common to many receptors including those for transport proteins and occurs in the absence of kinase activation and receptor autophosphorylation, but requires a juxta-membrane cytoplasmic segment of the beta-subunit of the receptor including a NPXY sequence. Thus, there are two independent mechanisms controlling insulin receptor internalization which depend on different domains of the beta-subunit.
In the title compound, [Co(C5HF6O2)2(C9H8N2)], the CoII centre exhibits a pseudooctahedral coordination geometry, comprising two N-atom donors from the bidentate aminoquinoline ligand and four O-atom donor atoms from two bidentate chelating 1,1,1,5,5,5-hexafluoropentane-2,4-dionate ligands. In the crystal, molecules are linked via pairs of N—H⋯O hydrogen bonds, forming inversion dimers. These dimers are further connected through π–π interactions between neighbouring quinoline rings [centroid–centroid distance = 3.472 (2) Å], and stack along the c axis.
Equatorial Africa has among the highest incidences of Kaposi’s sarcoma (KS) in the world, thus earning the name “KS Belt.” This was the case even prior to the HIV . To date, there is no clear evidence that HHV-8 seroprevalence is higher in this region, but interpretation of the available literature is tempered by differences in serologic assays used across studies. We examined representatively sampled ambulatory adults in Uganda, which is in the “KS Belt”, and in Zimbabwe and South Africa which are outside the Belt, for HHV-8 antibodies. All serologic assays were uniformly performed in the same reference laboratory by the same personnel. In the base-case serologic algorithm, seropositivity was defined by reactivity in an immunofluorescence assay or in two enzyme immunoassays. A total of 2375 participants were examined. In Uganda, HHV-8 seroprevalence was high early in adulthood (35.5% by age 21) without significant change thereafter. In contrast, HHV-8 seroprevalence early in adulthood was lower in Zimbabwe and South Africa (13.7% and 10.8%, respectively), but increased with age. After age adjustment, Ugandans had 3.24-fold greater odds of being HHV-8-infected than South Africans (p<0.001) and 2.22-fold greater odds than Zimbabweans (p<0.001). Inferences were unchanged using all other serologic algorithms evaluated. In conclusion, HHV-8 infection is substantially more common in Uganda than in Zimbabwe and South Africa. These findings help explain the high KS incidence in the “KS Belt” and underscore the importance of a uniform approach to HHV-8 antibody testing.
The ability to penetrate the dermal basement membrane and subsequently proliferate in the underlying mesenchyme is one of the key steps in malignant progression of human melanomas. We previously undertook studies aimed at assessing how normal dermal fibroblasts (one of the main cellular components of mesenchyme) may affect the growth of human melanoma cells and facilitate the overgrowth of malignant subpopulations (Cornil, I., D. Theodorescu, S. Man, M. Herlyn, J. Jambrosic, and R. S. Kerbel. 1991. Proc. Natl. Acad. Sci. USA. 88:6028- 6032). We found that melanoma cell lines from early-stage (metastatically incompetent) lesions were growth inhibited whereas those from advanced-stage (metastatically competent) lesions were stimulated under the same conditions by co-culture with fibroblasts; conditioned medium from such cells gave the same result. Subsequent studies using biochemical purification and neutralizing antibodies revealed the inhibitory activity to be identical to interleukin-6 (IL- 6). We now report that addition of purified recombinant human IL-6 resulted in a growth inhibition in vitro by G1/G0 arrest of early, but not advanced stage melanoma cells. Despite this alteration in response there was no significant difference in melanoma cell lines of varying malignancy in respect to their expression of genes encoding the IL-6 receptor, or gp130, the IL-6 signal transducer. Scatchard analysis also revealed similar [125I]IL-6 binding activities in both IL-6 sensitive and resistant groups. However, studies of IL-6 production indicated that five out eight IL-6 melanoma cell lines known to be resistant to exogenous IL-6-mediated growth inhibition constitutively expressed mRNA for IL-6; they also secreted bioactive IL-6 into culture medium. To assess the possible role of this endogenous IL-6 in melanoma cell growth, antisense oligonucleotides to the IL-6 gene were added to cultures of melanoma cells. This resulted in a significant growth inhibition only in cell lines that produced endogenous IL-6. In contrast, neutralizing antibodies to IL-6 were ineffective in causing such growth inhibition. This indicates that endogenous IL-6 may behave as a growth stimulator by an intracellular ("private") autocrine mechanism. Thus, a single cytokine, IL-6, can switch from behaving as a paracrine growth inhibitor to an autocrine growth stimulator within the same cell lineage during malignant tumor progression. Such a switch may contribute to the growth advantage of metastatically competent melanoma cells at the primary or distant organ sites and thereby facilitate progression of disease.
Environmental parameters drive phenotypic and genotypic frequency variations in microbial communities and thus control the extent and structure of microbial diversity. We tested the extent to which microbial community composition changes are controlled by shifting physiochemical properties within a hypersaline lagoon. We sequenced four sediment metagenomes from the Coorong, South Australia from samples which varied in salinity by 99 Practical Salinity Units (PSU), an order of magnitude in ammonia concentration and two orders of magnitude in microbial abundance. Despite the marked divergence in environmental parameters observed between samples, hierarchical clustering of taxonomic and metabolic profiles of these metagenomes showed striking similarity between the samples (>89%). Comparison of these profiles to those derived from a wide variety of publically available datasets demonstrated that the Coorong sediment metagenomes were similar to other sediment, soil, biofilm and microbial mat samples regardless of salinity (>85% similarity). Overall, clustering of solid substrate and water metagenomes into discrete similarity groups based on functional potential indicated that the dichotomy between water and solid matrices is a fundamental determinant of community microbial metabolism that is not masked by salinity, nutrient concentration or microbial abundance.
Polar and alpine microbial communities experience a variety of environmental stresses, including perennial cold and freezing; however, knowledge of genomic responses to such conditions is still rudimentary. We analyzed the metagenomes of cyanobacterial mats from Arctic and Antarctic ice shelves, using high-throughput pyrosequencing to test the hypotheses that consortia from these extreme polar habitats were similar in terms of major phyla and subphyla and consequently in their potential responses to environmental stresses. Statistical comparisons of the protein-coding genes showed similarities between the mats from the two poles, with the majority of genes derived from Proteobacteria and Cyanobacteria; however, the relative proportions differed, with cyanobacterial genes more prevalent in the Antarctic mat metagenome. Other differences included a higher representation of Actinobacteria and Alphaproteobacteria in the Arctic metagenomes, which may reflect the greater access to diasporas from both adjacent ice-free lands and the open ocean. Genes coding for functional responses to environmental stress (exopolysaccharides, cold shock proteins, and membrane modifications) were found in all of the metagenomes. However, in keeping with the greater exposure of the Arctic to long-range pollutants, sequences assigned to copper homeostasis genes were statistically (30%) more abundant in the Arctic samples. In contrast, more reads matching the sigma B genes were identified in the Antarctic mat, likely reflecting the more severe osmotic stress during freeze-up of the Antarctic ponds. This study underscores the presence of diverse mechanisms of adaptation to cold and other stresses in polar mats, consistent with the proportional representation of major bacterial groups.
Sulphur and carbon isotopic analyses on small samples of kerogens and sulphide minerals from biogenic and non-biogenic sediments of the 2.7 x 10(9) years(Ga)-old Belingwe Greenstone Belt (Zimbabwe) imply that a complex biological sulphur cycle was in operation. Sulphur isotopic compositions display a wider range of biological fractionation than hitherto reported from the Archaean. Carbon isotopic values in kerogen record fractionations characteristic of rubisco activity methanogenesis and methylotrophy and possibly anoxygenic photosynthesis. Carbon and sulphur isotopic fractionations have been interpreted in terms of metabolic processes in 2.7 Ga prokaryote mat communities, and indicate the operation of a diverse array of metabolic processes. The results are consistent with models of early molecular evolution derived from ribosomal RNA.
HIV-1 resistance data to inform treatment sequencing are limited for children with virological failure on first- and second-line antiretroviral therapy (ART) in Sub-Saharan Africa.
HIV-1-infected children aged ≤15 years were retrospectively identified from an ART cohort in Cape Town, South Africa (2003 to 2010). First-line ART was either non-nucleoside reverse transcriptase inhibitor (NNRTI) or lopinavir/ritonavir-based (with the exception of children <6 months old who received full-dose ritonavir as the sole protease inhibitor (PI) from 2004 to 2007). Second-line ART was the alternative regimen. Treatment outcomes, including virological failure, loss to care, death or remaining in care, were determined. Genotypic resistance testing was conducted on stored serum from children at first- or second-line virological failure (two consecutive HIV-1 RNA levels >1000 copies/ml). International AIDS Society criteria defined resistance mutations.
Of 472 children starting first-line ART, 352 (75%) remained in care, 45 (9%) were lost and 4 (1%) died on first-line treatment. Seventy-one (15%) had observed virological failure, and 37 of these children had specimens available for genotype testing. Eight children (22%) had wild-type virus, seven (19%) had thymidine analog mutations (TAMs), 24 (65%) had NNRTI resistance and two (5.4%) had multiple protease resistance (PR). Of the 78 children who received second-line ART, 54 (71%) remained in care, 6 (8%) were lost and 1 (1%) died during second-line treatment. Fifteen (20%) had observed virological failure; 13 had samples available for genotype. Three (23%) had wild-type virus, eight (62%) had TAMs, nine (69%) had NNRTI resistance, and five (38%) had multiple PI resistance all of whom had received full-dose ritonavir.
Although virological failure was infrequent in children on first- and second-line ART, rates of observed resistance including multiple PR resistance after failure were high. Reasons for high rates of resistance include use of full-dose ritonavir and continued viremia. Wild-type virus was common, suggesting poor adherence or challenges in correct dosing. Genotype resistance testing in children with virological failure may optimize selection of subsequent regimens and inform recommendations for sequencing of existing ART.
Polyamine analogues show antitumor activity in experimental models and their ability to alter activity of cytotoxic chemotherapeutic agents in breast cancer is well documented. Association of polyamines with nucleic acids and protein is included in their mechanism of action. The aim of this study was to examine the interaction of human serum albumin (HSA) with several polyamine analogues such as 1,11-diamino-4,8-diazaundecane (333), 3,7,11,15-tetrazaheptadecane.4HCl (BE-333) and 3,7,11,15,19-pentazahenicosane.5HCl (BE-3333) in aqueous solution at physiological conditions, using a constant protein concentration and various polyamine contents (μM to mM). FTIR, UV-visible and CD spectroscopic methods were used to determine the polyamine binding mode and the effects of polyamine complexation on protein stability and secondary structure.
Structural analysis showed that polyamines bind non-specifically (H-bonding) via polypeptide polar groups with binding constants of K333 = 9.30 × 103 M−1, KBE-333 = 5.63 × 102 M−1 and KBE-3333 = 3.66 × 102 M−1. The protein secondary structure showed major alterations with reduction of α-helix from 55% (free protein) to 43–50% and increase of β-sheet from 17% (free protein) to 29–36% in the 333-, BE-333- and BE-3333 complexes, indicating a partial protein unfolding upon polyamine interaction. HSA structure was less perturbed by polyamine analogues than those of the biogenic polyamines.
polyamine analogues; protein; HSA; binding mode; secondary structure; FTIR; CD spectroscopy
Toxoplasma gondii seroprevalence was determined in 3333 sheep sera from 125 distinct sheep flocks in Scotland, with the majority of flocks being represented by 27 samples, which were collected between July 2006 and August 2008. The selected farms give a representative sample of 14 400 sheep holdings identified in the Scottish Government census data from 2004. Overall T. gondii seroprevalence, at individual sheep level, was determined to be 56.6%; each flock tested, had at least a single positive animal and in four flocks all ewes tested positive. The seroprevalence of sheep increased from 37.7% in one year old stock to 73.8% in ewes that were older than six years, showing that acquired infections during the life of the animals is frequent and that environmental contamination by T. gondii oocysts must be significant. The median within-flock seroprevalence varied significantly across Scotland, with the lowest seroprevalence of 42.3% in the South and the highest seroprevalence of 69.2% in the far North of Scotland and the Scottish Islands, while the central part of Scotland had a seroprevalence of 57.7%. This distribution disequilibrium may be due to the spread and survival of oocysts on pasture and lambing areas. A questionnaire accompanying sampling of flocks identified farms that used Toxovax®, a commercial vaccine that protects sheep from abortion due to T. gondii infection. Only 24.7% of farmers used the vaccine and the vaccine did not significantly affect the within flock seroprevalence for T. gondii. The implications for food safety and human infection are discussed.
Stromatolites are laminated carbonate build-ups formed by the metabolic activity of microbial mats and represent one of the oldest known ecosystems on Earth. In this study, we examined a living stromatolite located within the Exuma Sound, The Bahamas and profiled the metagenome and metabolic potential underlying these complex microbial communities.
The metagenomes of the two dominant stromatolitic mat types, a nonlithifying (Type 1) and lithifying (Type 3) microbial mat, were partially sequenced and compared. This deep-sequencing approach was complemented by profiling the substrate utilization patterns of the mats using metabolic microarrays. Taxonomic assessment of the protein-encoding genes confirmed previous SSU rRNA analyses that bacteria dominate the metagenome of both mat types. Eukaryotes comprised less than 13% of the metagenomes and were rich in sequences associated with nematodes and heterotrophic protists. Comparative genomic analyses of the functional genes revealed extensive similarities in most of the subsystems between the nonlithifying and lithifying mat types. The one exception was an increase in the relative abundance of certain genes associated with carbohydrate metabolism in the lithifying Type 3 mats. Specifically, genes associated with the degradation of carbohydrates commonly found in exopolymeric substances, such as hexoses, deoxy- and acidic sugars were found. The genetic differences in carbohydrate metabolisms between the two mat types were confirmed using metabolic microarrays. Lithifying mats had a significant increase in diversity and utilization of carbon, nitrogen, phosphorus and sulfur substrates.
The two stromatolitic mat types retained similar microbial communities, functional diversity and many genetic components within their metagenomes. However, there were major differences detected in the activity and genetic pathways of organic carbon utilization. These differences provide a strong link between the metagenome and the physiology of the mats, as well as new insights into the biological processes associated with carbonate precipitation in modern marine stromatolites.
While the oceans cover more than 70% of the Earth’s surface, marine derived microbial natural products have been largely unexplored. The marine environment is a habitat for many unique microorganisms, which produce biologically active compounds (“bioactives”) to adapt to particular environmental conditions. For example, marine surface associated microorganisms have proven to be a rich source for novel bioactives because of the necessity to evolve allelochemicals capable of protecting the producer from the fierce competition that exists between microorganisms on the surfaces of marine eukaryotes. Chemically driven interactions are also important for the establishment of cross-relationships between microbes and their eukaryotic hosts, in which organisms producing antimicrobial compounds (“antimicrobials”), may protect the host surface against over colonisation in return for a nutrient rich environment. As is the case for bioactive discovery in general, progress in the detection and characterization of marine microbial bioactives has been limited by a number of obstacles, such as unsuitable culture conditions, laborious purification processes, and a lack of de-replication. However many of these limitations are now being overcome due to improved microbial cultivation techniques, microbial (meta-) genomic analysis and novel sensitive analytical tools for structural elucidation. Here we discuss how these technical advances, together with a better understanding of microbial and chemical ecology, will inevitably translate into an increase in the discovery and development of novel drugs from marine microbial sources in the future.
marine epibiotic microoorganisms; bioactive; antimicrobial; natural products
In this study, the composition of the metabolically active fraction of the microbial community occurring in Gulf of Mexico marine sediments (water depth, 550 to 575 m) with overlying filamentous bacterial mats was determined. The mats were mainly composed of either orange- or white-pigmented Beggiatoa spp. Complementary 16S ribosomal DNA (crDNA) was obtained from rRNA extracted from three different sediment depths (0 to 2, 6 to 8, and 10 to 12 cm) that had been subjected to reverse transcription-PCR amplification. Domain-specific 16S PCR primers were used to construct 12 different 16S crDNA libraries containing 333 Archaea and 329 Bacteria clones. Analysis of the Archaea clones indicated that all sediment depths associated with overlying orange- and white-pigmented microbial mats were almost exclusively dominated by ANME-2 (95% of total Archaea clones), a lineage related to the methanogenic order Methanosarcinales. In contrast, bacterial diversity was considerably higher, with the dominant phylotype varying by sediment depth. An equivalent number of clones detected at 0 to 2 cm, representing a total of 93%, were related to the γ and δ classes of Proteobacteria, whereas clones related to δ-Proteobacteria dominated the metabolically active fraction of the bacterial community occurring at 6 to 8 cm (79%) and 10 to 12 cm (85%). This is the first phylogenetics-based evaluation of the presumptive metabolically active fraction of the Bacteria and Archaea community structure investigated along a sediment depth profile in the northern Gulf of Mexico, a hydrocarbon-rich cold-seep region.
The main mechanism of fluoroquinolone (FQ) resistance in Mycobacterium tuberculosis is mutation in DNA gyrase (GyrA2GyrB2), especially in gyrA. However, the discovery of unknown mutations in gyrB whose implication in FQ resistance is unclear has become more frequent. We investigated the impact on FQ susceptibility of eight gyrB mutations in M. tuberculosis clinical strains, three of which were previously identified in an FQ-resistant strain. We measured FQ MICs and also DNA gyrase inhibition by FQs in order to clarify the role of these mutations in FQ resistance. Wild-type GyrA, wild-type GyrB, and mutant GyrB subunits produced from engineered gyrB alleles by mutagenesis were overexpressed in Escherichia coli, purified to homogeneity, and used to reconstitute highly active gyrase complexes. MICs and DNA gyrase inhibition were determined for moxifloxacin, gatifloxacin, ofloxacin, levofloxacin, and enoxacin. We demonstrated that the eight substitutions in GyrB (D473N, P478A, R485H, S486F, A506G, A547V, G551R, and G559A), recently identified in FQ-resistant clinical strains or encountered in M. tuberculosis strains isolated in France, are not implicated in FQ resistance. These results underline that, as opposed to phenotypic FQ susceptibility testing, the DNA gyrase inhibition assay is the only way to prove the role of a DNA gyrase mutation in FQ resistance. Therefore, the use of FQ in the treatment of tuberculosis (TB) patients should not be ruled out only on the basis of the presence of mutations in gyrB.
Biogenic polyamines are found to modulate protein synthesis at different levels, while polyamine analogues have shown major antitumor activity in multiple experimental models, including breast cancer. The aim of this study was to examine the interaction of bovine serum albumin (BSA) with biogenic polyamines, spermine and spermidine, and polyamine analogues 3,7,11,15-tetrazaheptadecane.4HCl (BE-333) and 3,7,11,15,19-pentazahenicosane.5HCl (BE-3333) in aqueous solution at physiological conditions. FTIR, UV-visible, CD and fluorescence spectroscopic methods were used to determine the polyamine binding mode and the effects of polyamine complexation on protein stability and secondary structure. Structural analysis showed that polyamines bind BSA via both hydrophilic and hydrophobic interactions. Stronger polyamine-protein complexes formed with biogenic than synthetic polyamines with overall binding constants of Kspm = 3.56 (±0.5) × 105 M-1, Kspmd = 1.77 (±0.4) × 105 M-1, KBE-333 = 1.11 (±0.3) × 104 M-1 and KBE-3333 = 3.90 (± 0.7) × 104 M-1 that correlate with their positively charged amino group contents. Major alterations of protein conformation were observed with reduction of α-helix from 63% (free protein) to 55-33% and increase of turn 12% (free protein) to 28-16% and random coil from 6% (free protein) to 24-17% in the polyamine-BSA complexes, indicating a partial protein unfolding. These data suggest that serum albumins might act as polyamine carrier proteins in delivering polyamine analogues to target tissues.
polyamine; BSA; binding mode; secondary structure; FTIR; CD; fluorescence spectroscopy
Smallpox was eradicated more than 10 years ago, but infection with another Orthopoxvirus, monkeypox virus, can result in a clinical picture resembling smallpox. Human infection with monkeypox virus is extremely rare, not easily transmitted, and confined to the rain forest belt of Africa (Z. Jezek and F. Fenner, p. 81-102, in Human Monkeypox, 1988). Evidence that variola virus, the causative agent of smallpox, might be readily derived from monkeypox virus was presented [S. S. Marennikova and E. M. Shelukhina, Nature (London) 276:291-292, 1978; S. S. Marennikova, E. M. Shelukhina, N. N. Maltseva, and G. R. Matsevich Intervirology 11:333-340, 1979], but this was not confirmed [K. R. Dumbell and L. C. Archard, Nature (London) 286:29-32, 1980] and was subsequently discounted (J. J. Esposito, J. H. Nakano, and J. F. Obijeski, Bull. W.H.O. 63:695-703, 1985). Although enough difference between the genomes of monkeypox and variola viruses to rule out a simple interconversion has been demonstrated [K. R. Dumbell and L. C. Archard, Nature (London) 286:29-32, 1980; J. J. Esposito and J. C. Knight, Virology 143:230-251, 1985; J. J. Esposito, J. H. Nakano, and J. F. Obijeski, Bull. W.H.O. 63:695-703, 1985; M. Mackett and L. C. Archard, J. Gen. Virol. 45:683-701, 1979], the possibility that monkeypox virus was a more remote ancestor of variola virus remained. We have now identified a sequence in monkeypox virus DNA which is a homolog of a 1,065-bp open reading frame in the conserved region of the variola virus genome but which has multiple deletions. This is strong evidence that monkeypox virus is not ancestral to variola virus and strengthens confidence in the long-term success of smallpox eradication.
Cyanobacterial mats are often a major biological component of extreme aquatic ecosystems, and in polar lakes and streams they may account for the dominant fraction of total ecosystem biomass and productivity. In this study we examined the vertical structure and physiology of Arctic microbial mats relative to the question of how these communities may respond to ongoing environmental change. The mats were sampled from Ward Hunt Lake (83°5.297′N, 74°9.985′W) at the northern coast of Arctic Canada, and were composed of three visibly distinct layers. Microsensor profiling showed that there were strong gradients in oxygen within each layer, with an overall decrease from 100% saturation at the mat surface to 0%, at the bottom, accompanied by an increase of 0.6 pH units down the profile. Gene clone libraries (16S rRNA) revealed the presence of Oscillatorian sequences throughout the mat, while Nostoc related species dominated the two upper layers, and Nostocales and Synechococcales sequences were common in the bottom layer. High performance liquid chromatography analyses showed a parallel gradient in pigments, from high concentrations of UV-screening scytonemin in the upper layer to increasing zeaxanthin and myxoxanthin in the bottom layer, and an overall shift from photoprotective to photosynthetic carotenoids down the profile. Climate change is likely to be accompanied by lake level fluctuations and evaporative concentration of salts, and thus increased osmotic stress of the littoral mat communities. To assess the cellular capacity to tolerate increasing osmolarity on physiology and cell membrane integrity, mat sections were exposed to a gradient of increasing salinities, and PAM measurements of in vivo chlorophyll fluorescence were made to assess changes in maximum quantum yield. The results showed that the mats were tolerant of up to a 46-fold increase in salinity. These features imply that cyanobacterial mats are resilient to ongoing climate change, and that in the absence of major biological perturbations, these vertically structured communities will continue to be a prominent feature of polar aquatic ecosystems.
microbial mats; cyanobacteria; polar lake; pigments; molecular phylogeny