Marla I. Tuffin

Molecular adaptations to psychrophily: the impact of ‘omic’ technologies

The ability of cold-adapted microorganisms (generally referred to as psychrophiles) to survive is the result of molecular evolution and adaptations which, together, counteract the potentially deleterious effects of low kinetic energy environments and the freezing of water. These physiological adaptations are seen at many levels. Against a background of detailed comparative protein structural analyses, the recent surge of psychrophile proteome, genome, metagenome and transcriptome sequence data has triggered a series of sophisticated analyses of changes in global protein composition. These studies have revealed consistent and statistically robust changes in amino acid composition, interpreted as evolutionary mechanisms designed to destabilise protein structures, as well as identifying the presence of novel genes involved in cold adaptation.

Extremophiles in biofuel synthesis

The current global energy situation has demonstrated an urgent need for the development of alternative fuel sources to the continually diminishing fossil fuel reserves. Much research to address this issue focuses on the development of financially viable technologies for the production of biofuels. The current market for biofuels, defined as fuel products obtained from organic substrates, is dominated by bioethanol, biodiesel, biobutanol and biogas, relying on the use of substrates such as sugars, starch and oil crops, agricultural and animal wastes, and lignocellulosic biomass. This conversion from biomass to biofuel through microbial catalysis has gained much momentum as biotechnology has evolved to its current status. Extremophiles are a robust group of organisms producing stable enzymes, which are often capable of tolerating changes in environmental conditions such as pH and temperature. The potential application of such organisms and their enzymes in biotechnology is enormous, and a particular application is in biofuel production. In this review an overview of the different biofuels is given, covering those already produced commercially as well as those under development. The past and present trends in biofuel production are discussed, and future prospects for the industry are highlighted. The focus is on the current and future application of extremophilic organisms and enzymes in technologies to develop and improve the biotechnological production of biofuels.

Abiotic factors influence microbial diversity in permanently cold soil horizons of a maritime-associated Antarctic Dry Valley

The McMurdo Dry Valleys collectively comprise the most extensive ice-free region in Antarctica and are considered one of the coldest arid environments on Earth. In low-altitude maritime-associated valleys, mineral soil profiles show distinct horizontal structuring, with a surface arid zone overlying a moist and biologically active zone generated by seasonally melted permafrost. In this study, long-term microenvironmental monitoring data show that temperature and soil humidity regimes vary in the soil horizons of north- and south-facing slopes within the Miers Valley, a maritime valley in the McMurdo Dry Valleys. We found that soil bacterial communities varied from the north to the south. The microbial assemblages at the surface and shallow subsurface depths displayed higher metabolic activity and diversity compared to the permafrost soil interface. Multivariate analysis indicated that K, C, Ca and moisture influenced the distribution and structure of microbial populations. Furthermore, because of the large % RH gradient between the frozen subsurface and the soil surface we propose that water transported to the surface as water vapour is available to microbial populations, either as a result of condensation processes or by direct adsorption from the vapour phase.

High-level diversity of tailed phages, eukaryote-associated viruses,and virophage-like elements in the metaviromes of Antarctic soils

ABSTRACT The metaviromes of two distinct Antarctic hyperarid desert soil communities have been characterized. Hypolithic communities, cyanobacterium-dominated assemblages situated on the ventral surfaces of quartz pebbles embedded in the desert pavement, showed higher virus diversity than surface soils, which correlated with previous bacterial community studies. Prokaryotic viruses (i.e., phages) represented the largest viral component (particularly Mycobacterium phages) in both habitats, with an identical hierarchical sequence abundance of families of tailed phages (Siphoviridae > Myoviridae > Podoviridae). No archaeal viruses were found. Unexpectedly, cyanophages were poorly represented in both metaviromes and were phylogenetically distant from currently characterized cyanophages. Putative phage genomes were assembled and showed a high level of unaffiliated genes, mostly from hypolithic viruses. Moreover, unusual gene arrangements in which eukaryotic and prokaryotic virus-derived genes were found within identical genome segments were observed. Phycodnaviridae and Mimiviridae viruses were the second-most-abundant taxa and more numerous within open soil. Novel virophage-like sequences (within the Sputnik clade) were identified. These findings highlight high-level virus diversity and novel species discovery potential within Antarctic hyperarid soils and may serve as a starting point for future studies targeting specific viral groups.

Metagenomic gene discovery: how far have we moved into novel sequence space?

Metagenomics emerged in the late 1990s as a tool for accessing and studying the collective microbial genetic material in the environment. The advent of the technology generated great excitement, as it has provided new opportunities and technologies for studying the wealth of microbial genetic diversity in the environment. Metagenomics has been widely predicted to access new dimensions of protein sequence space. A decade on, we review how far we have actually moved into new sequence space (and other aspects of protein space) using metagenomic tools. While several novel enzyme activities and protein structures have been identified through metagenomic strategies, the greatest advancement has been made in the isolation of novel protein sequences, some of which have no close relatives, form deeply branched lineages and even represent novel families. This is particularly true for glycosyl hydrolases and lipase/esterases, despite the fact that these activities are frequently screened for in metagenomic studies. However, there is much room for improvement in the methods employed and they will need to be addressed so that access to novel biocatalytic activities can be widened.

Hypolithic microbial communities of quartz rocks from Miers Valley, McMurdo Dry Valleys, Antarctica

The McMurdo Dry Valleys region of eastern Antarctica is a cold desert that presents extreme challenges to life. Hypolithic microbial colonisation of the subsoil surfaces of translucent quartz rocks represent a significant source of terrestrial biomass and productivity in this region. Previous studies have described hypoliths as dominated by cyanobacteria. However, hypoliths that occur in the lower Dry Valleys such as the Miers, Garwood and Marshall Valleys are unusual as they are not necessarily cyanobacteria-dominated. These hypoliths support significant eukaryal colonisation by fungi and mosses in addition to cyanobacteria-dominated bacterial assemblages and so have considerable ecological value in this barren landscape. Here, we characterise these novel hypoliths by analysis of environmental rRNA gene sequences. The hypolithic community was demonstrated to be distinct from the surrounding soil and non-translucent rocks. Hypoliths supported cyanobacterial signatures from the Oscillatoriales and Nostocales. Other heterotrophic bacterial signatures were also recovered, and these were phylogenetically diverse and spanned 8 other bacterial phyla. Archaeal phylotypes recovered were phylogenetically affiliated with the large group of unclassified, uncultured Crenarcheota. Eukaryal phylotypes indicated that free-living ascomycetous fungi, chlorophytes and mosses (Bryum sp.) were all supported by these hypoliths, and these are thought to be responsible for the extensive eukaryotic biomass that develops around quartz rocks.

Metagenomic analysis of the viral community in Namib Desert hypoliths

Hypolithic microbial communities are specialized desert communities inhabiting the underside of translucent rocks. Here, we present the first study of the viral fraction of these communities isolated from the hyperarid Namib Desert. The taxonomic composition of the hypolithic viral communities was investigated and a functional assessment of the sequences determined. Phylotypic analysis showed that bacteriophages belonging to the order Caudovirales, in particular the family Siphoviridae, were most prevalent. Functional analysis and comparison with other metaviromes revealed a relatively high frequency of cell wall-degrading enzymes, ribonucleotide reductases (RNRs) and phage-associated genes. Phylogenetic analyses of terL and phoH marker genes indicated that many of the sequences were novel and distinct from known isolates, and the class distribution of the RNRs suggests that this is a novel environment. The composition of the viral hypolith fraction containing many Bacillus-infecting phages was not completely consistent with Namib hypolith phylotypic surveys of the bacterial hosts, in which the cyanobacterial genus Chroococcidiopsis was found to be dominant. This could be attributed to the lack of sequence information about hypolith viruses/bacteria in public databases or the possibility that hypolithic communities incorporate viruses from the surrounding soil.

Microbial responses to solvent and alcohol stress.

Increasing fuel prices and doubts over the long‐term availability of oil are currently major global concerns. Such concerns have led to national policies and objectives to develop microbially produced alcohols as fuel additives or substitutes. However, in South Africa this solution poses the further dilemma of sourcing a suitable fermentative carbohydrate that will not impact negatively on the availability of staple foods. The solution lies in the use of lignocellulosic materials, currently a waste product of the food and agriculture industries, which could be used in conjunction with a catabolically suitable production strain. In the pursuit of lignocellulosic biofuel production, conventional fermentation strains have been shown to have limited catabolic versatility. However, catabolically versatile engineered strains and novel isolates engineered with ethanologenic pathways have subsequently been shown to exhibit limitations in solvent tolerance, hindering their full potential as economically viable production strains. A considerable volume of research has been reported on the general cellular mechanisms and physiological responses to solvent shock as well as adaptive changes responsible for solvent tolerant phenotypes in mutant progeny. Here we review a number of the more common cell responses to solvents with particular focus on alcohol tolerance.

Evidence of variability in the structure and recruitment of rhizospheric and endophytic bacterial communities associated with arable sweet sorghum (Sorghum bicolor (L) Moench)

Background and aimsSorghum is the second most cultivated crop in Africa and is a staple food source in many African communities. Exploiting the associated plant growth-promoting bacteria (PGPB) has potential as an agricultural biotechnology strategy to enhance sorghum growth, yield and nutritional properties. Therefore this study aimed to evaluate factors that shape bacterial communities associated with sorghum farmed in South Africa, and to detect bacteria consistently associated with sorghum which may impart PGP activities.MethodsTerminal-Restriction Fragment Length Polymorphism (T-RFLP) was used to assess factors that potentially shape rhizospheric (rhizosphere and rhizoplane) and endophytic (root, shoot, stem) bacterial communities associated with South African sorghum, and together with Denaturing Gradient Gel Electrophoresis (DGGE) to identify consistently sorghum-associated bacterial taxa.ResultsThe sorghum rhizospheric communities were less variable than the endophytic ones. Geographical location was the main driver in describing bacterial community assemblages found in rhizospheric sorghum-linked niches, with total NO3-N, NH4-N, nitrogen, carbon, pH and, to a lesser extent, clay content identified as the main abiotic factors shaping sorghum-associated soil communities. Endophytic communities presented rather stochastic assemblages, with pH being the main variable explaining their structures. Despite community variations, specific bacterial taxa were consistently detected in sorghum-created rhizospheric and endophytic environments, irrespective of environmental factor effects.ConclusionsSoil structure and composition, which are influenced by agricultural practices, played major roles in shaping sorghum-associated edaphic bacterial communities. In contrast, endophytic bacterial communities displayed more variation. Nevertheless, potentially agronomically relevant (cyano)bacterial taxa constantly associated with sorghum were identified which is suggestive of their deterministic recruitment.

Assessment of temporal and spatial evolution of bacterial communities in a biological sand filter mesocosm treating winery wastewater

To assess the impact of winery wastewater (WW) on biological sand filter (BSF) bacterial community structures, and to evaluate whether BSFs can constitute alternative and valuable treatment‐ processes to remediate WW.