Bacterial 16S rRNA Gene Copies Research Articles

Simulated soil erosion predominantly affects fungal abundance in the rapeseed rhizosphere

Eroded agricultural soils have reduced soil organic carbon (SOC) levels that may affect the plant-microbiome interactions in the rhizosphere. We explored the impact of simulated erosion on major microbial groups in a pot experiment with rapeseed (Brassica napus L.) grown on arable soil with the potential to capture SOC. An erosion gradient was simulated by admixture (0%, 12%, 24%) of subsoil horizon (Bt) to topsoil (Ap) material. Rapeseed plants were pulse-labeled with 14CO2 at three growth stages and two soil compartments (bulk and rhizosphere soil) were sampled. Fungal ITS copy numbers were consistently higher in the rhizosphere and decreased with progressing plant growth stages. A significant increase of bacterial 16S rRNA gene copies in the rhizosphere only occurred at flowering. A response of fungal abundance to subsoil admixture was found detectable based on fungi:Bacteria and fungi:Archaea ratio at flowering. Archaea were neither affected by soil compartment nor subsoil admixture. 14C activity of microbial biomass, an indicator for relative input of freshly assimilated C into soil microbiome, was impacted by growth stage and compartment and decreased with ongoing growth stage. During the rosette growth stage, the 14C activity of the microbial biomass was elevated in the rhizosphere of the eroded soil indicating a plant response to the erosion factor. Our experiment revealed a compositional separation of the fungal community along the simulated erosion gradient and a selection of fungi for the two different soil compartments at flowering. Olpidimycetes, Fusarium and Rhizopus and putative pathogens were enriched in the rhizosphere at flowering. Fungi may have a competitive advantage in the rhizosphere of strongly eroded and nutrient diluted soils due to ecological adaptation and morphological traits i.e. hyphae that can bypass soil areas with low nutrient availability.

Cover crop monocultures and mixtures enhance bacterial abundance and functionality in the maize root zone.

Cover cropping is an effective method to protect agricultural soils from erosion, promote nutrient and moisture retention, encourage beneficial microbial activity, and maintain soil structure. Re-utilization of winter cover crop root channels by maize roots during summer allows the cash crop to extract resources from distal regions in the soil horizon. In this study, we investigated how cover cropping during winter followed by maize (Zea mays L.) during summer affects the spatiotemporal composition and function of the bacterial communities in the maize rhizosphere and surrounding soil samples using quantitative polymerase chain reaction (PCR), 16S ribosomal ribonucleic acid (rRNA) gene amplicon sequencing, and metaproteomics. We found that the bacterial community differed significantly among cover crop species, soil depths, and maize growth stages. Bacterial abundance increased in reused root channels, and it continued to increase as cover crop diversity changed from monocultures to mixtures. Mixing Fabaceae with Brassicaceae or Poaceae enhanced the overall contributions of several steps of the bacterial carbon and nitrogen cycles, especially glycolysis and the pentose phosphate pathway. The deeper root channels of Fabaceae and Brassicaceae as compared to Poaceae corresponded to higher bacterial 16S rRNA gene copy numbers and improved community presence in the subsoil regimes, likely due to the increased availability of root exudates secreted by maize roots. In conclusion, root channel reuse improved the expression of metabolic pathways of the carbon and nitrogen cycles and the bacterial communities, which is beneficial to the soil and to the growing crops.

Hyperarid soil microbial community response to simulated rainfall.

The exceptionally long and protracted aridity in the Atacama Desert (AD), Chile, provides an extreme, terrestrial ecosystem that is ideal for studying microbial community dynamics under hyperarid conditions. Our aim was to characterize the temporal response of hyperarid soil AD microbial communities to ex situ simulated rainfall (5% g water/g dry soil for 4 weeks) without nutrient amendment. We conducted replicated microcosm experiments with surface soils from two previously well-characterized AD hyperarid locations near Yungay at 1242 and 1609 masl (YUN1242 and YUN1609) with distinct microbial community compositions and average soil relative humidity levels of 21 and 17%, respectively. The bacterial and archaeal response to soil wetting was evaluated by 16S rRNA gene qPCR, and amplicon sequencing. Initial YUN1242 bacterial and archaeal 16S rRNA gene copy numbers were significantly higher than for YUN1609. Over the next 4 weeks, qPCR results showed significant increases in viable bacterial abundance, whereas archaeal abundance decreased. Both communities were dominated by 10 prokaryotic phyla (Actinobacteriota, Proteobacteria, Chloroflexota, Gemmatimonadota, Firmicutes, Bacteroidota, Planctomycetota, Nitrospirota, Cyanobacteriota, and Crenarchaeota) but there were significant site differences in the relative abundances of Gemmatimonadota and Chloroflexota, and specific actinobacterial orders. The response to simulated rainfall was distinct for the two communities. The actinobacterial taxa in the YUN1242 community showed rapid changes while the same taxa in the YUN1609 community remained relatively stable until day 30. Analysis of inferred function of the YUN1242 microbiome response implied an increase in the relative abundance of known spore-forming taxa with the capacity for mixotrophy at the expense of more oligotrophic taxa, whereas the YUN1609 community retained a stable profile of oligotrophic, facultative chemolithoautotrophic and mixotrophic taxa. These results indicate that bacterial communities in extreme hyperarid soils have the capacity for growth in response to simulated rainfall; however, historic variations in long-term hyperaridity exposure produce communities with distinct putative metabolic capacities.

Distribution of microbial abundance in long-term copper contaminated soils from Topolnitsa-Pirdop valley, Southern Bulgaria

This study presents the distribution of bacterial and fungal abundances in long-term copper (Cu) contaminated soils in Topolnitsa-Pirdop valley – a highly industrialized zone with a number of mines and processing plants for copper and other non-ferrous metals. The bacterial (16S rRNA gene copies) and fungal (ITS rRNA gene copies) were estimated using quantitative PCR technique in five topsoils, differently Cu contaminated (ranging from 28.05 to 198.9 mg kg-1). Bacterial abundance varied in a range of 1.68 × 1011 to 3.24 × 101116S rRNA genes, whereas fungi amounted from 1.95 × 108 to 6.71 × 108 ITS rRNA genes. Fungal and bacterial abundances were significantly (fungi) and insignificantly (bacteria) influenced by Cu contamination. The fungal/bacterial ratio related negatively with soil Cu, which shifted microbial communities’ structure towards bacterial dominance. Since the ratio between bacteria and fungi are vital in explaining many soil functions, the calculated changes in this ratio indicated deterioration in soil quality, being of primary importance for plant production.

Response of microbial communities to nutrient removal in coastal sediment by using ecological concrete.

Ecological concrete (eco-concrete) is a kind of environment-friendly material with porous characteristics. In this study, the eco-concrete was used to remove the total nitrogen (TN), total phosphorus (TP), and total organic carbon (TOC) in marine coastal sediment. The bacterial communities in sediment and on eco-concrete surface were also investigated by using high-throughput sequencing and quantitative PCR of 16S rRNA gene. We found that the mean removal efficiencies of TN, TP, and TOC in treatment group were 8.3%, 8.4%, and 12.3% after 28days. The bacterial community composition in the treatment group was significantly different from that in the control group on day 28. In addition, the bacterial community composition on eco-concrete surface was slightly different from that in sediment, and the copy numbers of 16S rRNA gene were higher on eco-concrete surface than in sediment. The types of eco-concrete aggregates (gravel, pebble, and zeolite) also had effects on the bacterial community composition and 16S rRNA gene copy numbers. Furthermore, we found the abundant genus Sulfurovum increased significantly on eco-concrete surface in the treatment group after 28days. Bacteria belonging to this genus were found having denitrification ability and were commonly detected in bioreactors for nitrate removal. Overall, our study expands the application scopes of eco-concrete and suggests that the bacterial communities in eco-concrete can potentially enhance the removal efficiency of nutrients in coastal sediment.

Progressive belowground soil development associated with sustainable plant establishment during copper mine waste revegetation

Critical to the environmental sustainability of hard rock mining is the reclamation of disturbed lands following mine closure through revegetation. Improved understanding of associations between above- and belowground processes that characterize successful plant establishment is critical to the implementation of more efficient revegetation strategies for nutrient-poor mine waste materials. The specific objective of this five-year temporal study was to identify progressive biotic and abiotic indicators of primary soil development on mine waste rock (WR) on a slope hydroseeded with native plant species, and to quantify comparative effects of plant lifeform on soil development. Aboveground plant diversity and belowground substrate properties were measured annually at 67 m intervals along transects following the slope contour. Seeded WR was compared to unseeded WR and the adjacent native ecosystem. A temporal increase in WR microbial biomass was observed in seeded WR relative to unseeded areas. Microbial community analysis found the unseeded WR to be defined by oligotrophic microbes, whereas targeted grass and shrub root zone samples demonstrated significant increases in specific cellulose and lignin degrading and N-cycling phylotypes. More extensive chemical and biological fertility development was observed in shrub root zones relative to grass. Ten chemical and biological indicators increased significantly in shrub WR relative to unseeded WR, whereas grass WR was only enriched in bacterial 16S rRNA gene copy number/g substrate and bacterial/archaeal and fungal diversity. In addition, the shrub root zone had significantly higher nitrogen-cycling potential than grass root zones or unseeded WR. Thus, both grasses and shrubs improve belowground WR development; however, shrub establishment had greater fertility outcomes. Concurrent belowground fertility development is critical to sustainable plant establishment. Coupled evaluation of above- and belowground metrics provides an improved quantitative assessment of revegetation progress and a valuable tool to guide management decisions.

How do biochar size fractions and organic fertilizers interactively influence nitrous oxide emission from a tropical vertisol?

AbstractBackgroundNitrous oxide (N2O) emission from agriculture is increasing alarmingly due to intensive application of inorganic and organic fertilizers. Recently, biochar has been identified as a promising additive to improve agriculture by enhancing soil function and mitigate greenhouse gas emission. However, it is unclear how biochar of different size fractions influences N2O emission from agricultural soil.AimsThe current experiment aims to understand how the size of biochar (BC) and organic fertilizers interactively influence N2O emission from a tropical vertisol.MethodsBC was prepared using pigeon pea (Cajanus cajan) stalks and further processed to obtain two size fractions (<0.25 mm or 0.25–2.00 mm). Organic fertilizers (vermicompost [VC], poultry manure [PM], or farmyard manure [FYM]) and BC were added to soil to evaluate N2O emission potential. BC was added to soil at 10% (w/w), whereas the organic fertilizers were added at 80 kg N ha–1. Emission of CO2 and the abundance of 16S rRNA and amoA gene copies were estimated after incubation period. The interactive effect of BC size fractions and organic fertilizers were statistically evaluated.ResultsBoth BC and organic fertilizers stimulated N2O emission in soil. BC of larger size stimulated N2O emission (µg N2O produced g–1 soil day–1) more than smaller size. Of the three organic fertilizers, PM resulted highest N2O (0.380) emission followed by FYM (0.240) and VC (0.210). BC (0.25 mm) + PM produced least N2O. Abundance of heterotrophic bacterial 16S rRNA gene copies and ammonia‐oxidizing bacterial (AOB) amoA gene copies were highest in PM + BC and lowest in control. Significant relation (p < 0.0001) existed among N2O emission, CO2 emission, and microbial abundance.ConclusionsBC of small size fraction along with organic fertilizers can be an effective strategy to mitigate N2O emission from tropical vertisol.

Exploring the effect of a microencapsulated citrus essential oil on in vitro fermentation kinetics of pig gut microbiota.

Essential oils (EOs) have emerged as a potential alternative to antibiotics in pig breeding due to their antimicrobial properties. Citrus EOs, a common by-product of the orange juice industry, can be an interesting alternative from a financial perspective due to their huge offer in the global market. Thus, the effect of a citrus EO, and specifically different formulations of Brazilian Orange Terpenes (BOT), on pig gut microbiota was evaluated by means of an in vitro fermentation model simulating different sections of the pig gut (stomach, ileum, and colon). Treatments consisted in: BOT in its unprotected form (BOT, 1.85 and 3.70 mg/mL), microencapsulated BOT (MBOT, 3.50 and 7.00 mg/mL), colistin (2 μg/mL), and a control. BOT and MBOT altered in a similar way the total bacterial 16S rRNA gene copies in the stomach only from 18 h of incubation onwards, and no metabolite production in terms of short-chain fatty acids (SCFAs) was detected. In ileal and colonic fermentations, BOT and MBOT affected ileal and colonic microbiota in terms of total bacterial 16S rRNA gene copies, reduced phylogenetic diversity, and altered composition (p < 0.05) as evidenced by the significant reduction of certain bacterial taxa. However, more pronounced effects were found for MBOT, indicating its higher antimicrobial effects compared to the unprotected BOT, and suggesting that the antibacterial efficiency of the unprotected BOT was probably enhanced by microencapsulation. Furthermore, MBOT stimulated lactate production in ileal fermentations and greatly stimulated overall SCFA production in colonic fermentations. This indicates that besides the shifts in ileal and colonic microbiota by the delivered EO (BOT), the wall material of microcapsules (chitosan/modified starch) might have worked as an additional carbon source with prebiotic functioning, stimulating growth and metabolic activity (SCFAs) of colonic bacteria.

Bacterial Necromass Is Rapidly Metabolized by Heterotrophic Bacteria and Supports Multiple Trophic Levels of the Groundwater Microbiome.

ABSTRACTPristine groundwater is a highly stable environment with microbes adapted to dark, oligotrophic conditions. Input events like heavy rainfalls can introduce the excess particulate organic matter, including surface-derived microorganisms, thereby disturbing the groundwater microbiome. Some surface-derived bacteria will not survive this translocation, leading to an input of necromass to the groundwater. Here, we investigated the effects of necromass addition to the microbial community in fractured bedrock groundwater, using groundwater mesocosms as model systems. We followed the uptake of 13C-labeled necromass by the bacterial and eukaryotic groundwater community quantitatively and over time using a complementary protein-stable and DNA-stable isotope probing approach. Necromass was rapidly depleted in the mesocosms within 4 days, accompanied by a strong decrease in Shannon diversity and a 10-fold increase in bacterial 16S rRNA gene copy numbers. Species of Flavobacterium, Massilia, Rheinheimera, Rhodoferax, and Undibacterium dominated the microbial community within 2 days and were identified as key players in necromass degradation, based on a 13C incorporation of >90% in their peptides. Their proteomes comprised various proteins for uptake and transport functions and amino acid metabolization. After 4 and 8 days, the autotrophic and mixotrophic taxa Nitrosomonas, Limnohabitans, Paucibacter, and Acidovorax increased in abundance with a 13C incorporation between 0.5% and 23%. Likewise, eukaryotes assimilated necromass-derived carbon either directly or indirectly. Our data point toward a fast and exclusive uptake of labeled necromass by a few specialists followed by a concerted action of groundwater microorganisms, including autotrophs presumably fueled by released, reduced nitrogen and sulfur compounds generated during necromass degradation.IMPORTANCE Subsurface microbiomes provide essential ecosystem services, like the generation of drinking water. These ecosystems are devoid of light-driven primary production, and microbial life is adapted to the resulting oligotrophic conditions. Modern groundwater is most vulnerable to anthropogenic and climatic impacts. Heavy rainfalls, which will increase with climate change, can result in high surface inputs into shallow aquifers by percolation or lateral flow. These inputs include terrestrial organic matter and surface-derived microbes that are not all capable to flourish in aquatic subsurface habitats. Here, we investigated the response of groundwater mesocosms to the addition of bacterial necromass, simulating event-driven surface input. We found that the groundwater microbiome responds with a rapid bloom of only a few primary degraders, followed by the activation of typical groundwater autotrophs and mixotrophs, as well as eukaryotes. Our results suggest that this multiphase strategy is essential to maintain the balance of the groundwater microbiome to provide ecosystem services.

Transformation of inherent microorganisms in Wyoming-type bentonite and their effects on structural iron

Bentonite is one of the materials used to construct engineered barriers in high-level radioactive nuclear waste geological disposal with its many advantageous features such as low hydraulic conductivity, self-sealing ability, durability, adsorption and immobilization of metals and radionuclides and reduction of microbial activity. Many of these properties are linked with the bentonite swelling capability. Transformations of indigenous microorganism communities from Wyoming-type bentonite and the Finnish repository site groundwater and their effects on the bentonite structural iron over five years were studied in repository relevant anoxic and oxic slurry conditions. Active sulfate reduction (0.06 nmol mL−1 day−1) was detected in the anoxic microcosm waters after a year, however after two years sulfate reduction was not active anymore. Microbial numbers determined by quantitative PCR in the bentonite slurry of both experiment types supported the finding of decrease of overall microbial activity after a year of incubation that was not maintained anymore by the dissolving organic carbon from the bentonite. Regular electron donor additions (final concentration of 2 mM for formate and acetate each, three times per year) activated the microbiome resulting in increasing numbers of bacterial 16S rRNA gene copies and sulfate reducers (dsrB gene copies) as well as detection of sulfide in the water phase of both experiment types. After 4.9 years the structural iron in the fine portion of the montmorillonite had become completely reduced in all microbial microcosms and minor smectite illitization was detected especially in anoxic microcosms. Dominating bacterial groups at the end of the experiment were mainly known sulfur/sulfate and iron reducers. Archaea and fungi constituted a minor part of the microbiomes. In originally oxic microcosms, the bacterial 16S RNA and dsrB gene copy numbers were lower than in the anoxic experiment but started to significantly increase after the electron donor additions. Microorganisms originating from the repository environment could reduce the bentonite structural iron in a few years to an extent likely to affect the bentonite swelling ability if sufficient amounts of suitable electron donors are available in localized areas where bentonite is not at high density and pressure in the geological disposal.

Subacute ruminal acidosis phenotypes in periparturient dairy cows differ in ruminal and salivary bacteria and in the in vitro fermentative activity of their ruminal microbiota

Both ruminal microbial structure and functionality might play a role in inter-individual variation in susceptibility for subacute rumen acidosis (SARA) observed in dairy cows. The aims of this study were to determine whether differences between cows with distinct SARA susceptibility were reflected in distinct (1) ruminal microbial communities, (2) salivary bacterial communities, and (3) fermentative capacity of ruminal microbiota assessed in vitro. To test this hypothesis, rumen samples were collected via an esophageal tube on 21 d postpartum from 38 multiparous Holstein cows, which were classified into 4 groups differing in median and mean time of reticular pH below 6 as well as area under the curve of pH below 6.0. During the 21 d postpartum, all cows within a group fulfilled following criteria: susceptible (S, n = 10; mean or median ≥180 min/d), moderately susceptible (MS, n = 7; 60 min/d < mean time of pH below 6 < 180 min/d, and median time of pH below 6 <180 min/d), moderately unsusceptible (MU, n = 11; 10 min/d < mean < 60 min/d, and median time of pH below 6 ≤30 min/d), or unsusceptible (U, n = 10; median = 0 min/d, and mean <10 min/d). Groups did not differ in total daily dry matter intake nor in total, roughage, or concentrate intake during daily 6-h time intervals. Rumen bacterial α-diversity did not differ among groups, but β-diversity varied and bacterial 16S rRNA gene copy numbers were lower in S compared with U cows. The relative abundance of genera Streptococcus, Sharpea, Prevotellaceae_YAB2003, Succinivibrionaceae_UCG-001, Ruminococcus, and Ruminococcaceae_UCG-001 were higher in S compared with U cows. In contrast, Lachnospiraceae_ND3007 and Oscillospiraceae_V9D2013 were more abundant in U cows. Although pH-associated, inter-animal differences were also observed in the salivary bacteria, common differences in ruminal and salivary bacterial genera were limited. The functionality of the rumen microbiota was evaluated in vitro through exposure of the microbial inoculum of S and U cows to an anaerobic buffer at pH 5.8 and 6.8, in the presence of sterile supernatant of their own and of dry cows' rumen fluid (2 × 2 design). Generally, the S inoculum produced more volatile fatty acids, except at low pH with dry cows' supernatant, where volatile fatty acid production was completely impaired and lactate accumulation was highest. Compared with the microbes of U cows, microbes of S cows showed less fermentative activity in situations with 2 stress factors (low pH and an unfamiliar environment, i.e., rumen fluid supernatant of dry cows).

Occurrence of opportunistic pathogens in private wells after major flooding events: A four state molecular survey

Private wells can become contaminated with waterborne pathogens during flooding events; however, testing efforts focus almost exclusively on fecal indicator bacteria. Opportunistic pathogens (OPs), which are the leading cause of identified waterborne disease in the United States, are understudied in private wells. We conducted a quantitative polymerase chain reaction survey of Legionella spp., L. pneumophila, Mycobacterium spp., M. avium, Naegleria fowleri, and shiga toxin-producing Escherichia coli gene markers and total coliform and E. coli in drinking water supplied by private wells following the Louisiana Floods (2016), Hurricane Harvey (2017), Hurricane Irma (2017), and Hurricane Florence (2018). Self-reported well characteristics and recovery status were collected via questionnaires. Of the 211 water samples collected, 40.3% and 5.2% were positive for total coliform and E. coli, which were slightly elevated positivity rates compared to prior work in coastal aquifers. DNA markers for Legionella and Mycobacterium were detected in 54.5% and 36.5% of samples, with L. pneumophila and M. avium detected in 15.6% and 17.1%, which was a similar positivity rate relative to municipal system surveys. Total bacterial 16S rRNA gene copies were positively associated with Legionella and Mycobacterium, indicating that conditions that favor occurrence of general bacteria can also favor OPs. N. fowleri DNA was detected in 6.6% of samples and was the only OP that was more prevalent in submerged wells compared to non-submerged wells. Self-reported well characteristics were not associated with OP occurrence. This study exposes the value of routine baseline monitoring and timely sampling after flooding events in order to effectively assess well water contamination risks.

Excellent excrement? Frass impacts on a soil's microbial community, processes and metal bioavailability

The commercial rearing of insects is a growing economic sector. Therefore, an assessment was made of the potential of its by-product, frass, to be a soil improver. Essential plant nutrients were extracted (using 0.01 M CaCl2 or Mehlich 3) from frass of mealworms (Tenebrio molitor), black soldier flies (Hermetia illucens) and buffalo worms (Alphitobius diaperinus). A 28-day incubation, in which frass was added to a sandy loam soil at application rates of 2.5% or 5% (w/w), assessed its effects on soil microbial biomass, abundance of bacteria, archaea and fungi, carbon mineralisation and nitrification. In a separate 56-day incubation, the impact of frass on heavy metal bioavailability in an artificially contaminated, carbon-poor substrate was tested. All frass types featured high electrical conductivity, a mildly acidic to neutral pH and C: N ratios between 11 and 16. Black soldier fly frass (BSFF) was richer in extractable ammonium, phosphorus, potassium and magnesium than mealworm frass (MWF) and buffalo worm frass (BWF) but poorer in extractable calcium. All frass types stimulated carbon mineralisation, nitrification, bacterial and archaeal 16S rRNA gene copy numbers, and fungal biomass as determined by ergosterol concentrations. Bacterial and particularly fungal abundances were stimulated by the 5% frass application rate whereas archaeal abundances were greater in the 2.5% application rate regimes. The 2.5% application rate of MWF and BWF led to a profound build-up of soil extractable nitrite. Correspondingly, these treatments featured the highest 16S rRNA gene copy numbers of archaea, a domain encompassing organisms which oxidise ammonium to nitrite. No nitrite was detectable in soil amended with BSFF. The 5% application rates induced microbial biomass growth (as determined by extractable DNA concentrations) only when BSFF was applied. This was possibly due to differences in the frass types' extractable nutrient or labile carbon contents. BSFF and BWF amendment led to significantly higher microbial biomass in a metal-contaminated substrate. This was likely due to frass providing nutrients, energy and reduced metal bioavailability: extractable zinc, copper, cadmium and nickel concentrations fell due to increased metal sorption and complexation. All frass types could be used as ameliorants in metal-contaminated soils, while BSFF shows most promise as an organic fertiliser as its use did not cause soil nitrite build-up.

Vertical Distribution of Bacterial Community in Water Columns of Reservoirs With Different Trophic Conditions During Thermal Stratification

Water eutrophication is a global ecological issue, and thermal stratification of water bodies can enable eutrophication. We examined bacterial communities in the stratified water columns and sediments in two different trophic reservoirs along the Wujiang River using quantitative real-time PCR and high-throughput sequencing. Bacterial 16S rRNA gene copies varied from 3.70 × 107to 5.27 × 108copies/L in the water column of Hongjiadu (HJD) Reservoir (60 m water depth) with slightly stratified variation; while in Wujiangdu (WJD) Reservoir (70 m water depth), bacterial abundance decreased markedly from the surface to the bottom(1.74 × 109to 2.38 × 107copies/L). The vertical distribution patterns of bacteria in both reservoirs resembled those of water Chlorophylla(Chla) concentrations. The abundance was negatively correlated with water depth (D), total nitrogen (TN), nitrate (NO3–-N), and positively correlated with water temperature (T) and dissolved oxygen (DO) level. In contrast, the alpha diversity of bacteria showed the opposite trend in the vertical water column.Proteobacteria,Actinobacteria, andBacteroideteswere the predominant phyla in the water column of both reservoirs. Compared to WJD Reservoir, HJD Reservoir displayed marked vertical spatial difference in bacterial community structure during thermal stratification. In particular,Pseudomonaswas frequently detected at the bottom of the HJD Reservoir. These results were consistent with predictive metagenomic profiling that revealed different vertical functional variation patterns of the bacterial communities in the two reservoirs. The bacterial community structure of HJD Reservoir was associated with water D, ammonium (NH4+-N), nitrite (NO2–-N), and total phosphorus (TP). The community structure of WJD Reservoir was related to water T, Chla, NO3–-N, and TN. The findings highlighted the important roles played by thermal stratification and nutrients in shaping the water bacterial community structure. Additionally, the absolute abundance of water nitrifiers (AOBgene copies) and denitrifiers (narG,nirS,norB, andnosZgene copies) displayed significant vertical differences in the water columns of both reservoirs. Gene copies involved in denitrification were significantly higher than those involved in nitrification. Water phosphorus and nitrogen contents were important variables influencing the absolute abundance of ammonia oxidizers and denitrifying bacteria, respectively. Our study revealed that the emergence of thermal stratification was responsible for the vertical stratification of bacteria in water and affected the bacterial community structure together with nutrients.

Assessment of two DNA extraction kits for profiling poultry respiratory microbiota from multiple sample types.

Characterization of poultry microbiota is becoming increasingly important due to the growing need for microbiome-based interventions to improve poultry health and production performance. However, the lack of standardized protocols for sampling, sample processing, DNA extraction, sequencing, and bioinformatic analysis can hinder data comparison between studies. Here, we investigated how the DNA extraction process affects microbial community compositions and diversity metrics in different chicken respiratory sample types including choanal and tracheal swabs, nasal cavity and tracheal washes, and lower respiratory lavage. We did a side-by-side comparison of the performances of Qiagen DNeasy blood and tissue (BT) and ZymoBIOMICS DNA Miniprep (ZB) kits. In general, samples extracted with the BT kit yielded higher concentrations of total DNA while those extracted with the ZB kit contained higher numbers of bacterial 16S rRNA gene copies per unit volume. Therefore, the samples were normalized to equal amounts of 16S rRNA gene copies prior to sequencing. For each sample type, all predominant bacterial taxa detected in samples extracted with one kit were present in replicate samples extracted with the other kit and did not show significant differences at the class level. However, a few differentially abundant shared taxa were observed at family and genus levels. Furthermore, between-kit differences in alpha and beta diversity metrics at the amplicon sequence variant level were statistically indistinguishable. Therefore, both kits perform similarly in terms of 16S rRNA gene-based poultry microbiome analysis for the sample types analyzed in this study.

Response of soil bacterial communities to high petroleum content in the absence of remediation procedures.

Oil spills are events that frequently lead to petroleum pollution. This pollution may cause stress to microbial communities, which require long adaption periods. Soil petroleum pollution is currently considered one of the most serious environmental problems. In the present work, processes occurring in the bacterial communities of three soil samples with different physicochemical characteristics, artificially polluted with 12% of crude oil, were investigated in 120-day laboratory experiment. It was found that the total petroleum hydrocarbon content did not decrease during this time; however, the proportion of petroleum fractions was altered. Petroleum pollution led to a short-term decrease in the bacterial 16S rRNA gene copy number. On the basis of amplicon sequencing analysis, it was concluded that bacterial community successions were similar in the three soils investigated. Thus, the phyla Actinobacteria and Proteobacteria and candidate TM7 phylum (Saccaribacteria) were predominant with relative abundances ranging from 35 to 58%, 25 to 30%, and 15 to 35% in different samples, respectively. The predominant operational taxonomic units (OTUs) after pollution belonged to the genera Rhodococcus and Mycobacterium, families Nocardioidaceae and Sinobacteraceae, and candidate class ТМ7-3. Genes from the alkIII group encoding monoxygenases were the most abundant compared with other catabolic genes from the alkI, alkII, GN-PAH, and GP-PAH groups, and their copy number significantly increased after pollution. The copy numbers of expressed genes involved in the horizontal transfer of catabolic genes, FlgC, TraG, and OmpF, also increased after pollution by 11-33, 16-63, and 11-71 times, respectively. The bacterial community structure after a high level of petroleum pollution changed because of proliferation of the cells that initially were able to decompose hydrocarbons, and in the second place, because proliferation of the cells that received these catabolic genes through horizontal transfer.

Comparison of the microbial communities of alpacas and sheep fed diets with three different ratios of corn stalk to concentrate.

The objective of this study was to investigate the characteristics of ruminal microbial communities of alpacas (Lama pacos) and sheep (Ovis aries) fed three diets with varying ratios of roughage (corn stalk) to concentrate, 3:7 (LS), 5:5 (MS) and 7:3 (HS). Six alpacas (one-year-old and weighing 29.5±7.1kg) and six sheep (one-year-old and weighing 27.9±2.7kg) were used in this study, in a replicated 3×3 Latin square experiment. Total protozoa concentration was determined under the microscope; total fungi and methanogens were assessed using quantitative polymerase chain reaction and expressed as a percentage of total bacterial 16S rRNA gene copies; bacterial communities were investigated by targeted 16S rRNA gene (V3-V4 region) sequencing. The percentage of fungi was significantly higher in alpacas than in sheep under the LS diet, while the concentration of protozoa was significantly lower in alpacas under HS, MS and LS diets. The alpha diversity including Shannon, Chao l and ACE indices of bacterial communities was higher in alpacas than in sheep, under the LS diet. A total of 299 genera belonging to 22 phyla were observed in the forestomach of alpaca and sheep, with Bacteroidetes and Firmicutes dominating both animal species. Phyla Armatimonadetes and Fusobacteria, as well as 64 genera, were detected only in alpacas, whereas phyla Acidobacteria and Nitrospira, as well as 44 genera, were found only in sheep. The abundance of cellulolytic bacteria, including Butyrivibrio and Pseudobutyrivibrio, was higher in alpacas than in sheep under all three diets. These differences in the forestomach microbial communities partly explained why alpacas displayed a higher poor-quality roughage digestibility, and a lower methane production. Results also revealed that the adverse effects of high-concentrate diets (70%) were lesser in alpacas than in sheep.

Microbial Communities of Stored Product Mites: Variation by Species and Population.

Arthropod-associated microorganisms are important because they affect host fitness, protect hosts from pathogens, and influence the host's ability to vector pathogens. Stored product mites (Astigmata) often establish large populations in various types of food items, damaging the food by direct feeding and introducing contaminants, including their own bodies, allergen-containing feces, and associated microorganisms. Here we access the microbial structure and abundance in rearing diets, eggs, feces fraction, and mite bodies of 16 mite populations belonging to three species (Carpoglyphus lactis, Acarus siro, and Tyrophagus putrescentiae) using quantitative PCR and 16S ribosomal RNA (rRNA) gene amplicon sequencing. The mite microbiomes had a complex structure dominated by the following bacterial taxa (OTUs): (a) intracellular symbionts of the genera Cardinium and Wolbachia in the mite bodies and eggs; (b) putative gut symbionts of the genera Solitalea, Bartonella, and Sodalis abundant in mite bodies and also present in mite feces; (c) feces-associated or environmental bacteria of the genera Bacillus, Staphylococcus, and Kocuria in the diet, mite bodies, and feces. Interestingly and counterintuitively, the differences between microbial communities in various conspecific mite populations were higher than those between different mite species. To explain some of these differences, we hypothesize that the intracellular bacterial symbionts can affect microbiome composition in mite bodies, causing differences between microbial profiles. Microbial profiles differed between various sample types, such as mite eggs, bodies, and the environment (spent growth medium-SPGM). Low bacterial abundances in eggs may result in stochastic effects in parent-offspring microbial transmission, except for the intracellular symbionts. Bacteria in the rearing diet had little effect on the microbial community structure in SPGM and mite bodies. Mite fitness was positively correlated with bacterial abundance in SPGM and negatively correlated with bacterial abundances in mite bodies. Our study demonstrates critical host-microbe interactions, affecting all stages of mite growth and leading to alteration of the environmental microbiome. Correlational evidence based on absolute quantitation of bacterial 16S rRNA gene copies suggests that mite-associated microorganisms are critical for modulating important pest properties of mites by altering population growth.

First insights to the microbial communities in the plant process water of the multi-metal Kevitsa mine

Metallurgical processes demand large quantities of water. However, in many locations, water is becoming scarce and process water recycling is needed. Closing water loops can be challenging due to build-up of flotation chemicals, metal ions and microorganisms in the recycled water affecting the flotation performance. Here, we have characterized the microbial communities over a 2-month period in different locations of the multi-metal Kevitsa mine in Northern Finland, by microbiome sequencing, enumeration of bacteria, archaea and fungi by qPCR, and cultivation. The microbial communities showed high diversity, but were dominated by Alpha- and Gammaproteobacteria. In addition, various fungal taxa were detected, whereas the archaeal taxa were only sparsely detected from the sequence data. The number of bacterial 16S rRNA gene copies in Process water and Ni thickener overflow varied between 0.5–3.3 × 105 mL−1, whereas the Flotation tailings showed two orders of magnitude lower amounts. Fungi were present at 3.0 × 102–8.1 × 104 5.8S rRNA gene copies mL−1 in all samples, while the number of archaea fluctuated between 8.8 × 101–3.2 × 105 16S rRNA gene copies mL−1. The number of all microbial groups were generally lower in September than in August. When tested on 8 different cultivation media, the microorganisms generally responded positively to organic carbon, and were also shown to oxidize thiosulfate, which may indicate that build-up of organic flotation chemicals and sulfur species from the ore may cause the microbial numbers to increase. This study is part of the H2020 ITERAMS project (Grant agreement# 730480), which strives to improve the recycling of water and minimize the environmental impact of mines.

A survey of extended-spectrum beta-lactamase-producing Enterobacteriaceae in urban wetlands in southwestern Nigeria as a step towards generating prevalence maps of antimicrobial resistance.

In many countries, emission of insufficiently treated wastewater into water bodies appears to be an important factor in spreading clinically relevant antimicrobial resistant bacteria. In this study, we looked for the presence of Enterobacteriaceae strains with resistance to 3rd generation cephalosporin antibiotics in four urban wetlands in southwestern Nigeria by isolation, whole genome sequencing and qPCR enumeration of marker genes. Genome analysis of multi-drug resistant and potentially pathogenic Escherichia coli isolates (members of the widely distributed ST10 complex) revealed the presence of the extended spectrum beta-lactamase gene blaCTX-M-15 on self-transmissible IncF plasmids. The gene was also present together with a blaTEM-1B gene on self-transmissible IncH plasmids in multi-drug resistant Enterobacter cloacae isolates. A Citrobacter freundii isolate carried blaTEM-1B on an IncR-type plasmid without discernable conjugation apparatus. All strains were isolated from a wetland for which previous qPCR enumeration of marker genes, in particular the ratio of intI1 to 16S rRNA gene copy numbers, had indicated a strong anthropogenic impact. Consistent with the isolation origin, qPCR analysis in this study showed that the blaCTX-M gene was present at an abundance of 1x10-4 relative to bacterial 16S rRNA gene copy numbers. The results indicate that contamination of these urban aquatic ecosystems with clinically relevant antibiotic resistant bacteria is substantial in some areas. Measures should therefore be put in place to mitigate the propagation of clinically relevant antimicrobial resistance within the Nigerian aquatic ecosystems.