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At least 19 records
All sourcesNASA NTRSDOE OSTI
DOE OSTI2022

Cryptosporulation in Kurthia spp. forces a rethinking of asporogenesis in Firmicutes

Endosporulation is a complex morphophysiological process resulting in a more resistant cellular structure that is produced within the mother cell and is called endospore. Endosporulation evolved in the common ancestor of Firmicutes, but it is lost in descendant lineages classified as asporogenic. While Kurthia spp. is considered to comprise only asporogenic species, we show here that strain 11kri321, which was isolated from an oligotrophic geothermal reservoir, produces phase-bright spore-like structures. Phylogenomics of strain 11kri321 and other Kurthia strains reveals little similarity to genetic determinants of sporulation known from endosporulating Bacilli. However, morphological hallmarks of endosporulation were observed in two of the four Kurthia strains tested, resulting in spore-like structures (cryptospores). In contrast to classic endospores, these cryptospores did not protect against heat or UV damage and successive sub-culturing led to the loss of the cryptosporulating phenotype. Our findings imply that a cryptosporulation phenotype may have been prevalent and subsequently lost by laboratory culturing in other Firmicutes currently considered as asporogenic. Cryptosporulation might thus represent an ancestral but unstable and adaptive developmental state in Firmicutes that is under selection under harsh environmental conditions.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

Examination of Genetic Control Elements in the Phototrophic Firmicute Heliomicrobium modesticaldum

Heliomicrobium modesticaldum has been used as a model organism for the Heliobacteria, the only phototrophic family in the Firmicutes. It is a moderately thermophilic anoxygenic phototrophic bacterium that is capable of fermentative growth in the dark. The genetic manipulation of H. modesticaldum is still in its infancy. Methods to introduce genes through the use of exogenous plasmids and to delete genes from the chromosome through the use of the native CRISPR/Cas system have been developed in the last several years. To expand our genetic toolkit, it was necessary to control gene expression. In this study, we analyzed constitutive and inducible promoters developed for clostridia for their use in H. modesticaldum and further tested two reporters, adhB and lacZ, as indicators of promoter strength. Alcohol dehydrogenase (AdhB) was unsuitable as a reporter in this species due to high endogenous activity and/or low activity of the reporter, but a thermostable LacZ worked well as a reporter. A set of constitutive promoters previously reported to work in Clostridium thermocellum was found to be reliable for controlling the expression of the lacZ reporter gene in H. modesticaldum at a range of activities spanning an order of magnitude. An anhydrotetracycline-inducible promoter was created by inserting tetO operators into a strong constitutive promoter, but it was not fully repressible. The implementation of a xylose-inducible promoter resulted in complete repression of β-gal in the absence of xylose, and reliable expression tunable through the concentration of xylose added to the culture.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2020

Controls on Iron Reduction and Biomineralization over Broad Environmental Conditions as Suggested by the Firmicutes Orenia metallireducens Strain Z6

Microbial iron reduction is a ubiquitous biogeochemical process driven by diverse microorganisms in a variety of environments. However, it is often difficult to separate the biological from the geochemical controls on bioreduction of Fe(III) oxides. In this study, we investigated the primary driving factor(s) that mediate secondary iron mineral formation over a broad range of environmental conditions using a single dissimilatory iron reducer, Orenia metallireducens strain Z6. A total of 17 distinct geochemical conditions were tested with differing pH (6.5–8.5), temperature (22–50 °C), salinity (2–20% NaCl), anions (phosphate and sulfate), electron shuttle (anthraquinone-2,6-disulfonate), and Fe(III) oxide mineralogy (ferrihydrite, lepidocrocite, goethite, hematite, and magnetite). The observed rates and extent of iron reduction differed significantly with k int between 0.186 and 1.702 mmol L –1 day –1 and Fe(II) production ranging from 6.3% to 83.7% of the initial Fe(III). Using X-ray absorption and scattering techniques (EXAFS and XRD), we identified and assessed the relationship between secondary minerals and the specific environmental conditions. It was inferred that the observed bifurcation of the mineralization pathways may be mediated by differing extents of Fe(II) sorption on the remaining Fe(III) minerals. These results expand our understanding of the controls on biomineralization during microbial iron reduction and aid the development of practical applications.

54 ENVIRONMENTAL SCIENCES↗
DOE OSTI2023

A comparative genome analysis of the Bacillota ( Firmicutes ) class Dehalobacteriia

Dehalobacterium formicoaceticum is recognized for its ability to anaerobically ferment dichloromethane (DCM), and a catabolic model has recently been proposed. D. formicoaceticum is currently the only axenic representative of its class, the Dehalobacteriia, according to the Genome Taxonomy Database. However, substantial additional diversity has been revealed in this lineage through culture-independent exploration of anoxic habitats. Here we performed a comparative analysis of 10 members of the Dehalobacteriia, representing three orders, and infer that anaerobic DCM degradation appears to be a recently acquired trait only present in some members of the order Dehalobacteriales. Inferred traits common to the class include the use of amino acids as carbon and energy sources for growth, energy generation via a remarkable range of putative electron-bifurcating protein complexes and the presence of S-layers. The ability of D. formicoaceticum to grow on serine without DCM was experimentally confirmed and a high abundance of the electron-bifurcating protein complexes and S-layer proteins was noted when this organism was grown on DCM. We suggest that members of the Dehalobacteriia are low-abundance fermentative scavengers in anoxic habitats.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2020

Identification of key bacterial populations affecting early embryonic development in cattle uterus

Abstract Superovulation is an important animal breeding biotechnology, while the quality of embryos obtained from superovulation is unstable in cattle. The relationship between the microorganisms in the cattle uterus and embryo qualities was determined to identify the key bacterial populations affecting early embryonic development. A total of 10 Xia Nan cows underwent superovulation, we collected cervical mucus and flush samples to investigated by 16S rDNA sequencing. Results showed that there were abundant microorganisms in cervical mucus, but no obvious relationship with the quality of embryos. The clustering results of flush samples were consistent with the grouping of embryo quality. Proteobacteria accounted for more than 95% of the total bacterial community in group A with the best embryo quality (qualified embryo ratio above 0.8), and as embryo quality decreased, the Proteobacteria proportion also decreased. In contrast to the proportion of Proteobacteria, the proportions of Firmicutes and Bacteroidetes significantly increased as embryo quality decreased. For group C with the worst embryo quality, the proportions of Firmicutes and Bacteroidetes increased to 4.7 times and 12.3 times of group A, respectively. These results showed that the quantities and proportions of Firmicutes and Bacteroidetes may be related to early embryonic development in cattle.

Lyu, Chenchen↗
DOE OSTI2021

Structure of the Extracellular Region of the Bacterial Type VIIb Secretion System Subunit EsaA

Gram-positive bacteria use type VII secretion systems (T7SSs) to export effector proteins that manipulate the physiology of nearby prokaryotic and eukaryotic cells. Several mycobacterial T7SSs have established roles in virulence. By contrast, the genetically distinct T7SSb pathway found in Firmicutes bacteria more often functions to mediate bacterial competition. A lack of structural information on the T7SSb has limited the understanding of effector export by this protein secretion apparatus. Here, we present the 2.4 angstrom crystal structure of the extracellular region of the T7SSb subunit EsaA from Streptococcus gallolyticus. Our structure reveals that homodimeric EsaA is an elongated, arrow-shaped protein with a surface-accessible "tip", which in some species of bacteria serves as a receptor for lytic bacteriophages. Because it is the only T7SSb subunit large enough to traverse the peptidoglycan layer of Firmicutes, we propose that EsaA plays a critical role in transporting effectors across the entirety of the Gram-positive cell envelope.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

The association between vaginal microbiota disorders and early missed abortion: A prospective study

Abstract Introduction The objective of this study was to explore the association between disordered vaginal microbiota and missed abortion to ascertain potential causes of missed abortion related to vaginal microbiology. Material and methods This study was a prospective cohort study with a comparison group (reference group). Vaginal secretions from the posterior fornix of women in the early stages of pregnancy were collected in the Xiangya Third Hospital of Central South University in Changsha, Hunan, China, from November 2018 to November 2019. A total of 54 cases of missed abortion (case group) and 50 cases of normal pregnancy requiring induced abortion (reference group) were analyzed. Bacterial DNA was extracted, hybridized with gene‐specific primers, and then detected using a bacterial chip. The composition and relative abundance of vaginal microbiota in the two groups were compared using α‐diversity analysis, β‐diversity analysis, and the linear discriminant analysis effect size method. Results The α‐diversity analysis showed that the Simpson index of the case group was lower than that of the reference group, whereas the Shannon index in the case group was higher. The relative abundance of Firmicutes in the case group (42.52%) was lower than in the reference group (51.03%, p < 0.05), as was the relative abundance of Lactobacillus (case group 16.51%, reference group 23.00%; p < 0.05). Interestingly, levels of Mycoplasma genitalium and Ureaplasma were lower in the case group ( p < 0.05). The relative abundance of Lactobacillus crispatus , Lactobacillus jensenii , and Lactobacillus gasseri was also significantly lower in the case group than in the reference group ( p < 0.05). The pathways enriched in the case group were predominantly related to metabolism, whereas the genetics‐related pathways were predominantly enriched in the reference group. Conclusions Bacteria are more diverse and unevenly distributed in patients with missed abortion. Decreases in the proportion of vaginal Lactobacillus and changes in Lactobacillus species in these patients may increase the chance of genital tract pathogenic bacterial infection. To our knowledge, our study was the first to observe that a decrease of Firmicutes levels in the vaginal microbiota might impair energy metabolism and have an association with missed abortion.

Sun, Dan↗
DOE OSTI2026

Data for Immediate Impacts of Soybean Cover Crop on Bacterial Community Composition and Diversity in Soil Under Long-Term Saccharum Monoculture

Saccharum yield decline results from long-term monoculture practices. Changes in cropping management can improve soil health and productivity. Below-ground bacterial community diversity and composition across soybean (Glycine max (L.) Merr) cover crop, Saccharum monoculture (30+ year) and fallowed soil were determined. Near full length (~1,400 base pairs) of 16S rRNA gene sequences were extracted from the rhizospheres of sugarcane and soybean and fallowed soil were compared. Higher soil bacterial diversity was observed in the soybean cover crop than sugarcane monoculture across all measured indices (observed operationational taxonomic units, Chao1, Shannon, reciprocal Simpson and Jackknife). Acidocateria, Proteobacteria, Bacteroidetes and Planctomycetes were the most abundant bacterial phyla across the treatments. Indicator species analysis identified nine indicator phyla. Planctomycetes, Armatimonadetes and candidate phylum FBP were associated with soybean; Proteobacteria and Firmicutes were linked with sugarcane and Gemmatimonadetes, Nitrospirae, Rokubacteria and unclassified bacteria were associated with fallowed soil. Non-metric multidimensional scaling analysis showed distinct groupings of bacterial operational taxonomic units (97% identity) according to management system (soybean, sugarcane or fallow) indicating compositional differences among treatments. This is confirmed by the results of the multi-response permutation procedures (A = 0.541, p = 0.00045716). No correlation between soil parameters and bacterial community structure was observed according to Mantel test (r = 211865, p = 0.14). Use of soybean cover-crop fostered bacterial diversity and altered community structure. This indicates cover crops could have a restorative effect and potentially promote sustainability in long-term Saccharum production systems.

Field Data↗
DOE OSTI2021

Diverse Profile of Fermentation Byproducts From Thin Stillage

The economy of biorefineries is influenced not only by biofuel production from carbohydrates but also by the production of valuable compounds from largely underutilized industrial residues. Currently, the demand for many chemicals that could be made in a biorefinery, such as succinic acid (SA), medium-chain fatty acids (MCFAs), and lactic acid (LA), is fulfilled using petroleum, palm oil, or pure carbohydrates as raw materials, respectively. Thin stillage (TS), the residual liquid material following distillation of ethanol, is an underutilized coproduct from the starch biofuel industry. This carbon-rich material has the potential for chemical upgrading by microorganisms. Here, we explored the formation of different fermentation products by microbial communities grown on TS using different bioreactor conditions. At the baseline operational condition (6-day retention time, pH 5.5, 35°C), we observed a mixture of MCFAs as the principal fermentation products. Operation of a bioreactor with a 1-day retention time induced an increase in SA production, and a temperature increase to 55°C resulted in the accumulation of lactic and propionic acids. In addition, a reactor operated with a 1-day retention time at 55°C conditions resulted in LA accumulation as the main fermentation product. The prominent members of the microbial community in each reactor were assessed by 16S rRNA gene amplicon sequencing and phylogenetic analysis. Under all operating conditions, members of the Lactobacillaceae family within Firmicutes and the Acetobacteraceae family within Proteobacteria were ubiquitous. Members of the Prevotellaceae family within Bacteroidetes and Lachnospiraceae family within the Clostridiales order of Firmicutes were mostly abundant at 35°C and not abundant in the microbial communities of the TS reactors incubated at 55°C. The ability to adjust bioreactor operating conditions to select for microbial communities with different fermentation product profiles offers new strategies to explore and compare potentially valuable fermentation products from TS and allows industries the flexibility to adapt and switch chemical production based on market prices and demands.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

A metagenome-level analysis of a microbial community fermenting ultra-filtered milk permeate

Fermentative microbial communities have the potential to serve as biocatalysts for the conversion of low-value dairy coproducts into renewable chemicals, contributing to a more sustainable global economy. To develop predictive tools for the design and operation of industrially relevant strategies that utilize fermentative microbial communities, there is a need to determine the genomic features of community members that are characteristic to the accumulation of different products. To address this knowledge gap, we performed a 282-day bioreactor experiment with a microbial community that was fed ultra-filtered milk permeate, a low-value coproduct from the dairy industry. The bioreactor was inoculated with a microbial community from an acid-phase digester. A metagenomic analysis was used to assess microbial community dynamics, construct metagenome-assembled genomes (MAGs), and evaluate the potential for lactose utilization and fermentation product synthesis of community members represented by the assembled MAGs. This analysis led us to propose that, in this reactor, members of the Actinobacteriota phylum are important in the degradation of lactose, via the Leloir pathway and the bifid shunt, and the production of acetic, lactic, and succinic acids. In addition, members of the Firmicutes phylum contribute to the chain-elongation-mediated production of butyric, hexanoic, and octanoic acids, with different microbes using either lactose, ethanol, or lactic acid as the growth substrate. We conclude that genes encoding carbohydrate utilization pathways, and genes encoding lactic acid transport into the cell, electron confurcating lactate dehydrogenase, and its associated electron transfer flavoproteins, are genomic features whose presence in Firmicutes needs to be established to infer the growth substrate used for chain elongation.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

Geochemistry and metagenomics analyses of bacterial community structure in selected waste dumpsites in Lagos Metropolis, Nigeria

Dumpsites are reservoirs of persistent organic pollutants (POPs) and heavy metals (HMs), constituting environmental hazards to humanity. Autochthonous microorganisms in dumpsites exhibit various degrees of responses to contaminants. Unfortunately, there is a dearth of information on the types and concentration of pollutants and the array of microorganisms in these dumpsites which may play important roles in the metabolism of such pollutants or other community processes. Therefore, determining the microbial community structure in such contaminated sites across a municipality is essential for profiling the taxa that would serve as consensus degraders of the pollutants. In this study, soil samples from three dumpsites (Cele, CS; Solous, SS; and Computer Village, CVS) were characterized for geochemical properties using GC-MS, MP-AES, and other analytical protocols, while the dynamics of bacterial communities were evaluated based on their 16S rRNA gene barcodes. A significant difference in the bacterial communities was observed among the dumpsites in relation to the extent of pollution caused by POPs and HMs. CVS, with the highest HM contamination, was rich in Actinobacteria (41.7%) and Acidobacteria (10.2%), in contrast to CS and SS. Proteobacteria (34.1%) and Firmicutes (20%) were the dominant phyla in CS (highest POP contamination), while Bacteroidetes (45.5%) and Proteobacteria (39.9%) were dominant in SS soil.Bacilluswas the dominant genus in the most polluted dumpsite. Canonical correspondence analysis revealed that polycyclic aromatic hydrocarbons (PAHs) and HMs shaped the structure of the bacterial operational taxonomic units (OTUs) in the most polluted dumpsite. Out of a total of 706 OTUs, 628 OTUs exhibited a significant correlation (>50%) with benzo(b)fluoranthene, azobenzene, dibenzofurans, pyrene, dibenzo(a,l)pyrene, Cu, and Zn. In particular, Proteobacteria (Achromobactersp. andSerratiasp.), Bacteroidetes (Zhouiasp.), and Firmicutes (Bacillussp.) were suggested to be pivotal to the ecophysiology of dumpsite soils contaminated with POPs and HMs. The results generally underscored the importance of metagenomic and physicochemical analyses of polluted systems in enabling correlations for useful prediction of drivers of such ecosystems. This will further improve our understanding of the metabolic potential and adaptation of organisms in such systems.

Environmental Sciences & Ecology↗
DOE OSTI2022

Effects of antibacterial peptide-producing Bacillus subtilis, gallic acid, and cellulase on fermentation quality and bacterial community of whole-plant corn silage

In the current study, we assessed the effects of antibacterial peptide-producing Bacillus subtilis (BS), gallic acid (GA) and cellulase (CL) on the fermentation quality and bacterial community of various varieties of whole-plant corn silage. Three different varieties of whole-plant corn (Yuqing386, Enxiai298, and Nonghe35) were treated with 0.02% BS (fresh material basis), 0.2% GA (fresh material basis) and 0.02% CL (fresh material basis), after which 45 days of anaerobic fermentation were conducted. With the exception of its low dry matter content, the results showed that Yuqing386’s crude protein, water-soluble carbohydrate, and lactic acid contents were significantly higher than those of the other two corn varieties. However, its acid detergent fiber and cellulose contents were significantly lower than those of the other two corn varieties. Among the three corn variety silages, Yuqing386 had the highest relative abundance of Lactobacillus at the genus level and the biggest relative abundance of Firmicutes at the phylum level. In addition, the three additives markedly enhanced the quantity of dry matter and crude protein as compared to the control group. The application of GA considerably decreased the level of neutral detergent fiber while significantly increasing the content of lactic acid and water-soluble carbohydrates. Even though all additives enhanced the structure of the bacterial community following silage, the GA group experienced the greatest enhancement. On a phylum and genus level, the GA group contains the highest relative abundance of Firmicutes and Lactobacillus , respectively. Overall, of the three corn varieties, Yuqing386 provides the best silage qualities. GA has the biggest impact among the additions employed in this experiment to enhance the nutritional preservation and fermentation quality of whole-plant corn silage.

Zhang, Zhiheng↗
DOE OSTI2022

Effects of high concentrate rations on ruminal fermentation and microbiota of yaks

Ruminal microflora is closely correlated with the ruminant’s diet. However, information regarding the effect of high concentrate diets on rumen microflora in yaks is lacking. In the current study, 24 healthy male yaks were randomly assigned to two groups, each fed with different diets: less concentrate (LC; concentrate: coarse = 40: 60) and high concentrate (HC; concentrate: coarse = 80: 20) diets. Subsequently, a 21-day feeding trial was performed with the yaks, and rumen fluid samples were collected and compared using 16 s rRNA sequencing. The results showed that NH 3 -N, total VFA, acetate, butyrate, isobutyrate, and isovalerate were significantly higher in the HC group than that in the LC group ( p < 0.05), while microbial diversity and richness were significantly lower in the HC group ( p < 0.05). Principal coordinate analysis indicated that rumen microflora was significantly different in LC and HC groups ( p < 0.05). In the rumen, phyla Firmicutes and Bacteroidota were the most abundant bacteria, with Firmicutes being more abundant, and Bacteroidota being less abundant in the HC group than those found in the LC group. Christensenellaceae_R-7_group and Prevotella are the highest abundant ones at the genus level. The relative abundance of Acetitomaculum , Ruminococcus , and Candidatus_Saccharimonas were significantly higher in the HC group than that in the LC group ( p < 0.05), while the relative abundance of Olsenella was significantly lower in the HC group than in the LC group ( p < 0.05). Compared to the LC group, the relative abundance of Prevotella , Ruminococcus , and Candidatus_Saccharimonas was significantly higher in the HC group. The relative abundances of Prevotella , Prevotellaceae_UCG-003 , Olsenella , Ruminococcus , Acetitomaculum , Candidatus_Saccharimonas , and NK4A214_group were correlated with ruminal fermentation parameters ( p < 0.05). Furthermore, PICRUSt 2 estimation indicated that microbial genes associated with valine, leucine, and isoleucine biosynthesis were overexpressed in the rumen microflora of yaks in the HC group ( p < 0.05). Conclusively, our results suggest that high concentrate diets affect the microflora composition and fermentation function in yak rumen. The present findings would provide new insights into the health of yaks under high concentrate feeding conditions and serve as a potent reference for the short-term fattening processes of yaks.

Pang, Kaiyue↗
DOE OSTI2023

BONCAT-FACS-Seq reveals the active fraction of a biocrust community undergoing a wet-up event

Determining which microorganisms are active within soil communities remains a major technical endeavor in microbial ecology research. One promising method to accomplish this is coupling bioorthogonal non-canonical amino acid tagging (BONCAT) with fluorescence activated cell sorting (FACS) which sorts cells based on whether or not they are producing new proteins. Combined with shotgun metagenomic sequencing (Seq), we apply this method to profile the diversity and potential functional capabilities of both active and inactive microorganisms in a biocrust community after being resuscitated by a simulated rain event. We find that BONCAT-FACS-Seq is capable of discerning the pools of active and inactive microorganisms, especially within hours of applying the BONCAT probe. The active and inactive components of the biocrust community differed in species richness and composition at both 4 and 21 h after the wetting event. The active fraction of the biocrust community is marked by taxa commonly observed in other biocrust communities, many of which play important roles in species interactions and nutrient transformations. Among these, 11 families within the Firmicutes are enriched in the active fraction, supporting previous reports indicating that the Firmicutes are key early responders to biocrust wetting. We highlight the apparent inactivity of many Actinobacteria and Proteobacteria through 21 h after wetting, and note that members of the Chitinophagaceae, enriched in the active fraction, may play important ecological roles following wetting. Based on the enrichment of COGs in the active fraction, predation by phage and other bacterial members, as well as scavenging and recycling of labile nutrients, appear to be important ecological processes soon after wetting. To our knowledge, this is the first time BONCAT-FACS-Seq has been applied to biocrust samples, and therefore we discuss the potential advantages and shortcomings of coupling metagenomics to BONCAT to intact soil communities such as biocrust. In all, by pairing BONCAT-FACS and metagenomics, we are capable of highlighting the taxa and potential functions that typifies the microbes actively responding to a rain event.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

Associations between Frequency of Culinary Herb Use and Gut Microbiota

While evidence suggests that culinary herbs have the potential to modulate gut microbiota, much of the current research investigating the interactions between diet and the human gut microbiome either largely excludes culinary herbs or does not assess use in standard culinary settings. As such, the primary objective of this study was to evaluate how the frequency of culinary herb use is related to microbiome diversity and the abundance of certain taxa, measured at the phylum level. In this secondary data analysis of the INCLD Health cohort, we examined survey responses assessing frequency of culinary herb use and microbiome analysis of collected stool samples. We did not observe any associations between frequency of culinary herb use and Shannon Index, a measure of alpha diversity. Regarding the abundance of certain taxa, the frequency of use of polyphenol-rich herbs and herbs with certain quantities of antibacterial compounds was positively associated with Firmicutes abundance, and negatively associated with Proteobacteria abundance. Additionally, the total number of herbs used with high frequency, defined as over three times per week, was also positively associated with Firmicutes abundance, independent of adjustments, and negatively associated with Proteobacteria abundance, after adjusting for dietary factors. Frequency of culinary herb use was not associated with Bacteroidota or Actinobacteria abundance.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2025

Microbial Enrichments Contribute to Characterization Of Desert Tortoise Gut Microbiota

Abstract Desert tortoises play ecologically significant roles, including plant seed dispersal and mineral cycling, and yet little is known about microbial members that are critical to their gut and overall health. Tortoises consume recalcitrant plant material, which their gut microbiota degrades and converts into usable metabolites and nutrients for the tortoise. Findings from tortoise gut microbiomes may translate well into biotechnological applications as these microbes have evolved to efficiently degrade recalcitrant substrates and generate useful products. In this study, we cultivated microbial communities from desert tortoise fecal samples following a targeted anaerobic enrichment for microbes involved in deconstruction and utilization of plant biomass. We employed 16S rRNA amplicon sequencing to compare cultivated communities to initial fecal source material and found high abundances of Firmicutes and Bacteroidota typically associated with biomass deconstruction in all cultivated samples. Significantly decreased microbial diversity was observed in the cultivated microbial communities, yet several key taxa thrived in lignocellulose enrichments, includingLachnospiraceaeandEnterococcus. Additionally, cultivated communities produced short-chain fatty acids under anaerobic conditions, and their growth and metabolic output provide evidence of their viability in the initial fecal communities. Overall, this study adds to the limited understanding of reptilian herbivore microbiota, and offers a path towards biotechnological translation based on the ability of the cultivated communities to convert lignocellulose directly to acetate, propionate, and butyrate.

Environmental Sciences & Ecology↗
DOE OSTI2024

A Comparative Metagenomic Analysis of Specified Microorganisms in Groundwater for Non-Sterilized Pharmaceutical Products

In pharmaceutical manufacturing, ensuring product safety involves the detection and identification of microorganisms with human pathogenic potential, including Burkholderia cepacia complex (BCC), Escherichia coli, Pseudomonas aeruginosa, Salmonella enterica, Staphylococcus aureus, Clostridium sporogenes, Candida albicans, and Mycoplasma spp., some of which may be missed or not identified by traditional culture-dependent methods. In this study, we employed a metagenomic approach to detect these taxa, avoiding the limitations of conventional cultivation methods. We assessed the groundwater microbiome’s taxonomic and functional features from samples collected at two locations in the spring and summer. All datasets comprised 436–557 genera with Proteobacteria, Bacteroidota, Firmicutes, Actinobacteria, and Cyanobacteria accounting for > 95% of microbial DNA sequences. The aforementioned species constituted less than 18.3% of relative abundance. Escherichia and Salmonella were mainly detected in Hot Springs, relative to Jefferson, while Clostridium and Pseudomonas were mainly found in Jefferson relative to Hot Springs. Multidrug resistance efflux pumps and BlaR1 family regulatory sensor-transducer disambiguation dominated in Hot Springs and in Jefferson. These initial results provide insight into the detection of specified microorganisms and could constitute a framework for the establishment of comprehensive metagenomic analysis for the microbiological evaluation of pharmaceutical-grade water and other non-sterile pharmaceutical products, ensuring public safety.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2021

Assessing Microbial Communities Related to Mercury Transformations in Contaminated Streambank Soils

In nature, the bioaccumulative potent neurotoxin methylmercury (MeHg) is produced from inorganic mercury (Hg) predominantly by anaerobic microorganisms. Hg-contaminated soils are a potential source of MeHg due to microbial activity. We examine streambank soils collected from the contaminated East Fork Poplar Creek (EFPC) in Tennessee, USA, where seasonal variations in MeHg levels have been observed throughout the year, suggesting active microbial Hg methylation. In this study, we characterized the microbial community in contaminated bank soil samples collected from two locations over a period of one year and compared the results to soil samples from an uncontaminated reference site with similar geochemistry (n = 12). Microbial community composition and diversity were assessed by 16S rRNA gene amplicon sequencing. Furthermore, to isolate potential methylators from soils, enrichment cultures were prepared using selective media. A set of three clade-specific primers targeting the gene hgcA were used to detect Hg methylators among the δ-Proteobacteria in EFPC bank soils across all seasons. Two families among the δ-Proteobacteria that have been previously associated with Hg methylation, Geobacteraceae and Syntrophobacteraceae, were found to be predominant with relative abundances of 0.13% and 4.0%, respectively. However, in soil enrichment cultures, Firmicutes were predominant among families associated with Hg methylation. Specifically, Clostridiaceae and Peptococcaceae and their genera Clostridium and Desulfosporosinus were among the ten most abundant genera with relative abundances of 2.6% and 1.7%, respectively. These results offer insights into the role of microbial communities on Hg transformation processes in contaminated bank soils in EFPC. Identifying the biogeochemical drivers of MeHg production is critical for future remediation efforts.

54 ENVIRONMENTAL SCIENCES↗