Selective carbon sources influence the end-products of microbial nitrate respiration

AbstractRespiratory and catabolic pathways are differentially distributed in microbial genomes. Thus, specific carbon sources may favor different respiratory processes. We profiled the influence of 94 carbon sources on the end-products of nitrate respiration in microbial enrichment cultures from diverse terrestrial environments. We found that some carbon sources consistently favor dissimilatory nitrate reduction to ammonium (DNRA/nitrate ammonification) while other carbon sources favor nitrite accumulation or denitrification. For an enrichment culture from aquatic sediment, we sequenced the genomes of the most abundant strains, matched these genomes to 16S rDNA exact sequence variants (ESVs), and used 16S rDNA amplicon sequencing to track the differential enrichment of functionally distinct ESVs on different carbon sources. We found that changes in the abundances of strains with different genetic potentials for nitrite accumulation, DNRA or denitrification were correlated with the nitrite or ammonium concentrations in the enrichment cultures recovered on different carbon sources. Specifically, we found that either L-sorbose or D-cellobiose enriched for a Klebsiella nitrite accumulator, other sugars enriched for an Escherichia nitrate ammonifier, and citrate or formate enriched for a Pseudomonas denitrifier and a Sulfurospirillum nitrate ammonifier. Our results add important nuance to the current paradigm that higher concentrations of carbon will always favor DNRA over denitrification or nitrite accumulation, and we propose that, in some cases, carbon composition can be as important as carbon concentration in determining nitrate respiratory end-products. Furthermore, our approach can be extended to other environments and metabolisms to characterize how selective parameters influence microbial community composition, gene content and function.

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Identifying Hidden Viable Bacterial Taxa in Tropical Forest Soils Using Amplicon Sequencing of Enrichment Cultures

Biology ◽

10.3390/biology10070569 ◽

2021 ◽

Vol 10 (7) ◽

pp. 569

Author(s):

Chakriya Sansupa ◽

Sara Fareed Mohamed Wahdan ◽

Terd Disayathanoowat ◽

Witoon Purahong

Keyword(s):

Bacterial Community ◽

Forest Soil ◽

Culture Media ◽

Enrichment Culture ◽

Sequence Similarity ◽

Illumina Miseq ◽

Amplicon Sequencing ◽

Soil Samples ◽

Enrichment Cultures ◽

Total Community

This study aims to estimate the proportion and diversity of soil bacteria derived from eDNA-based and culture-based methods. Specifically, we used Illumina Miseq to sequence and characterize the bacterial communities from (i) DNA extracted directly from forest soil and (ii) DNA extracted from a mixture of bacterial colonies obtained by enrichment cultures on agar plates of the same forest soil samples. The amplicon sequencing of enrichment cultures allowed us to rapidly screen a culturable community in an environmental sample. In comparison with an eDNA community (based on a 97% sequence similarity threshold), the fact that enrichment cultures could capture both rare and abundant bacterial taxa in forest soil samples was demonstrated. Enrichment culture and eDNA communities shared 2% of OTUs detected in total community, whereas 88% of enrichment cultures community (15% of total community) could not be detected by eDNA. The enrichment culture-based methods observed 17% of the bacteria in total community. FAPROTAX functional prediction showed that the rare and unique taxa, which were detected with the enrichment cultures, have potential to perform important functions in soil systems. We suggest that enrichment culture-based amplicon sequencing could be a beneficial approach to evaluate a cultured bacterial community. Combining this approach together with the eDNA method could provide more comprehensive information of a bacterial community. We expected that more unique cultured taxa could be detected if further studies used both selective and non-selective culture media to enrich bacteria at the first step.

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Assessing Microbial Communities Related to Mercury Transformations in Contaminated Streambank Soils

Water Air & Soil Pollution ◽

10.1007/s11270-020-04978-0 ◽

2021 ◽

Vol 232 (1) ◽

Author(s):

Yazeed Abdelmageed ◽

Carrie Miller ◽

Carrie Sanders ◽

Timothy Egbo ◽

Alexander Johs ◽

...

Keyword(s):

Microbial Community ◽

Microbial Communities ◽

Contaminated Soils ◽

Microbial Community Composition ◽

Inorganic Mercury ◽

Amplicon Sequencing ◽

Soil Samples ◽

Rrna Gene ◽

Enrichment Cultures ◽

Relative Abundances

AbstractIn 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.

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Selective carbon sources influence the end products of microbial nitrate respiration

The ISME Journal ◽

10.1038/s41396-020-0666-7 ◽

2020 ◽

Vol 14 (8) ◽

pp. 2034-2045 ◽

Cited By ~ 1

Author(s):

Hans K. Carlson ◽

Lauren M. Lui ◽

Morgan N. Price ◽

Alexey E. Kazakov ◽

Alex V. Carr ◽

...

Keyword(s):

Carbon Sources ◽

Nitrate Respiration ◽

End Products

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Volatile and Dissolved Organic Carbon Sources Have Distinct Effects on Microbial Activity, Nitrogen Content, and Bacterial Communities in Soil.

10.21203/rs.3.rs-1041309/v1 ◽

2021 ◽

Author(s):

Steven Glynn McBride ◽

Ernest D Osburn ◽

Jane L Lucas ◽

Julia S Simpson ◽

Taylor Brown ◽

...

Keyword(s):

Organic Carbon ◽

Dissolved Organic Carbon ◽

Microbial Activity ◽

Microbial Community Composition ◽

Carbon Sources ◽

Amplicon Sequencing ◽

Moisture Regimes ◽

Available Carbon ◽

Moisture Conditions ◽

Extractable Nitrogen

Abstract Variation in microbial use of soil carbon compounds is a major driver of biogeochemical processes and microbial community composition. Available carbon substrates in soil include both low molecular weight dissolved organic carbon (LMW-DOC), and volatile organic compounds (VOCs). To compare the effects of LMW-DOC and VOCs on soil chemistry and microbial communities under different moisture regimes, we performed a microcosm experiment with five levels of soil water content (ranging from 25-70% water-holding capacity) and five levels of carbon amendment: a no carbon control, two dissolved compounds (glucose and oxalate), and two volatile compounds (methanol and α-pinene). Microbial activity was measured throughout as soil respiration; at the end of the experiment, we measured extractable soil organic carbon and total extractable nitrogen and characterized prokaryotic communities using amplicon sequencing. All C amendments increased microbial activity, and all except oxalate decreased total extractable nitrogen. Likewise, individual phyla responded to specific C amendments – e.g., Proteobacteria increased under addition of glucose, and both VOCs. Further, we observed an interaction between moisture and C amendment, where both VOC treatments had higher microbial activity than LMW-DOC treatments and controls at low moisture. Across moisture and C treatments, we identified that Chloroflexi, Nitrospirae, Proteobacteria, and Verrucomicrobia were strong predictors of microbial activity, while Actinobacteria, Bacteroidetes, and Thaumarcheota strongly predicted soil extractable nitrogen. These results indicate that the type of labile C source available to soil prokaryotes can influence both microbial diversity and ecosystem function and that VOCs may drive microbial functions and composition under low moisture conditions.

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Reduced metagenome sequencing for strain-resolution taxonomic profiles

Microbiome ◽

10.1186/s40168-021-01019-8 ◽

2021 ◽

Vol 9 (1) ◽

Author(s):

Lars Snipen ◽

Inga-Leena Angell ◽

Torbjørn Rognes ◽

Knut Rudi

Keyword(s):

Microbial Community ◽

Microbial Community Composition ◽

Amplicon Sequencing ◽

Good Alternative ◽

Shotgun Sequencing ◽

Least Square ◽

Full Potential ◽

Shotgun Metagenomics ◽

Metagenome Sequencing ◽

Reference Genomes

Abstract Background Studies of shifts in microbial community composition has many applications. For studies at species or subspecies levels, the 16S amplicon sequencing lacks resolution and is often replaced by full shotgun sequencing. Due to higher costs, this restricts the number of samples sequenced. As an alternative to a full shotgun sequencing we have investigated the use of Reduced Metagenome Sequencing (RMS) to estimate the composition of a microbial community. This involves the use of double-digested restriction-associated DNA sequencing, which means only a smaller fraction of the genomes are sequenced. The read sets obtained by this approach have properties different from both amplicon and shotgun data, and analysis pipelines for both can either not be used at all or not explore the full potential of RMS data. Results We suggest a procedure for analyzing such data, based on fragment clustering and the use of a constrained ordinary least square de-convolution for estimating the relative abundance of all community members. Mock community datasets show the potential to clearly separate strains even when the 16S is 100% identical, and genome-wide differences is < 0.02, indicating RMS has a very high resolution. From a simulation study, we compare RMS to shotgun sequencing and show that we get improved abundance estimates when the community has many very closely related genomes. From a real dataset of infant guts, we show that RMS is capable of detecting a strain diversity gradient for Escherichia coli across time. Conclusion We find that RMS is a good alternative to either metabarcoding or shotgun sequencing when it comes to resolving microbial communities at the strain level. Like shotgun metagenomics, it requires a good database of reference genomes and is well suited for studies of the human gut or other communities where many reference genomes exist. A data analysis pipeline is offered, as an R package at https://github.com/larssnip/microRMS.

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Do initial concentration and activated sludge seasonality affect pharmaceutical biotransformation rate constants?

Applied Microbiology and Biotechnology ◽

10.1007/s00253-021-11475-9 ◽

2021 ◽

Author(s):

Tamara J. H. M. van Bergen ◽

Ana B. Rios-Miguel ◽

Tom M. Nolte ◽

Ad M. J. Ragas ◽

Rosalie van Zelm ◽

...

Keyword(s):

Microbial Community ◽

Activated Sludge ◽

Community Composition ◽

Rate Constants ◽

Wastewater Treatment Plants ◽

Microbial Community Composition ◽

Amplicon Sequencing ◽

Rrna Gene ◽

Initial Concentration ◽

Different Seasons

Abstract Pharmaceuticals find their way to the aquatic environment via wastewater treatment plants (WWTPs). Biotransformation plays an important role in mitigating environmental risks; however, a mechanistic understanding of involved processes is limited. The aim of this study was to evaluate potential relationships between first-order biotransformation rate constants (kb) of nine pharmaceuticals and initial concentration of the selected compounds, and sampling season of the used activated sludge inocula. Four-day bottle experiments were performed with activated sludge from WWTP Groesbeek (The Netherlands) of two different seasons, summer and winter, spiked with two environmentally relevant concentrations (3 and 30 nM) of pharmaceuticals. Concentrations of the compounds were measured by LC–MS/MS, microbial community composition was assessed by 16S rRNA gene amplicon sequencing, and kb values were calculated. The biodegradable pharmaceuticals were acetaminophen, metformin, metoprolol, terbutaline, and phenazone (ranked from high to low biotransformation rates). Carbamazepine, diatrizoic acid, diclofenac, and fluoxetine were not converted. Summer and winter inocula did not show significant differences in microbial community composition, but resulted in a slightly different kb for some pharmaceuticals. Likely microbial activity was responsible instead of community composition. In the same inoculum, different kb values were measured, depending on initial concentration. In general, biodegradable compounds had a higher kb when the initial concentration was higher. This demonstrates that Michealis-Menten kinetic theory has shortcomings for some pharmaceuticals at low, environmentally relevant concentrations and that the pharmaceutical concentration should be taken into account when measuring the kb in order to reliably predict the fate of pharmaceuticals in the WWTP. Key points • Biotransformation and sorption of pharmaceuticals were assessed in activated sludge. • Higher initial concentrations resulted in higher biotransformation rate constants for biodegradable pharmaceuticals. • Summer and winter inocula produced slightly different biotransformation rate constants although microbial community composition did not significantly change. Graphical abstract

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Cervicovaginal Microbiome Composition Is Associated with Metabolic Profiles in Healthy Pregnancy

mBio ◽

10.1128/mbio.01851-20 ◽

2020 ◽

Vol 11 (4) ◽

Author(s):

Andrew Oliver ◽

Brandon LaMere ◽

Claudia Weihe ◽

Stephen Wandro ◽

Karen L. Lindsay ◽

...

Keyword(s):

Mass Spectrometry ◽

Reproductive Health ◽

Microbial Communities ◽

Carbon Sources ◽

Amplicon Sequencing ◽

Vaginal Microbiome ◽

Content Type ◽

Healthy Women ◽

Healthy Pregnancy ◽

Tof Ms

ABSTRACT Microbes and their metabolic products influence early-life immune and microbiome development, yet remain understudied during pregnancy. Vaginal microbial communities are typically dominated by one or a few well-adapted microbes which are able to survive in a narrow pH range and are adapted to live on host-derived carbon sources, likely sourced from glycogen and mucin present in the vaginal environment. We characterized the cervicovaginal microbiomes of 16 healthy women throughout the three trimesters of pregnancy. Additionally, we analyzed saliva and urine metabolomes using gas chromatography-time of flight mass spectrometry (GC-TOF MS) and liquid chromatography-tandem mass spectrometry (LC-MS/MS) lipidomics approaches for samples from mothers and their infants through the first year of life. Amplicon sequencing revealed most women had either a simple community with one highly abundant species of Lactobacillus or a more diverse community characterized by a high abundance of Gardnerella, as has also been previously described in several independent cohorts. Integrating GC-TOF MS and lipidomics data with amplicon sequencing, we found metabolites that distinctly associate with particular communities. For example, cervicovaginal microbial communities dominated by Lactobacillus crispatus have high mannitol levels, which is unexpected given the characterization of L. crispatus as a homofermentative Lactobacillus species. It may be that fluctuations in which Lactobacillus dominate a particular vaginal microbiome are dictated by the availability of host sugars, such as fructose, which is the most likely substrate being converted to mannitol. Overall, using a multi-“omic” approach, we begin to address the genetic and molecular means by which a particular vaginal microbiome becomes vulnerable to large changes in composition. IMPORTANCE Humans have a unique vaginal microbiome compared to other mammals, characterized by low diversity and often dominated by Lactobacillus spp. Dramatic shifts in vaginal microbial communities sometimes contribute to the presence of a polymicrobial overgrowth condition called bacterial vaginosis (BV). However, many healthy women lacking BV symptoms have vaginal microbiomes dominated by microbes associated with BV, resulting in debate about the definition of a healthy vaginal microbiome. Despite substantial evidence that the reproductive health of a woman depends on the vaginal microbiota, future therapies that may improve reproductive health outcomes are stalled due to limited understanding surrounding the ecology of the vaginal microbiome. Here, we use sequencing and metabolomic techniques to show novel associations between vaginal microbes and metabolites during healthy pregnancy. We speculate these associations underlie microbiome dynamics and may contribute to a better understanding of transitions between alternative vaginal microbiome compositions.

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An Improved Screening Method for the Detection and Isolation of Escherichia coli 0157:H7 From Meat, Incorporating the 3M Petrifilm™ Test Kit - HEC - for Hemorrhagic Escherichia coli 0157:H7

Journal of Food Protection ◽

10.4315/0362-028x-53.11.936 ◽

1990 ◽

Vol 53 (11) ◽

pp. 936-940 ◽

Cited By ~ 41

Author(s):

ANITA J. G. OKREND ◽

BONNIE E. ROSE ◽

RICHARD MATNER

Keyword(s):

Escherichia Coli ◽

Screening Program ◽

Enrichment Culture ◽

Screening Method ◽

Meat Sample ◽

E Coli ◽

Enrichment Cultures ◽

Poultry Products ◽

Test Kit ◽

Cooked Meat

A screening method was devised incorporating a commercially available reactive disc blot ELISA for Escherichia coli 0157 antigen, into a cultural screening program for the isolation of E. coli 0157:H7 from meat and poultry products. The method includes the inoculation of a raw or cooked meat sample into an enrichment broth, incubation with shaking at 37°C for 6 to 8 h, followed by inoculation of 3M Petrifilm™ E. coli Count plates with dilutions of the enrichment culture. The Petrifilm plates were incubated at 42°C for 18 h and tested for the presence of the 0157 antigen. The enrichment cultures were reincubated static at 35°C after the initial shaken incubation. Isolation was attempted from the positive Petrifilm plates by both a direct picking and streaking method and by the 3M Prompt™ isolation method. Isolation also was attempted from the 24-h enrichment cultures by spread plating serial dilutions on 150 × 15 mm MacConkey sorbitol agar (MSA) and MSA with 5-bromo-4-chloro-3-indoxyl-β-D-glucuronic acid cyclohexylammonium salt (BCIG). This fast and efficient screening procedure identifies negative and presumptive positive samples in 26–28 h. Isolation and confirmation of the presumptive positive isolates require an additional 3 to 4 d.

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Effects of pH and alkalinity on sulfur-denitrification in a biological granular filter

Water Science & Technology ◽

10.2166/wst.1996.0391 ◽

1996 ◽

Vol 34 (1-2) ◽

pp. 355-362 ◽

Cited By ~ 16

Author(s):

Hiroaki Furumai ◽

Hideki Tagui ◽

Kenji Fujita

Keyword(s):

Enrichment Culture ◽

Nitrite Reduction ◽

The Other ◽

Nitrite Accumulation ◽

Ph Dependency ◽

Effects Of Ph ◽

Rapid Removal ◽

Nitrate And Nitrite ◽

The Relationship ◽

Biological Filters

Two laboratory-scale biological filters were operated to investigate the effects of alkalinity and pH on removal of nitrate and nitrite in sulfur denitrification filter processes. The concentration of sodium bicarbonate in the feed media was changed from 120 to 240 mg/l during about 3 months in a filter (Run A). The other filter was initially fed with 300 mg/l and then with 240 mg/l (Run B). The performance of the filter was monitored by measuring pH, nitrate, nitrite, sulfate, alkalinity, and thiosulfate. Nitrate concentration in effluent rapidly decreased to lower levels within several days for both filters after inoculation of enrichment culture of sulfur denitrifiers. However there was a large difference in removal of nitrite. When rapid removal of nitrate took place, nitrite accumulation was observed and remained while the bicarbonate concentration was 120 and 150 mg/l. On the other hand the nitrite accumulation disappeared when more bicarbonate (240 and 300 mg/l) was supplied. The experimental results indicated that the nitrite accumulation was closely related to pH condition and alkalinity level in the filter. The stable data of effluent water quality for 5 cases were collected and the relationship discussed between nitrite concentration and pH in effluents. The relationship indicated a strong pH dependency on nitrite accumulation below pH of 7.4. The pH condition around 7 is not so inhibitory to biological activity. Therefore, the pH within the biofilm would be low enough to suppress the nitrite reduction by sulfur denitrifiers, while the pH in effluent was not in the inhibitory range. It was recommended to keep the pH higher than 7.4 to prevent nitrite accumulation in the sulfur denitrification filter.

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Evaluation of the microbial community structure of potable water samples from occupied and unoccupied buildings using16S rRNA amplicon sequencing

10.1101/2020.07.17.209346 ◽

2020 ◽

Author(s):

Kimothy L Smith ◽

Howard A Shuman ◽

Douglas Findeisen

Keyword(s):

Microbial Community ◽

16S Rrna ◽

Water Samples ◽

Ad Hoc ◽

Microbial Community Composition ◽

Amplicon Sequencing ◽

The Other ◽

Water Usage ◽

16S Rrna Amplicon Sequencing ◽

Oxford Nanopore

AbstractWe conducted two studies of water samples from buildings with normal occupancy and water usage compared to water from buildings that were unoccupied with little or no water usage due to the COVID-19 shutdown. Study 1 had 52 water samples obtained ad hoc from buildings in four metropolitan locations in different states in the US and a range of building types. Study 2 had 36 water samples obtained from two buildings in one metropolitan location with matched water sample types. One of the buildings had been continuously occupied, and the other substantially vacant for approximately 3 months. All water samples were analyzed using 16S rRNA amplicon sequencing with a MinION from Oxford Nanopore Technologies. More than 127 genera of bacteria were identified, including genera with members that are known to include more than 50 putative frank and opportunistic pathogens. While specific results varied among sample locations, 16S rRNA amplicon abundance and the diversity of bacteria were higher in water samples from unoccupied buildings than normally occupied buildings as was the abundance of sequenced amplicons of genera known to include pathogenic bacterial members. In both studies Legionella amplicon abundance was relatively small compared to the abundance of the other bacteria in the samples. Indeed, when present, the relative abundance of Legionella amplicons was lower in samples from unoccupied buildings. Legionella did not predominate in any of the water samples and were found, on average, in 9.6% of samples in Study 1 and 8.3% of samples in Study 2.SynopsisComparison of microbial community composition in the plumbing of occupied and unoccupied buildings during the COVID-19 pandemic shutdown.

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