Analysis Of Gene Diversity Research Articles

Analysis of Starch Gene Diversity in the Wild Relatives of Oryza sativa

Genetic loci influencing traits important to humans have been selected during crop domestication. The starch properties of rice influence the ease of cooking and attractiveness of rice as a human food. Starch biosynthesis genes likely to influence starch properties in the grain were compared in wild and domesticated rice genotypes. Sequence variation was investigated in starch biosynthesis gene exons that have been reported to have a direct influence on rice amylose content, gelatinization temperature, and amylopectin chain length. Exons 6 and 10 of GBSSI, exon 8 of SSIIa and exons 11, 13, 14 and 16 of SBEIIb were amplified and sequenced from 13 wild Oryza species encompassing genome types AA to HHJJ. Thirty two single nucleotide polymorphisms (SNPs) were identified in the exons of GBSSI; 176 in exon 8 of SSIIa, and 43 in SBEIIb, giving a total of 251 SNPs among the species. Eighty six of these SNP caused changes in the encoded amino acid, of which 28 were missense mutations that affected highly conserved amino acids within the protein sequence of GBSSI, SSIIa or SBEIIb. Two indels were identified in Potamophila parviflora, a close relative of Zizania palustris, a North American native wild rice. Most of the nucleotide variations and non-conservative changes were observed in the genomes other than the AA genome species. This represents a genetic resource for use in rice starch manipulation. The impact of human selection at these loci can be deduced by comparison of modern cultivated genotypes with their wild progenitors.

GeoChip-Based Analysis of Microbial Functional Gene Diversity in a Landfill Leachate-Contaminated Aquifer

The functional gene diversity and structure of microbial communities in a shallow landfill leachate-contaminated aquifer were assessed using a comprehensive functional gene array (GeoChip 3.0). Water samples were obtained from eight wells at the same aquifer depth immediately below a municipal landfill or along the predominant downgradient groundwater flowpath. Functional gene richness and diversity immediately below the landfill and the closest well were considerably lower than those in downgradient wells. Mantel tests and canonical correspondence analysis (CCA) suggested that various geochemical parameters had a significant impact on the subsurface microbial community structure. That is, leachate from the unlined landfill impacted the diversity, composition, structure, and functional potential of groundwater microbial communities as a function of groundwater pH, and concentrations of sulfate, ammonia, and dissolved organic carbon (DOC). Historical geochemical records indicate that all sampled wells chronically received leachate, and the increase in microbial diversity as a function of distance from the landfill is consistent with mitigation of the impact of leachate on the groundwater system by natural attenuation mechanisms.

Microbially Induced Iron Precipitation Associated with a Neutrophilic Spring at Borra Caves, Vishakhapatnam, India

The present investigation uncovers various pieces of evidence for the possible biologically induced mineralization in iron mats associated with a pH-neutral spring in the Borra caves, Vishakhapatnam, India. Electron microscopy [scanning electron microscopy (SEM) and transmission electron microscopy (TEM)] demonstrated large numbers of (i) hollow tubes (diameter ∼1 μm) resembling sheaths of the iron-oxidizing bacteria Leptothrix, (ii) thin (diameter <<1 μm) solid fibers of uncertain origin, (iii) nanoscale subspherical to irregularly shaped particles encrusting tubes and fibers, and (iv) aggregates of broken and partially disintegrated sheaths, fibers, and particles embedded in extracellular polymeric substances (EPS) occasionally including microbial cells. X-ray microanalyses by energy dispersive spectroscopy (EDS) revealed that the mat accumulated largely Fe but also smaller amounts of Si and traces of P and Ca. Particles rich in Si and Al (possibly kaolinite) and Ca (carbonate) were also observed. High-resolution TEM/EDS of unstained ultrathin sections suggests that microbial sheaths were highly mineralized by amorphous to cryptocrystalline Fe-rich phases and less frequently by other fine-grained and fibrous authigenic claylike minerals. Total number of microorganisms in the iron mats was 5.8×10(5) cells, g sed(-1) (wet weight). Analysis of the 16S rRNA gene diversity revealed microorganisms assigned to eight different phyla [Proteobacteria (62%), Chloroflexi (8%), Bacteroidetes (7%), Planctomycetes (1%), Actinobacteria (5%), Acidobacteria (6%), Nitrospira (1%), Firmicutes (5%)]. Within the Proteobacteria, Betaproteobacteria was the predominant class, which accounted for 28% of the sequences. Within this class some obvious similarities between the obtained sequences and sequences from other cave systems could be seen, especially sequences affiliated with Leptothrix, Siderooxidans, Crenothrix, Comamonadaceae, Dechloromonas, and many uncultured Betaproteobacteria. Four (4%) of the sequences could not be assigned to phylum level but were affiliating with the candidate division TM7 (2%), candidate division OP11 (1%), and candidate division WWE3 (1%). The results allow us to infer a possible relationship of microbial sheaths, EPS, and the iron precipitates to microbial community diversity in the Borra cave springs. Understanding biogenic iron oxides in caves has important astrobiological applications as it provides a potential tool for the detection of extraterrestrial life.

Cloning of SCA Gene Related to Pollen Tube Adhesion and Oriented Growth and Analysis of Gene Diversity in &lt;i&gt;Lilium spp.&lt;/i&gt;

Stigma/style cysteine-rich adhesin genes ( SCA ) were cloned from ‘Tresor’ ( Lilium Asiatic Hybrids), ‘White heaven’ ( Lilium Longiflorum Hybrids) and ‘Caruso’ ( Lilium Oriental Hybrids) leaves by PCR approaches. In this research we obtained four gene copies in both of ‘Tresor’ and ‘Caruse’ genomes and three gene copies in ‘White heaven’ genome. The identities of the cloned SCA among the copies were in the ranges from 72.97% to 99.68%. There are two exons and one intron in all of cloned SCA sequence, which could be deduced immature protein with 113 or 115 amino acid residues (encoding mature protein with 91 amino acid residues) SCA protein contains an N-terminal signal peptide of typical LTP protein, eight conserved cysteine residues and two consensus pentapeptide motifs. The identities among the amino acid sequences were in range from 87.91% to 98.90% higher than that of the similarities of gene sequences. Alignment analysis of amino acid sequences of cloned SCAs showed there were some distinct differences of amino acid site existing in the tested three species of lily hybrids. The phylogenetic tree revealed the relationship of the cloned 11 amino acid sequences of SCA were all more close to LTP family of monocotyledons than that of dicotyledons, which were consistent with the conclusions of kinship in plant taxonomy.

Longitudinal and Cross-Sectional Genetic Diversity in the Korean Peninsula Based on the P vivax Merozoite Surface Protein Gene

ObjectivesVivax malaria has reemerged and become endemic in Korea. Our study aimed to analyze by both longitudinal and cross-sectional genetic diversity of this malaria based on the P vivax Merozoite Surface Protein (PvMSP) gene parasites recently found in the Korean peninsula. MethodsPvMSP-1 gene sequence analysis from P vivax isolates (n = 835) during the 1996-2010 period were longitudinally analyzed and the isolates from the Korean peninsula through South Korea, the demilitarized zone and North Korea collected in 2008-2010 were enrolled in an overall analysis of MSP-1 gene diversity. ResultsNew recombinant subtypes and severe multiple-cloneinfection rates were observed in recent vivax parasites. Regional variation was also observed in the study sites. ConclusionThis study revealed the great complexity of genetic variation and rapid dissemination of genes in P vivax. It also showed interesting patterns of diversity depending, on the region in the Korean Peninsula. Understanding the parasiteninsula. Under genetic variation may help to analyze trends and assess the extent of endemic malaria in Korea.

Genetic variability and differentiation of Caragana microphylla populations as revealed by RAPD markers

Genetic variability in random amplified polymorphic DNA (RAPD) was studied in 90 individuals of Caragana microphylla, an outcrossing perennial shrub species, from five natural populations sampled in Inner Mongolia steppe of China on a small scale. Nineteen selected primers were used to amplify DNA samples, and totally 225 bands were detected. The percentage of polymorphic bands within populations ranged form 58.22% to 63.56%, with an average of 60% at the population level and 71.11% at the species level, indicating relatively high genetic variations in C. microphylla species. Shannon's information index (I) and Nei's gene diversity (h) showed the similar trend with each other. According to the analysis of Nei's gene diversity, the percentage of genetic variation among populations was 7.13%, indicating a low level of genetic differentiation among populations. There existed a strong gene flow (Nm = 3.26) among populations. Although AMOVA analysis also revealed most variation was within populations (phi(ST) = 4.1%), a significant proportion was observed among populations (P<0.001) in the present study, suggesting genetic differentiation occurred among populations at a certain extent. Based on Mantel's tests and the results of previous studies, the genetic structure pattern of C. microphylla accorded with the isolation-by-distance model on a very large scale, however, on a small scale, the significant genetic differentiation among populations might be enhanced by the micro-environmental divergence among the sampling sites, rather than by geographic factors. Analysis of the genetic variations of C. microphylla populations provided useful information for the adaptive strategy of Caragana species.

GeoChip-Based Analysis of the Functional Gene Diversity and Metabolic Potential of Microbial Communities in Acid Mine Drainage

Acid mine drainage (AMD) is an extreme environment, usually with low pH and high concentrations of metals. Although the phylogenetic diversity of AMD microbial communities has been examined extensively, little is known about their functional gene diversity and metabolic potential. In this study, a comprehensive functional gene array (GeoChip 2.0) was used to analyze the functional diversity, composition, structure, and metabolic potential of AMD microbial communities from three copper mines in China. GeoChip data indicated that these microbial communities were functionally diverse as measured by the number of genes detected, gene overlapping, unique genes, and various diversity indices. Almost all key functional gene categories targeted by GeoChip 2.0 were detected in the AMD microbial communities, including carbon fixation, carbon degradation, methane generation, nitrogen fixation, nitrification, denitrification, ammonification, nitrogen reduction, sulfur metabolism, metal resistance, and organic contaminant degradation, which suggested that the functional gene diversity was higher than was previously thought. Mantel test results indicated that AMD microbial communities are shaped largely by surrounding environmental factors (e.g., S, Mg, and Cu). Functional genes (e.g., narG and norB) and several key functional processes (e.g., methane generation, ammonification, denitrification, sulfite reduction, and organic contaminant degradation) were significantly (P < 0.10) correlated with environmental variables. This study presents an overview of functional gene diversity and the structure of AMD microbial communities and also provides insights into our understanding of metabolic potential in AMD ecosystems.

Analysis of Malus S-RNase gene diversity based on a comparative study of old and modern apple cultivars and European wild apple

Malus S-RNase genetic diversity was analyzed in Malus × domestica cultivars and compared to European wild apple (Malus sylvestris). Using PCR-based approaches, the S-RNase genotype of 140 M. × domestica cultivars, 196 M. sylvestris trees and 27 M. sylvestris—M. × domestica hybrids was determined. S-RNase allelic richness in M. sylvestris was much higher than in M. × domestica, indicating the negative influence of domestication on S-RNase diversity. Heterogeneity of the S-RNase allelic distribution is much higher in cultivated apple than in wild apple, which shows that breeding leads to strong departure from the expected homogeneity of genes under negative frequency-dependent selection. The majority of the M. × domesticaS-alleles has been found in M. sylvestris as well, which points to strong conservation of the S-locus gene structure. Based on the sequence of all different SCAR-fragments, which comprise both the hypervariable PS1 region and the single intron, S-RNase genetic diversity was further explored. It provided some clues to the occurrence of new S-alleles among the multitude of novel S-RNase sequences that have been identified, which were mostly unique for the group of M. sylvestris individuals. The determination of the S-RNase genotypes of old cultivars and M. sylvestris will enable their introduction into new breeding strategies. As M. sylvestris has become an endangered species in Belgium, the knowledge gathered in this study will be an important tool for selecting useful genotypes for a core collection.

Microsatellite-based genetic diversity and differentiation of foreign common carp ( Cyprinus carpio) strains farmed in the Czech Republic

This study set out to improve our understanding of the molecular genetic characteristics of imported strains of common carp reared in the Czech Republic. To do this we characterized the genetic diversity and population structure of 11 carp strains including two farmed strains from Germany (Scheuerman and Glinzig mirror carp) and France (Forez and Dombez scaly carp), one farmed population from Amur river basins, and one feral population from the Ebro River (Spain), by microsatellite analysis of 10 loci. Mean heterozygosity within populations ranged from 0.584 to 0.700, and the mean number of alleles per population ranged from 5.0 to 9.8. Of the analysed loci, 92 out of 130 possible tests were found to deviate significantly ( P < 0.05) from the Hardy–Weinberg equilibrium revealing significant heterozygote deficit after Bonferroni correction. Global analysis of hierarchical gene diversity (AMOVA) further revealed that the total genetic variation among populations was 21%, whilst variation within populations was estimated to be 79%. After grouping, the majority of total microsatellite loci variation appeared to be due to variation within populations. A relationship tree based on Nei's D A genetic distance and the UPGMA algorithm grouped the strains and populations into two major clusters, reflecting their affiliation to the European/Central Asian or East-Asian subspecies of common carp, rather than their country of origin. An individual assignment test (self-classification) using a Bayesian method demonstrated 89.6% accuracy in assigning individuals into their correct strains/population of origin at P < 0.05. Thus, the set of 10 highly polymorphic microsatellite markers used in the present study demonstrated significant worth in monitoring the genetic diversity of cultured common carp. Additionally, our results proved the efficiency of the conservation programme and highlighted the need for better control of genetic variability in common carp strains. This could also be useful for the conservation of carp strains in the Czech Republic.

Variable correlation of genome GC% with transfer RNA number as well as with transfer RNA diversity among bacterial groups: α-Proteobacteria and Tenericutes exhibit strong positive correlation

Mainly two selective forces are thought to affect transfer RNA (tRNA) gene numbers in cells: firstly, transfer RNA genes have coevolved with codon usage to optimize the translation rate in organisms; and secondly, tRNA gene number is positively correlated with growth rate of organisms. Since a codon ending with 'G' is recognized with lower efficiency than the synonymous codon ending with 'A' by a tRNA, organisms with high genome GC% are expected to possess tRNA molecules with 'C' at the first anticodon position to decode efficiently the former codon. This is in concordance with the observation of higher tRNA diversity in several bacteria with higher genome GC%. In addition to this, tRNA gene redundancy has been reported to be correlated with genome size. In this paper we have carried out a comprehensive correlation analysis of tRNA gene diversity, total tRNA number, genome GC% and genome size, in 626 bacterial strains (belonging to 433 species). A common positive trend of correlation in bacteria of GC% with tRNA diversity, of GC% with genome size and of genome size with total tRNA number, is observed, even within individual bacterial groups. Comparison of organisms' growth rate data with their genome GC% indicated that organisms with higher growth rate are most likely with <50% GC. This work extends the list of features in organisms that relate to genome GC%.

Bacterial community composition and chitinase gene diversity of vermicompost with antifungal activity

Bacterial communities and chitinase gene diversity of vermicompost (VC) were investigated to clarify the influence of earthworms on the inhibition of plant pathogenic fungi in VC. The spore germination of Fusarium moniliforme was reduced in VC aqueous extracts prepared from paper sludge and dairy sludge (fresh sludge, FS). The bacterial communities were examined by culture-dependent and -independent analyses. Unique clones selected from 16S rRNA libraries of FS and VC on the basis of restriction fragment length polymorphism (RFLP) fell into the major lineages of the domain bacteria Proteobacteria, Bacteroidetes, Verrucomicrobia, Actinobacteria and Firmicutes. Among culture isolates, Actinobacteria dominated in VC, while almost equal numbers of Actinobacteria and Proteobacteria were present in FS. Analysis of chitinolytic isolates and chitinase gene diversity revealed that chitinolytic bacterial communities were enriched in VC. Populations of bacteria that inhibited plant fungal pathogens were higher in VC than in FS and particularly chitinolytic isolates were most active against the target fungi.

Analysis of nifH gene diversity in red soil amended with manure in Jiangxi, south China

In order to understand the community structure of diazotrophs in red soil and effects of organic manure Application on the structure, four nifH gene libraries were constructed: the control (CK), low manure (LM), High manure (HM), and high manure adding lime (ML). Totally 150 nifH gene clones were screened and grouped into 21 clusters by RFLP analysis. Existence of dominant patterns was observed in all libraries, which counted for over 96% of clones in library HM and about 56 approximately 72% in other three libraries. The nifH sequences of the dominant patterns in all libraries were most similar to sequences of the cyanobacteria. nifH genes showed high diversity in red soil, dispersing throughout the nifH clades (alpha-, beta-, and gamma-Proteobacteria, Firmicutes, cyanobacteria, Verrucomicrobia, and posited group). Bradyrhizobium and Burkholderia were also important diaxotrophs in low fertility soil samples. Low manure treatment increased the Diversity of nifH genes compared with CK and high manure treatments. Manure and lime treatment led to obvious community succession. Total N to available P ratio, total carbon, and K concentrations were the main factors affecting the diversity of diazotrophs in red soil.

Local scale population genetic structure of Entomophthora muscae epidemics

Abstract Isolates of the entomopathogenic fungus Entomophthora muscae were obtained from houseflies sampled at five stables on Zealand during an epidemic in fall 2005. DNA fingerprints were generated from single conidia from 40 E. muscae isolates using the PCR-based method of Inter-Simple Sequence Repeats (ISSR). This resulted in fingerprint patterns consisting of about 50 fragments, of which, 14 were polymorphic. From the polymorphic loci we estimated the reproductive mode, genetic differentiation and gene diversity of E. muscae populations using the statistics of index of association, Weir & Cockerhams theta and Nei's analysis of gene diversity in subdivided populations. Our results revealed no significant differences in allele frequencies among the five populations. The index of association test rejected the null hypothesis of random mating, but the test for paired locus compatibility showed weak or no linkage between loci indicating recombination or homoplasy in the dataset. From this study we cannot exclude the possibility that the population genetic structure underlying E. muscae epidemics could be panmictic consisting of several lineages with a high level of reciprocal migration.

Genetic differentiation in endangered Gynostemma pentaphyllum (Thunb.) Makino based on ISSR polymorphism and its implications for conservation

Studies were performed to investigate the genetic variation of 14 natural populations of Gynostemma pentaphyllum (Thunb.) Makino, an outcrossing clonal plant species in China, using inter-simple sequence repeat (ISSR) markers. Fourteen selected primers were used to amplify DNA samples from 140 individuals, and totally 194 loci were detected. The percentage of polymorphic bands (PPBs) showed that the genetic diversity was pretty high at the species level (PPB = 96.39%) but quite low at the population level (PPB = 1.03–25.26%). Shannon's information index ( I) and Nei's gene diversity ( h) displayed a similar trend to PPB. According to the hierarchical analysis of molecular variance (AMOVA) and Nei's analysis of gene diversity, the percentages of genetic variation among populations were 88.66 and 88.94%, respectively, indicating a high level of inter-population genetic differentiation. The low levels of genetic diversity within populations and high genetic differentiation among populations were assumed to result from the limited gene flow, the clonal nature and genetic drift. Based on the genetic data, effective conservation strategies were proposed for conserving this traditional Chinese medicinal herb. Concerning the management of G. pentaphyllum, we suggested that in situ conservation be an important and practical measure for maintaining the genetic diversity and that a possibly maximum number of populations be conserved. Populations EMS and HLT, in which particularly low levels of genetic variation were characterized, should be given the priority for ex situ conservation.

Genetic structure of arbuscular mycorrhizal populations in fallow and cultivated soils

The impact of fallowing on the genetic structure of arbuscular mycorrhizal fungi (AMF) was studied by hierarchical sampling of spores from four plots in a fallow and a cultivated field. A nested multiplex PCR approach was used to assign the spores to genotypes. Variable introns of the two protein-coding genes GmFOX2 and GmTOR2 were used as co-dominant genetic markers together with the large subunit (LSU) rDNA. The gene diversity and genetic structure of Glomus mosseae, Glomus geosporum and Glomus caledonium were compared within and between the fields. Spores of G. caledonium and G. geosporum were more abundant in the cultivated field, whereas G. mosseae was more frequent in the fallow field. The number of genotypes was not different between the two fields. Analysis of gene diversity of G. caledonium in the fallow field indicated that a larger part of the heterogeneity could be attributed to variation between plots rather than subplots, suggesting that the lack of soil cultivation resulted in more heterogeneous population genetic structures. Analyses of haplotype networks of the fungi suggested a subdivision of G. mosseae haplotypes between the two fields, whereas no such division was seen in G. geosporum and G. caledonium. The results show that agricultural practices differently affect both the abundance and the population structure of different AMF species.

Ecological genetics of Reaumuria soongorica (Pall.) Maxim. population in the oasis–desert ecotone in Fukang, Xinjiang, and its implications for molecular evolution

Using inter-simple sequence repeat (ISSR) markers, studies were performed to characterize the population genetic diversity and structure of Reaumuria soongorica (Pall.) Maxim. in the oasis–desert ecotone in Fukang, Xinjiang. Eleven primers were screened to amplify DNA sequences from 132 individuals, which corresponded to seven subpopulations. Totally, 176 loci, all of them polymorphic, were detected, and the percentage of polymorphic bands (PPB) was 100%, indicating a relatively high genetic diversity within the R. soongorica population. According to the hierarchical analysis of molecular variances (AMOVA), the analysis of Shannon's diversity and Nei's analysis of gene diversity, the percentages of genetic variation among subpopulations were 15.87%, 16.28% and 14.58%, respectively, which meant that 83.72–85.42% of total genetic variation occurred within subpopulations. Besides, POPGENE analysis also revealed a relatively high gene flow ( N m = 2.9290) among subpopulations. Correlation analysis showed that there existed no significant correlation between the ISSR-based genetic diversity of the seven R. soongorica subpopulations and their ecological factors (mainly in soil) ( P > 0.05). However, on the other hand, Mantel test revealed a significant correlation between inter-subpopulation genetic distances and geographic distances ( r = 0.637, P < 0.01), indicating that geographic distance was among the important factors affecting the population genetic structure of R. soongorica. Meanwhile, a comparative analysis was performed between the present ISSR-based and the previously published RAPD-based results, and revealed obvious discrepancies, implying the different evolutionary patterns of the genomic regions sampled by ISSR and RAPD markers.

Identity of active methanotrophs in landfill cover soil as revealed by DNA-stable isotope probing

A considerable amount of methane produced during decomposition of landfill waste can be oxidized in landfill cover soil by methane-oxidizing bacteria (methanotrophs) thus reducing greenhouse gas emissions to the atmosphere. The identity of active methanotrophs in Roscommon landfill cover soil, a slightly acidic peat soil, was assessed by DNA-stable isotope probing (SIP). Landfill cover soil slurries were incubated with (13)C-labelled methane and under either nutrient-rich nitrate mineral salt medium or water. The identity of active methanotrophs was revealed by analysis of (13)C-labelled DNA fractions. The diversity of functional genes (pmoA and mmoX) and 16S rRNA genes was analyzed using clone libraries, microarrays and denaturing gradient gel electrophoresis. 16S rRNA gene analysis revealed that the cover soil was mainly dominated by Type II methanotrophs closely related to the genera Methylocella and Methylocapsa and to Methylocystis species. These results were supported by analysis of mmoX genes in (13)C-DNA. Analysis of pmoA gene diversity indicated that a significant proportion of active bacteria were also closely related to the Type I methanotrophs, Methylobacter and Methylomonas species. Environmental conditions in the slightly acidic peat soil from Roscommon landfill cover allow establishment of both Type I and Type II methanotrophs.

Genetic diversity for aluminum tolerance in sorghum

Genetic variation for aluminum (Al) tolerance in plants has allowed the development of cultivars that are high yielding on acidic, Al toxic soils. However, knowledge of intraspecific variation for Al tolerance control is needed in order to assess the potential for further Al tolerance improvement. Here we focused on the major sorghum Al tolerance gene, Alt ( SB ), from the highly Al tolerant standard SC283 to investigate the range of genetic diversity for Al tolerance control in sorghum accessions from diverse origins. Two tightly linked STS markers flanking Alt ( SB ) were used to study the role of this locus in the segregation for Al tolerance in mapping populations derived from different sources of Al tolerance crossed with a common Al sensitive tester, BR012, as well as to isolate the allelic effects of Alt ( SB ) in near-isogenic lines. The results indicated the existence not only of multiple alleles at the Alt ( SB ) locus, which conditioned a wide range of tolerance levels, but also of novel sorghum Al tolerance genes. Transgressive segregation was observed in a highly Al tolerant breeding line, indicating that potential exists to exploit the additive or codominant effects of distinct Al tolerance loci. A global, SSR-based, genetic diversity analysis using a broader sorghum set revealed the presence of both multiple Alt ( SB ) alleles and different Al tolerance genes within highly related accessions. This suggests that efforts toward broadening the genetic basis for Al tolerance in sorghum may benefit from a detailed analysis of Al tolerance gene diversity within subgroups across a target population.

Chitinase Genes in Lake Sediments of Ardley Island, Antarctica

A sediment core spanning approximately 1,600 years was collected from a lake on Ardley Island, Antarctica. The sediment core had been greatly influenced by penguin guano. Using molecular methods, the chitinolytic bacterial community along the sediment core was studied over its entire length. Primers targeting conserved sequences of the catalytic domains of family 18 subgroup A chitinases detected group A chitinases from a wide taxonomic range of bacteria. Using quantitative competitive PCR (QC-PCR), chitinase gene copies in each 1-cm section of the whole sediment column were quantified. QC-PCR determination of the chitinase gene copies indicated significant correlation with phosphorus and total organic carbon concentration, suggesting a historical connection between chitinase gene copies and the amount of penguin guano input into the lake sediment. Most of the chitinase genes cloned from the historic sediment core were novel. Analysis of the chitinase gene diversity in selected sediment layers and in the fresh penguin deposits indicated frequent shifts in the chitinolytic bacterial community over time. Sequence analysis of the 16S rRNA genes of chitinolytic bacteria isolated from the lake sediment revealed that the isolates belonged to Janthinobacterium species, Stenotrophomonas species of gamma-Proteobacteria, Cytophaga species of the Cytophaga-Flexibacter-Bacteroides group, and Streptomyces and Norcardiopsis species of Actinobacteria. Chitinase gene fragments were cloned and sequenced from these cultivated chitinolytic bacteria. The phylogeny of the chitinase genes obtained from the isolates did not correspond well to that of the isolates, suggesting acquisition via horizontal gene transfer.

Investigation of the methanogen population structure and activity in a brackish lake sediment

The methanogen community in sediment from the edge of a small brackish lake connected to the Beaulieu Estuary (Hampshire, UK) was investigated by analysis of 16S rRNA gene diversity using new methanogen-specific primers plus Archaea-specific primers. 16S rRNA gene primers previously used for polymerase chain reaction (PCR) detection of methanogenic Archaea from a variety of environments were evaluated by in silico testing. The primers displayed variable coverage of the four main orders of methanogens, highlighting the importance of this type of primer evaluation. Three PCR primer sets were designed using novel reverse primers to facilitate specific amplification of the orders Methanomicrobiales/Methanosarcinales, Methanobacteriales and Methanococcales. Diversity of the methanogen functional gene, methyl coenzyme M reductase (mcrA), was also studied. All gene libraries constructed from this sediment indicated that Methanomicrobiales and Methanosarcinales were the only methanogens detected. There was good agreement between the relative sequence abundances in the methanogen-specific 16S rRNA gene library and terminal restriction fragment length polymorphism (T-RFLP) profiling, suggesting that the population was dominated by putative H2 CO2 utilizing Methanomicrobiales, although acetate-utilizing methanogens were also present. The methanogen population analyses were in agreement with methanogenic activity measurements, which indicated that bicarbonate methanogenesis was higher than acetate methanogenesis at all depths measured and overall there was a significant difference (P = 0.001) between the rates of the two pathways. This study demonstrates the utility of new 16S rRNA gene PCR primers targeting specific methanogenic orders, and the combined results suggest that the CO2 reduction pathway dominates methanogenesis in the brackish sediment investigated.