Microbial Ecology Studies Research Articles

Fungal Community Structure of Macroalga Ulva intestinalis Revealed by Miseq Sequencing

Since algae are well known as the host of fungi, the information about total fungal community associated with alga is needed to give the initial data that is useful for natural product discovery. The study on total fungal community associated with marine macroalga U. intestinalis from Xiamen-China was carried out using MiSeq, a powerful platform for microbial ecology studies. Metagenomics DNA was isolated from surfaces of the U. intestinalis thallus and rocks colonized by this alga and 18S ribosomal RNA region was sequenced using MiSeq Illumina to yield 81360 sequences from 162720 reads, which average read length is 245.61 bp. The sequences were assigned to operational taxonomic units (OTUs), where 101 genera represent 5 phyla, 17 classes and 64 families of fungal. Ascomycota was the highest abundance share of fungal community at phylum level. Meanwhile Aspergillus, Blastobotrys, Alternaria, Knufia, and Fusarium were some of dominant genera found associated with U. intestinalis which have been reported as the promising source of natural products. By using MiSeq platform, this study revealed the total community structure of algicolous fungi associated with U. intestinalis.

HIGH-QUALITY DNA FROM PEAT SOIL FOR METAGENOMIC STUDIES: A MINIREVIEW ON DNA EXTRACTION METHODS

The urge of understanding soil microbial function and diversity of peatland soils either forested or non-forested has emerged since over a decade ago concerning the ongoing the ecosystem deterioration and its environmental impacts it causes. Like any other soil environments, this unique ecosystem possesses great challenges in isolating quality DNA prerequisite for microbial ecology studies in virtue of its physicochemical characteristics. Two general approaches have been extensively applied for soil microbial DNA extraction which are termed as direct DNA extraction protocol and indirect DNA extraction protocol. The only step differentiate between these two protocols is the later includes additional cell separation method from soil microaggregates preceding DNA extraction process. Therefore, several improved and modified methods in conventional DNA extraction and purification methods are reviewed in this paper to cater all the highlighted issues as to obtain high-quality DNA for soil peatland metagenomics studies.

Bulk soil bacterial community mediated by plant community in Mediterranean ecosystem, Israel

The Israeli Mediterranean ecosystem has a distinctive flora with high levels of plant diversity. Previous studies of soil microbial ecology from the region have paid little attention to the possibility of distinctive soil bacterial communities associated with particular habitats. This study looked at bacterial communities present in the bulk soil from the closed canopy of four indigenous plants – Quercus ithaburensis, Calicotome villosa, Sarcopoterium spinosum, and Rhamnus puncata. Soil DNA was amplified for the 16S rRNA gene and sequenced using MiSeq Illumina platform. Results indicated that the bacterial communities differed between open and closed canopy with no significant difference in community structure within closed canopy, with soil moisture and organic matter content significantly influenced the community. Only the relative abundance of Gemmatimonadetes varied significantly with the open canopy having the highest abundance, and the relative abundance of Spartobacteria and Saprospirae was unusually high compared to other Mediterranean soils. There was no difference in OTU richness between plant species. Additionally, this study revealed that open canopy community between plants was nested within closed canopy bacterial community implying that open canopy community potentially serves as a source for the closed canopy. It is then possible that the rhizospheric influence extends beyond the root-soil interface and to the surrounding bulk soil. Overall, this study indicates that in Israeli Mediterranean ecosystems native plants extend a selection beyond the rhizosphere, and like other studies from Mediterranean ecosystems pH does not seem to play an integral role in structuring bacterial communities.

Preface Squalen Bulletin Vol. 12 No. 2 Tahun 2017

Since algae are well known as the host of fungi, the information about total fungal community associated with alga is needed to give the initial data that is useful for natural product discovery. The study on total fungal community associated with marine macroalga U. intestinalis from Xiamen-China was carried out using MiSeq, a powerful platform for microbial ecology studies. Metagenomics DNA was isolated from surfaces of the U. intestinalis thallus and rocks colonized by this alga and 18S ribosomal RNA region was sequenced using MiSeq Illumina to yield 81360 sequences from 162720 reads, which average read length is 245.61 bp. The sequences were assigned to operational taxonomic units (OTUs), where 101 genera represent 5 phyla, 17 classes and 64 families of fungal. Ascomycota was the highest abundance share of fungal community at phylum level. Meanwhile Aspergillus, Blastobotrys, Alternaria, Knufia, and Fusarium were some of dominant genera found associated with U. intestinalis which have been reported as the promising source of natural products. By using MiSeq platform, this study revealed the total community structure of algicolous fungi associated with U. intestinalis.

Distinct taxonomic and functional composition of soil microbiomes along the gradient forest-restinga-mangrove in southeastern Brazil.

Soil microorganisms play crucial roles in ecosystem functioning, and the central goal in microbial ecology studies is to elucidate which factors shape community structure. A better understanding of the relationship between microbial diversity, functions and environmental parameters would increase our ability to set conservation priorities. Here, the bacterial and archaeal community structure in Atlantic Forest, restinga and mangrove soils was described and compared based on shotgun metagenomics. We hypothesized that each distinct site would harbor a distinct taxonomic and functional soil community, which is influenced by environmental parameters. Our data showed that the microbiome is shaped by soil properties, with pH, base saturation, boron and iron content significantly correlated to overall community structure. When data of specific phyla were correlated to specific soil properties, we demonstrated that parameters such as boron, copper, sulfur, potassium and aluminum presented significant correlation with the most number of bacterial groups. Mangrove soil was the most distinct site and presented the highest taxonomic and functional diversity in comparison with forest and restinga soils. From the total 34 microbial phyla identified, 14 were overrepresented in mangrove soils, including several archaeal groups. Mangrove soils hosted a high abundance of sequences related to replication, survival and adaptation; forest soils included high numbers of sequences related to the metabolism of nutrients and other composts; while restinga soils included abundant genes related to the metabolism of carbohydrates. Overall, our finds show that the microbial community structure and functional potential were clearly different across the environmental gradient, followed by functional adaptation and both were related to the soil properties.

Diversity of bacterial communities in French chicken cuts stored under modified atmosphere packaging

Poultry meat, the second most consumed meat in France, is commercialized mainly as portions of chicken cuts with various quality labels, stored under various modified atmosphere packaging (MAP), with shelf-life ranging from 9 to 17 days. We used 16S rDNA pyrosequencing to describe microbiota of chicken legs. Ten samples representing a wide diversity of labels and MAP available on the market were collected from local supermarkets and stored at 4 °C. Microbiota were collected, total DNA was extracted, and V1-V3 fragment of 16S rRNA genes were amplified and sequenced. For data analysis several pipelines were compared. The Qiime pipeline was chosen to cluster reads and we used a database previously developed for a meat and fish microbial ecology study. Variability between samples was observed and a listing of bacteria present on chicken meat was established. The structure of the bacterial communities were compared with traditional cultural methods and validated with quantitative real time PCR. Brochothrix thermosphacta, Pseudomonas sp., and Carnobacterium sp. were dominant and the nature of the gas used for packaging influenced the relative abundance of each suggesting a MAP gas composition dependent competition between these species. We also noticed that slaughterhouse environment may influence the nature of the contaminants.

Trends in soil microbial communities during secondary succession

Succession, a central theme throughout the history of ecology, has been well studied predominantly in plant communities, but the general trends in soil microbial communities during succession remain unclear. Here, we compiled a comprehensive data set of 85 age sequences worldwide with the aims to (1) examine the trends in soil microbial composition, bioenergetics, and activity during secondary succession, and (2) explore their coordinating changes with soil properties. The results showed that the fungi to bacteria ratio (fungi:bacteria) increased, while the microbial respiration per unit biomass carbon (R/Cmic) decreased as the succession proceeds. Secondary succession had the rising trends in microbial biomass carbon to soil carbon ratio (Cmic/Csoil) and microbial biomass nitrogen to soil nitrogen ratio (Nmic/Nsoil). These successional trends in microbes were coincident with the macro-ecological succession theory in plants and animals. Specifically, early successional stages tended to be dominated by r-strategists (bacteria) that had higher R/Cmic and lower Cmic/Csoil and Nmic/Nsoil, whereas late successional stages tended to be dominated by K-strategists (fungi) that behaved oppositely. The soil C to N ratio (C:Nsoil) increased significantly with the successional stage, with a fast increasing C:Nsoil ratio being accompanied by a fast increase of fungi:bacteria, a slow decrease of R/Cmic, and a slow increase of Cmic/Csoil. This result suggests that the stoichiometry theory may provide a feasible approach to explain the divergent successional trends in microbial communities. In conclusion, our global synthesis highlights the application of the existing macro-ecological theory to soil microbial ecology studies.

Different Amplicon Targets for Sequencing-Based Studies of Fungal Diversity

Target-gene amplicon sequencing is the most exploited high-throughput sequencing application in microbial ecology. The targets are taxonomically relevant genes, with 16S rRNA being the gold standard for bacteria. As for fungi, the most commonly used target is the internal transcribed spacer (ITS). However, the uneven ITS length among species may promote preferential amplification and sequencing and incorrect estimation of their abundance. Therefore, the use of different targets is desirable. We evaluated the use of three different target amplicons for the characterization of fungal diversity. After an in silico primer evaluation, we compared three amplicons (the ITS1-ITS2 region [ITS1-2], 18S ribosomal small subunit RNA, and the D1/D2 domain of the 26S ribosomal large subunit RNA), using biological samples and a mock community of common fungal species. All three targets allowed for accurate identification of the species present. Nevertheless, high heterogeneity in ITS1-2 length was found, and this caused an overestimation of the abundance of species with a shorter ITS, while both 18S and 26S amplicons allowed for more reliable quantification. We demonstrated that ITS1-2 amplicon sequencing, although widely used, may lead to an incorrect evaluation of fungal communities, and efforts should be made to promote the use of different targets in sequencing-based microbial ecology studies.IMPORTANCE Amplicon-sequencing approaches for fungi may rely on different targets affecting the diversity and abundance of the fungal species. An increasing number of studies will address fungal diversity by high-throughput amplicon sequencing. The description of the communities must be accurate and reliable in order to draw useful insights and to address both ecological and biological questions. By analyzing a mock community and several biological samples, we demonstrate that using different amplicon targets may change the results of fungal microbiota analysis, and we highlight how a careful choice of the target is fundamental for a thorough description of the fungal communities.

Microbial ecology-based engineering of Microbial Electrochemical Technologies.

SummaryMicrobial ecology is devoted to the understanding of dynamics, activity and interaction of microorganisms in natural and technical ecosystems. Bioelectrochemical systems represent important technical ecosystems, where microbial ecology is of highest importance for their function. However, whereas aspects of, for example, materials and reactor engineering are commonly perceived as highly relevant, the study and engineering of microbial ecology are significantly underrepresented in bioelectrochemical systems. This shortfall may be assigned to a deficit on knowledge and power of these methods as well as the prerequisites for their thorough application. This article discusses not only the importance of microbial ecology for microbial electrochemical technologies but also shows which information can be derived for a knowledge‐driven engineering. Instead of providing a comprehensive list of techniques from which it is hard to judge the applicability and value of information for a respective one, this review illustrates the suitability of selected techniques on a case study. Thereby, best practice for different research questions is provided and a set of key questions for experimental design, data acquisition and analysis is suggested.

Microbial community profiling of fresh basil and pitfalls in taxonomic assignment of enterobacterial pathogenic species based upon 16S rRNA amplicon sequencing

Application of 16S rRNA (gene) amplicon sequencing on food samples is increasingly applied for assessing microbial diversity but may as unintended advantage also enable simultaneous detection of any human pathogens without a priori definition. In the present study high-throughput next-generation sequencing (NGS) of the V1–V2–V3 regions of the 16S rRNA gene was applied to identify the bacteria present on fresh basil leaves. However, results were strongly impacted by variations in the bioinformatics analysis pipelines (MEGAN, SILVAngs, QIIME and MG-RAST), including the database choice (Greengenes, RDP and M5RNA) and the annotation algorithm (best hit, representative hit and lowest common ancestor). The use of pipelines with default parameters will lead to discrepancies. The estimate of microbial diversity of fresh basil using 16S rRNA (gene) amplicon sequencing is thus indicative but subject to biases.Salmonella enterica was detected at low frequencies, between 0.1% and 0.4% of bacterial sequences, corresponding with 37 to 166 reads. However, this result was dependent upon the pipeline used: Salmonella was detected by MEGAN, SILVAngs and MG-RAST, but not by QIIME. Confirmation of Salmonella sequences by real-time PCR was unsuccessful. It was shown that taxonomic resolution obtained from the short (500bp) sequence reads of the 16S rRNA gene containing the hypervariable regions V1–V3 cannot allow distinction of Salmonella with closely related enterobacterial species.In conclusion 16S amplicon sequencing, getting the status of standard method in microbial ecology studies of foods, needs expertise on both bioinformatics and microbiology for analysis of results. It is a powerful tool to estimate bacterial diversity but amenable to biases. Limitations concerning taxonomic resolution for some bacterial species or its inability to detect sub-dominant (pathogenic) species should be acknowledged in order to avoid overinterpretation of results.

Genome Sequences for Streptomyces spp. Isolated from Disease-Suppressive Soils and Long-Term Ecological Research Sites.

ABSTRACTWe report here the high-quality genome sequences of three Streptomyces spp. isolated as part of a long-term study of microbial soil ecology. Streptomyces sp. strain GS93-23 was isolated from naturally disease-suppressive soil (DSS) in Grand Rapids, MN, and Streptomyces sp. strains S3-4 and 3211-3 were isolated from experimental plots in the Cedar Creek Ecosystem Science Reserve (CCESR).

Chemotaxonomic characterisation of the thaumarchaeal lipidome.

Thaumarchaeota are globally distributed and abundant microorganisms occurring in diverse habitats and thus represent a major source of archaeal lipids. The scope of lipids as taxonomic markers in microbial ecological studies is limited by the scarcity of comparative data on the membrane lipid composition of cultivated representatives, including the phylum Thaumarchaeota. Here, we comprehensively describe the core and intact polar lipid (IPL) inventory of ten ammonia-oxidising thaumarchaeal cultures representing all four characterized phylogenetic clades. IPLs of these thaumarchaeal strains are generally similar and consist of membrane-spanning, glycerol dibiphytanyl glycerol tetraethers with monoglycosyl, diglycosyl, phosphohexose and hexose-phosphohexose headgroups. However, the relative abundances of these IPLs and their core lipid compositions differ systematically between the phylogenetic subgroups, indicating high potential for chemotaxonomic distinction of thaumarchaeal clades. Comparative lipidomic analyses of 19 euryarchaeal and crenarchaeal strains suggested that the lipid methoxy archaeol is synthesized exclusively by Thaumarchaeota and may thus represent a diagnostic lipid biomarker for this phylum. The unprecedented diversity of the thaumarchaeal lipidome with 118 different lipids suggests that membrane lipid composition and adaptation mechanisms in Thaumarchaeota are more complex than previously thought and include unique lipids with as yet unresolved properties.

Bioinformatic Approaches Including Predictive Metagenomic Profiling Reveal Characteristics of Bacterial Response to Petroleum Hydrocarbon Contamination in Diverse Environments

Microbial remediation of oil polluted habitats remains one of the foremost methods for restoration of petroleum hydrocarbon contaminated environments. The development of effective bioremediation strategies however, require an extensive understanding of the resident microbiome of these habitats. Recent developments such as high-throughput sequencing has greatly facilitated the advancement of microbial ecological studies in oil polluted habitats. However, effective interpretation of biological characteristics from these large datasets remain a considerable challenge. In this study, we have implemented recently developed bioinformatic tools for analyzing 65 16S rRNA datasets from 12 diverse hydrocarbon polluted habitats to decipher metagenomic characteristics of the resident bacterial communities. Using metagenomes predicted from 16S rRNA gene sequences through PICRUSt, we have comprehensively described phylogenetic and functional compositions of these habitats and additionally inferred a multitude of metagenomic features including 255 taxa and 414 functional modules which can be used as biomarkers for effective distinction between the 12 oil polluted sites. Additionally, we show that significantly over-represented taxa often contribute to either or both, hydrocarbon degradation and additional important functions. Our findings reveal significant differences between hydrocarbon contaminated sites and establishes the importance of endemic factors in addition to petroleum hydrocarbons as driving factors for sculpting hydrocarbon contaminated bacteriomes.

Microdiversity shapes the traits, niche space, and biogeography of microbial taxa.

With rapidly improving sequencing technologies, scientists have recently gained the ability to examine diverse microbial communities at high genomic resolution, revealing that both free-living and host-associated microbes partition their environment at fine phylogenetic scales. This 'microdiversity,' or closely related (> 97% similar 16S rRNA gene) but ecologically and physiologically distinct sub-taxonomic groups, appears to be an intrinsic property of microorganisms. However, the functional implications of microdiversity as well as its effects on microbial biogeography are poorly understood. Here, we present two theoretical models outlining the evolutionary mechanisms that drive the formation of microdiverse 'sub-taxa.' Additionally, we review recent literature and reveal that microdiversity influences a wide range of functional traits across diverse ecosystems and microbes. Moving to higher levels of organization, we use laboratory data from marine, soil, and host-associated bacteria to demonstrate that the aggregated trait-based response of microdiverse sub-taxa modifies the fundamental niche of microbes. The correspondence between microdiversity and niche space represents a critical tool for future studies of microbial ecology. By combining growth experiments on diverse isolates with examinations of environmental abundance patterns, researchers can better quantify the fundamental and realized niches of microbes and improve understanding of microbial biogeography and response to future environmental change.

Methods for the estimation of the biovolume of microorganisms: A critical review

AbstractBiomass estimation is central in most microbial plankton ecology studies. Numerous advances have been made that rendered this task more and more effective, from the introduction of allometric volume‐to‐carbon conversion factors to the development of submersible flow‐through imaging devices. Although several ingenious methods have been suggested for this purpose, the problem of biovolume extrapolation from two‐dimensional pictures remains. This study offers a historical and critical review of the methods for the estimation of the biovolume of microorganisms that have been proposed throughout the last four decades. It covers three main approaches—respectively based on a single equation, a set of simple or composite geometric shapes or a computational algorithm—as well as several methods featuring exclusive characteristics. Both strengths and shortcomings of each technique are analyzed, with particular emphasis on compatibility with the most recent flow‐through devices coupled with in situ imaging systems. A composite picture emerges with no such unique method, at present, suitable for every situation. This review can thus be a valuable guide for the microbial plankton ecologist who seeks the most suitable technique for a specific circumstance.

Draft Genome Sequence of Deep-Sea Alteromonas sp. Strain V450 Isolated from the Marine Sponge Leiodermatium sp.

ABSTRACTThe proteobacterium Alteromonas sp. strain V450 was isolated from the Atlantic deep-sea sponge Leiodermatium sp. Here, we report the draft genome sequence of this strain, with a genome size of approx. 4.39 Mb and a G+C content of 44.01%. The results will aid deep-sea microbial ecology, evolution, and sponge-microbe association studies.

Assessing Species Diversity Using Metavirome Data: Methods and Challenges

Assessing biodiversity is an important step in the study of microbial ecology associated with a given environment. Multiple indices have been used to quantify species diversity, which is a key biodiversity measure. Measuring species diversity of viruses in different environments remains a challenge relative to measuring the diversity of other microbial communities. Metagenomics has played an important role in elucidating viral diversity by conducting metavirome studies; however, metavirome data are of high complexity requiring robust data preprocessing and analysis methods. In this review, existing bioinformatics methods for measuring species diversity using metavirome data are categorised broadly as either sequence similarity-dependent methods or sequence similarity-independent methods. The former includes a comparison of DNA fragments or assemblies generated in the experiment against reference databases for quantifying species diversity, whereas estimates from the latter are independent of the knowledge of existing sequence data. Current methods and tools are discussed in detail, including their applications and limitations. Drawbacks of the state-of-the-art method are demonstrated through results from a simulation. In addition, alternative approaches are proposed to overcome the challenges in estimating species diversity measures using metavirome data.

Analysis of 16S rRNA Gene Amplicon Sequences Using the QIIME Software Package.

The study of microbial ecology has undergone a paradigm shift in recent years, with rapid advances in molecular and bioinformatic tools allowing researchers with wide-ranging interests and backgrounds access to community profiling methods. While these advances have undoubtedly led to exciting new understanding of many systems, the array of protocols available and the idiosyncrasies of particular approaches can lead to confusion or, at worst, erroneous interpretation of results. Here, we describe a workflow from raw 16S rRNA gene amplicon sequence data, generated on an Illumina MiSeq instrument, to microbial community taxonomy profiles and basic diversity measures. The workflow can be adapted to input from major sequence platforms and uses freely available open source software that can be implemented on a range of operating systems.

Microbial Community Patterns Associated with Automated Teller Machine Keypads in New York City.

In densely populated urban environments, the distribution of microbes and the drivers of microbial community assemblages are not well understood. In sprawling metropolitan habitats, the "urban microbiome" may represent a mix of human-associated and environmental taxa. Here we carried out a baseline study of automated teller machine (ATM) keypads in New York City (NYC). Our goal was to describe the biodiversity and biogeography of both prokaryotic and eukaryotic microbes in an urban setting while assessing the potential source of microbial assemblages on ATM keypads. Microbial swab samples were collected from three boroughs (Manhattan, Queens, and Brooklyn) during June and July 2014, followed by generation of Illumina MiSeq datasets for bacterial (16S rRNA) and eukaryotic (18S rRNA) marker genes. Downstream analysis was carried out in the QIIME pipeline, in conjunction with neighborhood metadata (ethnicity, population, age groups) from the NYC Open Data portal. Neither the 16S nor 18S rRNA datasets showed any clustering patterns related to geography or neighborhood demographics. Bacterial assemblages on ATM keypads were dominated by taxonomic groups known to be associated with human skin communities (Actinobacteria, Bacteroides, Firmicutes, and Proteobacteria), although SourceTracker analysis was unable to identify the source habitat for the majority of taxa. Eukaryotic assemblages were dominated by fungal taxa as well as by a low-diversity protist community containing both free-living and potentially pathogenic taxa (Toxoplasma, Trichomonas). Our results suggest that ATM keypads amalgamate microbial assemblages from different sources, including the human microbiome, eukaryotic food species, and potentially novel extremophilic taxa adapted to air or surfaces in the built environment. DNA obtained from ATM keypads may thus provide a record of both human behavior and environmental sources of microbes. IMPORTANCE Automated teller machine (ATM) keypads represent a specific and unexplored microhabitat for microbial communities. Although the number of built environment and urban microbial ecology studies has expanded greatly in recent years, the majority of research to date has focused on mass transit systems, city soils, and plumbing and ventilation systems in buildings. ATM surfaces, potentially retaining microbial signatures of human inhabitants, including both commensal taxa and pathogens, are interesting from both a biodiversity perspective and a public health perspective. By focusing on ATM keypads in different geographic areas of New York City with distinct population demographics, we aimed to characterize the diversity and distribution of both prokaryotic and eukaryotic microbes, thus making a unique contribution to the growing body of work focused on the "urban microbiome." In New York City, the surface area of urban surfaces in Manhattan far exceeds the geographic area of the island itself. We have only just begun to describe the vast array of microbial taxa that are likely to be present across diverse types of urban habitats.

Metagenomics insights into food fermentations.

SummaryThis review describes the recent advances in the study of food microbial ecology, with a focus on food fermentations. High‐throughput sequencing (HTS) technologies have been widely applied to the study of food microbial consortia and the different applications of HTS technologies were exploited in order to monitor microbial dynamics in food fermentative processes. Phylobiomics was the most explored application in the past decade. Metagenomics and metatranscriptomics, although still underexploited, promise to uncover the functionality of complex microbial consortia. The new knowledge acquired will help to understand how to make a profitable use of microbial genetic resources and modulate key activities of beneficial microbes in order to ensure process efficiency, product quality and safety.