Community Analysis Research Articles

Insights into anode substrate optimization in bioelectrochemical systems for efficient cathodic lanthanum recovery during wastewater treatment

Wastewater containing lanthanum (La) is a significant global environmental concern, prompting the development of various treatment technologies. Existing methods are often costly, generate secondary waste streams, and prioritize removal over recovery. This study investigated La recovery in bioelectrochemical systems (BESs) using diverse substrates (acetate, lactate, methanol, and glucose) with anaerobic sludge as the inoculum. The results showed that acetate-fueled BESs achieved the highest power density (439.29 mW/m2), a chemical oxygen demand (COD) reduction efficiency of 96.52 ± 1.56 %, and a coulombic efficiency of 35.18 ± 1.86 %. Acetate and lactate-fed biofilms achieved high La recovery rates (98.96 ± 1.64 % and 98.24 ± 2.16 %, respectively) by the 36th hour. After 48 h, La was undetectable in these systems, surpassing the environmental guidelines. In contrast, methanol and glucose-fed systems yielded lower recoveries (80 ± 0.04 % and 32.8 ± 1.4 %, respectively) after 48 h, with final La concentrations remaining above the guideline levels. Acetate also enhanced the production of extracellular polymeric substances (EPS). Microbial community analysis revealed that Desulfovibrio and Geobacter predominated in acetate-fed microbial fuel cells (MFCs), while Klebsiella and Simplicispira were dominant in glucose and methanol-fed MFCs, respectively. Recovered La deposits, identified as La2O3 and La(OH)3, on the cathodic surfaces were confirmed via SEM-EDX, XRD, and XPS analyses. This study demonstrates the potential of BESs for efficient La recovery from wastewater.

Insight into the autochthonous lactic acid bacteria as starter culture for improving the quality of Sichuan radish paocai: Changes in microbial diversity and metabolic profiles

Paocai is a traditional Chinese fermented vegetable product popular in Asian countries. Recently, functional starters were used to control the fermentation process and improve the quality of paocai. In this study, three autochthonous lactic acid bacteria including Lactiplantibacillus plantarum LB6, Lactiplantibacillus pentosus LB3, and Weissella cibaria W51 were selected as starters and the effect of the starters on the fermentation of paocai was investigated. The results suggested that the inoculated fermentation led to a lower nitrite peak and more pronounced changes in pH and total titratable acid in the early stage of fermentation, compared with natural fermentation. Analysis of the flavor compounds indicated that the total content of volatile organic compounds of paocai through natural fermentation was significantly lower than that in inoculated fermentation. As for free amino acids, in the early stage of fermentation, the types and contents of free amino acids in the inoculated fermentation paocai were higher than those in the blank group. In the later stage of fermentation, the contents of amino acids representing umami and sweet tastes were also higher than those in the blank group. The bacterial community analysis showed that Lactobacillus and Lactococcus were the dominant bacteria in both inoculated fermentation and natural fermentation. Then, the correlations among physicochemical properties, microbial community and flavor compounds were revealed, and it was found that the dominant bacteria such as Lactococcus, Leuconostoc, Lactobacillus and Weissella displayed a considerable impact on the physical and chemical properties and flavor of paocai. In addition, the metabolic pathways involved in flavor formation and the abundance of related enzymes were elucidated. The abundance of enzymes involved in generating prephenic acid, 2-methylbutanoic acid, L-lactic acid, D-lactic acid, butanoic acid, etc., and in the pathway of producing flavor substances (His, Met, ethyl hexanoate, etc.) was up-regulated in the inoculated fermentation. Results presented in this study may provide a reference for the development of paocai starters and further guidance for the flavor improvement of Sichuan paocai.

Longitudinal Analysis of the Microbial Community on the Surface of Bloodstains Under Different Environmental Conditions in Areas with Minimal Human Interference.

The association between human metabolites and the environmental microbiome has primarily been investigated in relation to disease. In this study, the associations between environmental conditions and microbial communities on the surface of bloodstains were analyzed from a forensic science approach. The composition of microbial communities can be affected by numerous variables. After exposing bloodstains to two different environments with limited airflow and human interference, the microbial communities of the bloodstain surfaces were subjected to longitudinal analysis. Various microbes showed increasing or decreasing trends at the phylum and species level. The microbes identified in this study are usually found in soil, freshwater, and seawater and are known to exhibit unique properties, such as sporulation. Longitudinal variation in temperature and humidity were associated with various changes and correlations with the blood surface microbial community. Understanding these changes could introduce a new perspective to forensic science and could be used to develop a forensic tool used at crime scenes to analyze blood stains in more detail.

Prediction of early remission after infliximab in Crohn’s disease using baseline microbiome and metabolomics

To characterize the microbiome and metabolic profile in Crohn's disease (CD) patients with different outcome after infliximab (IFX) treatment. The clinical data of a cohort of 35 patients with moderate-to-severe CD admitted at Jinling hospital between Oct 2022 and Dec 2023 were collected. Stool samples at baseline were collected to perform 16SrRNA and ITS2 sequencing and LC-MS untargeted metabolomics. Of these, seven discontinued IFX and underwent surgery during the induction period, and 28 received IFX at weeks 0, 2, and 6, each administered intravenously. Clinical remission was assessed based on the clinical symptoms and HBI at baseline and week 14. Baseline microbial richness and evenness was not significantly different between remission and non-remission group. The taxonomic community analysis identified decrease of Ruminococcus, Lachnoclostridium, Akkermansia in bacterial community and decrease of Asterotremella and Wallemia in fungal community in the non-remission group. LC–MS analysis showed that histamine, creatinine and L-proline significantly increased in remission group, while androsterone, berberine and episterol significantly decreased. The combined prediction model of histamine, androsterone, and episterol demonstrated a high predictive value of remission in patients after IFX treatment (AUC=0.898, p<0.001). Together, these data might facilitate a priori determination of optimal therapeutics for CD patients.

First evidence for temperature’s influence on the enrichment, assembly, and activity of polyhydroxyalkanoate-synthesizing mixed microbial communities

Polyhydroxyalkanoates (PHA) are popular biopolymers due to their potential use as biodegradable thermoplastics. In this study, three aerobic sequencing batch reactors were operated identically except for their temperatures, which were set at 15 °C, 35 °C, and 48 °C. The reactors were subjected to a feast–famine feeding regime, where carbon sources are supplied intermittently, to enrich PHA-accumulating microbial consortia. The biomass was sampled for 16S rRNA gene amplicon sequencing of both DNA (during the enrichment phase) and cDNA (during the enrichment and accumulation phases). All temperatures yielded highly enriched PHA-accumulating consortia. Thermophilic communities were significantly less diverse than those at low or mesophilic temperatures. In particular, Thauera was highly adaptable, abundant, and active at all temperatures. Low temperatures resulted in reduced PHA production rates and yields. Analysis of the microbial community revealed a collapse of community diversity during low-temperature PHA accumulation, suggesting that the substrate dosing strategy was unsuccessful at low temperatures. This points to future possibilities for optimizing low-temperature PHA accumulation.

Boosting conversion of waste activated sludge to methane during anaerobic digestion via facilitating direct interspecies electron transfer with glycerol

Possibility of boosting conversion of waste activated sludge to methane via facilitating direct interspecies electron transfer with glycerol was explored. Methane yields with 5 %/8 % glycerol were 1.18/1.31 folds higher than that without glycerol. After removing methane yields from glycerol, methane yields with 5 %/8 % glycerol was still 17 %/19 % higher than that without glycerol. Electron transfer coefficients for charge/discharge in sludge with 8 % glycerol were 1.12/1.39 folds higher than that without glycerol. Analysis of conductivity-temperature responses found that conductivity of sludge with 8 % glycerol exhibited an exponential increase when temperature was declined, similar to that with 8 % ethanol. However, conductivity of sludge with 8 % glycerol was one order of magnitude higher than that with 8 % ethanol. Electrochemical Fourier transform infrared spectra showed that reversible changes of bands associated with c-type cytochrome in sludge with 8 % glycerol were more intense than that with 8 % ethanol. Analysis of microbial communities showed that glycerol feeding increased the abundance of Methanospirillum hungatei JF-1, and specially enriched Tangfeifania diversioriginum, Syntrophomonas zehnderi OL-4 and Trichococcus pasteurii. These results indicated that glycerol feeding facilitated the better direct interspecies electron transfer than ethanol feeding that had been previously documented during anaerobic digestion of waste activated sludge.

Prenatal ozone exposure and variations of the gut microbiome: Evidence from a Chinese mother-infant cohort

BackgroundThe gut microbiome is central to human health, but the potential impact of ozone (O3) exposure on its establishment in early life has not been thoroughly examined. Therefore, this study aimed to investigate the relationship between prenatal O3 exposure and the variations of the human gut microbiome during the first two years of life. DesignA cohort study design was used. Pregnant women in the third trimester were recruited from an obstetric clinic, and long-term follow-ups were conducted after delivery. The gut microbiome was analyzed using the 16 S rRNA V3-V4 gene regions. Functional pathway analyses of gut microbial communities in neonates were performed using Tax4fun. The average concentrations of ambient O3 and other air pollutants from pregnancy to delivery were calculated using the China High Air Pollutants (CHAP) dataset, based on the permanent residential addresses of participants. Multiple linear regression and mixed linear models were utilized to investigate the associations between prenatal O3 exposure and gut microbiome features. ResultsPrenatal O3 exposure did not significantly affect the gut microbial alpha diversity of mothers and neonates. However, it was found to be positively associated with the gut microbial alpha diversity in 24-month-old infants. Prenatal O3 exposure explained 13.1 % of the variation in neonatal gut microbial composition. After controlling for potential covariates, prenatal O3 exposure was associated with neonatal-specific gut microbial taxa and functional pathways. Furthermore, the mixed linear models showed that prenatal O3 exposure was negatively associated with variations of Streptococcus (p-value = 0.001, q-value = 0.005), Enterococcus (p-value = 0.001, q-value = 0.005), Escherichia-Shigella (p-value = 0.010, q-value = 0.025), and Bifidobacterium (p-value = 0.003, q-value = 0.010). ConclusionsThis study is the first to examine the effects of prenatal O3 exposure on gut microbial homeostasis and variations. It demonstrates that prenatal O3 exposure is associated with variations in certain aspects of the gut microbiome. These findings provide novel insights into the dynamics and establishment of the human microbiome during the first two years of life.

Enhancing reductive dechlorination of trichloroethylene in bioelectrochemical systems with conductive materials

The incorporation of conductive materials to enhance electron transfer in bioelectrochemical systems (BES) is considered a promising approach. However, the specific effects and mechanisms of these materials on trichloroethylene (TCE) reductive dechlorination in BES remains are not fully understood. This study investigated the use of magnetite nanoparticles (MNP) and biochars (BC) as coatings on biocathodes for TCE reduction. Results demonstrated that the average dechlorination rates of MNP-Biocathode (122.89 μM Cl·d−1) and BC-Biocathode (102.88 μM Cl·d−1) were greatly higher than that of Biocathode (78.17 μM Cl·d−1). Based on MATLAB calculation, the dechlorination rate exhibited a more significantly increase in TCE-to-DCE step than the other dechlorination steps. Microbial community analyses revealed an increase in the relative abundance of electroactive and dechlorinating populations (e.g., Pseudomonas, Geobacter, and Desulfovibrio) in MNP-Biocathode and BC-Biocathode. Functional gene analysis via RT-qPCR showed the expression of dehalogenase (RDase) and direct electron transfer (DET) related genes was upregulated with the addition of MNP and BC. These findings suggest that conductive materials might accelerate reductive dechlorination by enhancing DET. The difference of physicochemical characteristics (e.g. particle size and specific surface area), electron transfer enhancement mechanism between MNP and BC as well as the reduction of Fe(III) by hydrogen may explain the superior dechlorination rate observed with MNP-Biocathode.

Epiphytic microbiome associated with intertidal seaweeds in the Mediterranean Sea: comparative analysis of bacterial communities across seaweed phyla

The complex interactions between epiphytic bacteria and marine macroalgae are still poorly understood, with limited knowledge about their community structure, interactions, and functions. This study focuses on comparing epiphytic prokaryotes community structure between three seaweed phyla; Chlorophyta, Rhodophyta, and Heterokontophyta in an easternmost rocky intertidal site of the Mediterranean Sea. By taking a snapshot approach and simultaneously collecting seaweed samples from the same habitat, we minimize environmental variations that could affect epiphytic bacterial assembly, thereby emphasizing host specificity. Through 16S rRNA gene amplicon sequencing, we identified that the microbial community composition was more similar within the same seaweed phylum host compared to seaweed host from other phyla. Furthermore, exclusive Amplicon Sequence Variants (ASVs) were identified for each algal phyla despite sharing higher taxonomic classifications across the other phyla. Analysis of niche breadth indices uncovers distinctive affinities and potential specialization among seaweed host phyla, with 39% of all ASVs identified as phylum specialists and 13% as generalists. Using taxonomy function prediction, we observed that the taxonomic variability does not significantly impact functional redundancy, suggesting resilience to disturbance. The study concludes that epiphytic bacteria composition is connected to host taxonomy, possibly influenced by shared morphological and chemical traits among genetically related hosts, implying a potential coevolutionary relationship between specific bacteria and their host seaweeds.

New insights into biologic interpretation of bioinformatic pipelines for fish eDNA metabarcoding: A case study in Pearl River estuary

Environmental DNA (eDNA) metabarcoding is an emerging tool for monitoring biological communities in aquatic ecosystems. The selection of bioinformatic pipelines significantly impacts the results of biodiversity assessments. However, there is currently no consensus on the appropriate bioinformatic pipelines for fish community analysis in eDNA metabarcoding. In this study, we compared three bioinformatic pipelines (Uparse, DADA2, and UNOISE3) using real and mock (constructed with 15/30 known fish) communities to investigate the differences in biological interpretation during the data analysis process in eDNA metabarcoding. Performance evaluation and diversity analyses revealed that the choice of bioinformatic pipeline could impact the biological results of metabarcoding experiments. Among the three pipelines, the operational taxonomic units (OTU)-based pipeline (Uparse) showed the best performance (sensitivity: 0.6250 ± 0.0166; compositional similarity: 0.4000 ± 0.0571), the highest richness (25–102) and minimal inter-group differences in alpha diversity. It suggested the OTU-based pipeline possessed superior capability in fish diversity monitoring compared to ASV/ZOTU-based pipeline. Additionally, the Bray-Curtis distance matrix achieved the highest discriminative effect in the PCoA (43.3%–53.89%) and inter-group analysis (P < 0.01), indicating it was better at distinguishing compositional differences or specific genera of fish community at different sampling sites than other distance matrices. These findings provide new insights into fish community monitoring through eDNA metabarcoding in estuarine environments.

The sediment of a river having ‘’good ecological status” turned out to be toxic to midge Chironomus riparius larvae: Implication for environmental monitoring?

Globally, monitoring of the surface waters is largely limited to the physico-chemical analysis of water in rivers and lakes. Sediment state in the aquatic systems including sediment chemical content or the structure and diversity of benthic communities or ecotoxicological studies with natural sediments remains largely overlooked by the monitoring programs. Hence we assessed the potential toxicity of three riverine sediments on the life-cycle traits (emergence and reproduction) of midge Chironomus riparius via an ecotoxicological testing method over two generations (according to OECD test 233 guidelines). In addition, the riverine sediments were spiked with polyamide (nylon) microplastic particles (1 g kg−1) to analyze an additive effect of microplastic on the sediment toxicity. As model river systems, three rivers (Karchaghbyur, Gavaraget, Argichi) in the Lake Sevan basin (Armenia) were selected. Results of ecotoxicity testing were compared with the indices of water quality (derived from the physico-chemical analysis) and the indices of the ecological status of the rivers (derived from the analysis of benthic communities). The results of testing demonstrated an unexpectedly low emergence of midges after the first generation exposed to the sediment of the river having ‘’good ecological status’’ - the Argichi. Sediments of the Karchaghbyur and Gavaraget rivers impeded the emergence and reproduction of midges after the second generation. An addition of polyamide particles to the sediments did not significantly affect the life-cycle traits of C. riparius indicating the primary effect of the sediments’ condition. The discrepancy of biotesting result with that of the other two methods (which indicated ‘’average water quality’’ and “good ecological status”) underlies the importance of designing more comprehensive monitoring programs for better assessment and protection of aquatic systems and resources.

Efficient nitrogen removal of decentralized rural sewage: Pilot-scale study of a novel push-flow coupled bio-ecological system

The performance and mechanisms for pollutant removal were investigated in an extended pilot-scale operation. Results demonstrated that the PF-EFB system maintained stable and efficient performance after over 450 days of continuous operation. Under various operating conditions, the total nitrogen (TN) concentration was reduced to <2 mg/L, and the removal rate exceeded 90 %. The effluent chemical oxygen demand (COD) concentration remained below 40 mg/L. Additionally, a combination of nitrogen transformation and molecular biology analyses was conducted to explore the mechanisms. The key bacterial groups driving system performance were aerobic and anoxic nitrifying bacteria, which dominated the microbial population. Microbial community analysis showed that the abundance of aerobic denitrifying bacteria in the middle of the system was 40.5 %, while the abundance of anoxic denitrifying bacteria at the end was 19.0 %. This spatial variation in the distribution of aerobic and anoxic denitrifying bacteria along the flow direction inside the reactor, driven by environmental factors, is key to the system's efficient denitrification. Overall, the PF-EFB system effectively addressed challenges posed by water flow fluctuations and enhanced nitrogen removal efficiency. This innovative system offers significant benefits and serves as a valuable reference for rural sewage treatment.

Strengthening bioremediation potential: Enterobacter ludwigii ES2 for combined nicosulfuron and Cd contamination through whole genome and microbial diversity community analysis

Nicosulfuron and Cd are common pollutants that pose significant threats to the environment and human health, particularly under combined stress. This study is the first to remediate environmental nicosulfuron and Cd under combined stress using microbiological techniques. Enterobacter ludwigii ES2 was isolated, characterized, and demonstrated to degrade 93.80 % of nicosulfuron and remove 59.64 % of Cd within 4 d. Potential functional genes, including nicosulfuron degradation genes gstA, gstB, glnQ, glnP, mreB, and sixA, and Cd tolerance/removal-related genes mntA, mntB, mntH, dnaK, znuA, and zupt, were predicted by sequencing the whole genome of strain ES2, and their expression was verified by qRT-PCR. Strain ES2 managed oxidative stress induced by Cd through superoxide dismutase, glutathione, catalase, peroxidase, and malondialdehyde. Furthermore, to repair compound stress, up to 90.48 % of nicosulfuron and 67.74 % of Cd were removed. The community structure analysis indicated that Enterobacteriaceae, Sphingomonadaceae, and Gemmatimonadaceae were dominant populations, with ES2 stably colonizing and becoming the dominant bacterium. In summary, ES2 demonstrated significant potential in remediating nicosulfuron and Cd pollution from various perspectives, providing a solid theoretical foundation.

Comparative Study of Community Analysis of Plant Parasitic and Free-living Nemic Fauna around Root Zones of Mango (Mangifera indica) Plant in Hapur Region

Nematodes are tiny rounded body, unsegmented worms. They occupy any niche of the environment, like soil, freshwater, and terrestrial (free-living and parasitic). They constitute a large group of all metazoans in the world: Plant-parasitic and free-living nematodes are well-recognized factors in soil productivity. Phytonematodes feed upon the cell sap of plants with the help of stylet. It causes many abnormalities in plants like gall formation on roots, stunting growth, wilting of plants, and serious damage to crop yield. In the present study, soil samples were collected from (0-30 cm) vertical depth and (30-60 cm) horizontal depth from around the root of the mango plant in the Hapur region from July to November. Nematodes samples were extracted by decanting and sieving Techniques (Cobb1918) were applied. Under the microscope examination nematodes were identified as free-living with minimum percentages of 14.2% and8.88% at both vertical and horizontal depth while plant parasitic nematodes were recorded as a higher percentage(53.90% vertical depth)and(64.91% horizontal depth) away from the host plant. This was variation in communities of plant nematodes at both depths due to seasonal changes, various other edaphic factors such as PH, temperature, moisture emergence of secondary rootlets, and availability on set of monsoon season and unavailability of various food resources.

16S amplicon-based microbiome biomapping of a commercial broiler hatchery

Hatcheries, where eggs from multiple breeder farms are incubated and hatched before being sent to different broiler farms, represent a nexus point in the commercial production of broilers in the United States. Considering all downstream microbial quality and safety aspects of broiler production (live production, processing, consumer use) can be potentially affected by the hatchery, a better understanding of microbial ecology within commercial hatcheries is essential. Therefore, a commercial broiler hatchery was biomapped using 16S rRNA amplicon-based microbiome analyses of four sample type categories (Air, Egg, Water, Facility) across five different places in the pre-hatch, hatch, and post-hatch areas. While distinct microbiota were found for each sample type category and hatchery area, microbial community analyses revealed that Egg microbiota trended towards clustering with the facility-related samples when moving from the prehatch to post-hatch areas, highlighting the potential effect of the hatchery environment in shaping the pre-harvest broiler-related microbiota. Prevalence analyses revealed 20 ASVs (Core20) present in the core microbiota of all sample types and areas, with each ASV possessing a unique distribution throughout the hatchery. Interestingly, three Enterobacteriaceae ASVs were in the Core20, including Salmonella. Subsequent analyses showed that Salmonella, while a minor prehatch and hatch Core20ASV, dominated the Enterobacteriaceae niche and total microbiota in the chick pad feces in the post-hatch area of the hatchery, and the presence of this Salmonella ASV in the post-hatch feces was associated with swabs of breakroom tables. These findings highlight the complexity of commercial hatchery microbiota, including identifying chick pad feces and breakroom tables as potentially important sampling or disinfection targets for hatchery managers to focus their Salmonella mitigation efforts to reduce loads entering live production farms.

Contrasted trends of intertidal macroalgal communities and sharp decline of canopy-forming species across two decades

Macroalgal communities are essential to coastal ecosystems, yet increasing effects of global change and anthropogenic pressures are leading to their global decline. Investigating the long-term dynamics of these communities across different localities appears crucial to better understand their responses to such pressures, as our knowledge of spatial heterogeneities in macroalgal trajectories remains elusive. To fill this gap, the community trajectory analysis framework provides a set of innovative multivariate metrics to characterize and quantitatively compare the temporal dynamics of different communities. Using long-term monitoring data (2004-2022), this method was applied to intertidal macroalgal communities across 10 locations distributed over more than 500 km of coastline in Brittany, France. Three distinct temporal dynamics were identified. High-shore communities exhibited minimal changes over time, while low-shore communities were characterized by a fluctuating understorey species composition but a general stability pattern. In contrast, the mid-shore community dominated by Ascophyllum nodosum underwent conspicuous changes in composition and structure. Further analysis of the latter community unveiled clear spatial patterns, with a significant deterioration of the structural state attributed to canopy loss in eastern Brittany, negatively impacting understorey species. This decline may ultimately lead to massive changes in coastal ecosystem functioning and services. This study emphasizes the importance of maintaining long-term ecological monitoring as well as the pertinence of temporal trajectories methods to identify and understand community changes at various spatial scales.

Isolation and characterization of novel acetogenic Moorella strains for employment as potential thermophilic biocatalysts.

Thermophilic acetogenic bacteria have attracted attention as promising candidates for biotechnological applications such as syngas fermentation, microbial electrosynthesis, and methanol conversion. Here, we aimed to isolate and characterize novel thermophilic acetogens from diverse environments. Enrichment of heterotrophic and autotrophic acetogens was monitored by 16S rRNA gene-based bacterial community analysis. Seven novel Moorella strains were isolated and characterized by genomic and physiological analyses. Two Moorella humiferrea isolates showed considerable differences during autotrophic growth. The M. humiferrea LNE isolate (DSM 117358) fermented carbon monoxide (CO) to acetate, while the M. humiferrea OCP isolate (DSM 117359) transformed CO to hydrogen and carbon dioxide (H2 + CO2), employing the water-gas shift reaction. Another carboxydotrophic hydrogenogenic Moorella strain was isolated from the covering soil of an active charcoal burning pile and proposed as the type strain (ACPsT) of the novel species Moorella carbonis (DSM 116161T and CCOS 2103T). The remaining four novel strains were affiliated with Moorella thermoacetica and showed, together with the type strain DSM 2955T, the production of small amounts of ethanol from H2 + CO2 in addition to acetate. The physiological analyses of the novel Moorella strains revealed isolate-specific differences that considerably increase the knowledge base on thermophilic acetogens for future applications.

Complete pipeline for Oxford Nanopore Technology amplicon sequencing (ONT-AmpSeq): from pre-processing to creating an operational taxonomic unit table.

Amplicon sequencing has long served as a robust method for characterising microbial communities, and despite inherent resolution limitations, it remains a preferred technique, offering cost- and time-effective insights into bacterial compositions. Here, we introduce ONT-AmpSeq, a user-friendly pipeline designed for processing amplicon sequencing data generated from Oxford Nanopore Technology (ONT) devices. Our pipeline enables efficient creation of taxonomically annotated operational taxonomic unit (OTU) tables from ONT sequencing data, with the flexibility to multiplex amplicons on the same barcode. The pipeline encompasses six main steps-statistics, quality filtering, alignment, clustering, polishing, and taxonomic classification-integrating various state-of-the-art software tools. We provide a detailed description of each step, along with performance tests and robustness evaluations using both test data and a ZymoBIOMICS® Microbial Community Standard mock community dataset. Our results demonstrate the ability of ONT-AmpSeq to effectively process ONT amplicon data, offering valuable insights into microbial community composition. Additionally, we discuss the influence of polishing tools on taxonomic insight and the impact of taxonomic annotation methods on the derived microbial composition. Overall, ONT-AmpSeq represents a comprehensive solution for analysing ONT amplicon sequencing data, facilitating streamlined and reliable microbial community analysis. The pipeline, along with test data, is freely available for public use.

Local signal from Trichoderma afroharzianum T22 induces host transcriptome and endophytic microbiome leading to growth promotion in sorghum.

Trichoderma, a highly abundant soil fungus, may benefit plants, yet it remains understudied in sorghum (Sorghum bicolor). In this study, sorghum plants were cultivated for five weeks in soil pots with or without the inoculation of T. afroharzianum T22. We found that plants inoculated with T. afroharzianum T22 exhibited significant increases in growth parameters and nutrient levels, demonstrating its beneficial role in sorghum. The split-root assay demonstrated that T. afroharzianum T22 is essential in both compartments of soil pots for promoting plant growth. This suggests that local signals from this fungus drive symbiotic benefits in sorghum. The RNA-seq analysis revealed the induction of genes responsible for mineral transport (such as nitrate and aquaporin transporters), auxin response, sugar assimilation (hexokinase), and disease resistance (thaumatin) in the roots of sorghum inoculated with T. afroharzianum T22. Microbial community analysis further unveiled the positive role of T. afroharzianum T22 in enriching Penicillium (ITS) and Streptomyces (16S) while reducing disease-causing Fusarium in the roots. The microbial consortium, consisting of enriched microbiomes from bacterial and fungal communities, showed disrupted morphological features in plants inoculated with T. afroharzianum T22 in the absence of S. griseus. However, this disruption was not observed in the absence of P. chrysogenum. These results suggest that S. griseus may act as a helper microbe in close association with T. afroharzianum T22 in the sorghum endosphere. This study provides the first comprehensive explanation of how T. afroharzianum T22 modulates host molecular determinants and endophytic helper microbes, thereby collectively promoting sorghum growth. These findings may encourage the formulation of a synthetic inoculum dominated by T. afroharzianum T22 to enhance growth and stress resilience in sorghum and other similar crops.

The impact of extraction method and pollen concentration on community composition for pollen metabarcoding

AbstractPremisePlants and pollinators closely interact with each other to form complex networks of species interactions. Metabarcoding of pollen collections has recently been proposed as an advantageous method for the construction of such networks, but the extent to which diversity and community analyses depend on the extraction method and pollen concentration used remains unclear.MethodsIn this study, we used a dilution series of two pollen mixtures (a mock community and pooled natural pollen loads from bumblebees) to assess the effect of mechanical homogenization and two DNA extraction kits (spin column DNA extraction kit and magnetic bead DNA extraction kit) on the detected pollen richness and community composition.ResultsAll species were successfully detected using the three methods, even in the most dilute samples. However, the extraction method had a significant effect on the detected pollen richness and community composition, with simple mechanical homogenization introducing an extraction bias.DiscussionOur findings suggest that all three methods are effective for detecting plant species in the pollen loads on insects, even in cases of very low pollen loads. However, our results also indicate that extraction methods can have a profound impact on the ability to correctly assess the community composition of the pollen loads on insects. The choice of extraction methodology should therefore be carefully considered to ensure reliable and unbiased results in pollen diversity and community analyses.