Sludge digestibility and functionally active microorganisms in methanogenic sludge digesters revealed by E. coli-fed digestion and microbial source tracking.

Methanogenic sludge digestion plays a pivotal role in attenuating and hygienizing the massively-produced waste activated sludge (WAS), which is predominantly composed of microbial cells and extracellular polymeric substances (EPS). The efficient sludge digestion requires a variety of functionally active microorganisms working together closely to convert sludge organic matter into biogas. Nonetheless, the digestion efficiency (or digestibility quantified as carbon removal efficiency) of major sludge constituents (i.e., microbial cells and EPS) and associated functionally active microorganisms in sludge digesters remain elusive. In this study, we identified the digestibility of sludge microbial cells and the associated functionally active microorganisms by using Escherichia coli (E. coli)-fed digestion and microbial source tracking. The average carbon removals in four digesters fed with fresh WAS (WAS-AD), thermal pretreated WAS (Thermal-WAS-AD), E. coli cells (E.coli-AD) and thermal pretreated E. coli cells (Thermal-E.coli-AD) were 30.6 ± 3.4%, 45.8 ± 2.9%, 69.0 ± 3.4% and 68.9 ± 4.6%, respectively. Compared to WAS-AD and Thermal-WAS-AD, the significantly higher carbon removals in E. coli-AD and Thermal-E. coli-AD suggested the remarkably higher digestibility of microbial cells than EPS, and releasing organic matter from EPS might be a rate-limiting step in sludge digestion. Functionally active microorganisms for microbial cell digestion predominantly included fermenters (e.g., Petrimonas and Lentimicrobium), syntrophic acetogens (e.g., Synergistaceae) and methanogens (e.g., Methanosaeta and Methanosarcina). Microbial source tracking estimation showed that the microbial cell-digesting populations accounted for 35.6 ± 9.1% and 70.3 ± 10.1% of total microbial communities in the WAS-AD and Thermal-WAS-AD, respectively. Accordingly, the functionally active microorganisms for digestion of both microbial cells and EPS accounted for 64.5 ± 12.1% and 97.3 ± 2.0% of total digestion sludge microbiome in WAS-AD and Thermal-WAS-AD, respectively. By contrast, feeding WAS-derived microorganisms accounted for 23.2 ± 4.4% and 2.3 ± 1.2% of total microbial communities in the WAS-AD and Thermal-WAS-AD, respectively.

Effects of transgenic Bt rice on the active rhizospheric methanogenic archaeal community as revealed by DNA-based stable isotope probing.

AIMS: This study aimed to investigate the influence of planting Cry1Ab/Cry1Ac gene expressing rice (Bt rice) on rhizospheric active methanogenic archaeal communities. METHODS AND RESULTS: The nontransgenic parental line was used as the control (Ck rice). DNA-based stable isotope probing (DNA-SIP) technology traced the rhizospheric active methanogens at the tillering stage. The results revealed significantly lower CH4 emission flux from Bt soil than that from Ck soil during the whole growth period. The active methanogenic community composition remained stable. The RC-I lineage (77·9-79·8%) and Methanosaetaceae (13·9-15·1%) were the predominant active methanogens in Bt and Ck rice rhizospheres. However, the abundance of functionally active methanogens in the Bt rice rhizosphere was significantly reduced. Lower levels of root exudates (that included carbohydrate and organic acids) from Bt rice were also detected at the tillering stage. CONCLUSION: This study found that the genetic modification of rice reduced the potential methanogenic substrates came from plant-derived root exudates, which represented an important factor in reducing CH4 generation and active methanogenic archaeal abundance in Bt rhizosphere soil. SIGNIFICANCE AND IMPACT OF THE STUDY: The effect of genetically modified (GM) insect-resistant crops on soil micro-organisms has become an issue of public concern, especially the indirect effect of plant metabolisms caused by the insertion of foreign genes. Methanogenesis, which is regarded as a critical ecological process in paddy soil, is influenced by plant root exudates; these are mainly derived from photosynthesis. The variations in root exudates across the Bt and Ck rice suggested the indirect influence of foreign gene insertion. DNA-SIP successfully traced the active methanogenic archaeal populations assimilating 13 C-labelled photosynthetic carbon and found a strong influence of planting Bt rice on active methanogens. As a consequence, we proposed that analysis of functionally active micro-organisms is more suitable for monitoring and predicting the environmental influence of GM plants.

The use of Yucca schidigera and microbial preparation for poultry manure deodorization and hygienization.

The aim of this study was to determine the effectiveness of microbial preparation and Yucca schidigera in the removal of odorous volatile compounds from poultry manure as well as to evaluate antimicrobial properties of these amendments. It was demonstrated that the combined treatment of poultry manure (PM) with the microbial preparation and Y. schidigera extract can reduce the concentration of odorants by 58%-73%, depending on the tested compound. When Y. schidigera extract and the microbial preparation were applied at a time interval of 48 h, the deodorization efficiency was improved by 6-24%. Furthermore, Y. schidigera extract has antimicrobial properties, which affect poultry manure hygienization. It was found that when the microbial preparation was enriched with Lactobacillus plantarum, it became insensitive to the antimicrobial properties of Y. schidigera.

Removal of odorous compounds from poultry manure by microorganisms on perlite--bentonite carrier.

Laboratory-scale experiments were conducted using poultry manure (PM) from a laying hen farm. Six strains of bacteria and one strain of yeast, selected on the base of the previous study, were investigated to evaluate their activity in the removal of odorous compounds from poultry manure: pure cultures of Bacillus subtilis subsp. spizizenii LOCK 0272, Bacillus megaterium LOCK 0963, Pseudomonas sp. LOCK 0961, Psychrobacter faecalis LOCK 0965, Leuconostoc mesenteroides LOCK 0964, Streptomyces violaceoruber LOCK 0967, and Candida inconspicua LOCK 0272 were suspended in water solution and applied for PM deodorization. The most active strains in the removal of volatile odorous compounds (ammonia, hydrogen sulfide, dimethylamine, trimethylamine, isobutyric acid) belonged to B. subtilis subsp. spizizenii, L. mesenteroides, C. inconspicua, and P. faecalis. In the next series of experiments, a mixed culture of all tested strains was immobilized on a mineral carrier being a mixture of perlite and bentonite (20:80 by weight). That mixed culture applied for PM deodorization was particularly active against ammonia and hydrogen sulfide, which were removed from the exhaust gas by 20.8% and 17.5%, respectively. The experiments also showed that during deodorization the microorganisms could reduce the concentrations of proteins and amino acids in PM. In particular, the mixed culture was active against cysteine and methionine, which were removed from PM by around 45% within 24 h of deodorization.

Bacterial dispersal enhances the elimination of active fecal coliforms during vermicomposting of fruit and vegetable wastes: The overlooked role of earthworm mucus.

Earthworms play a pivotal role in the elimination of fecal coliforms during vermicomposting of fruit and vegetable waste (FVWs). However, the specific mechanisms underlying the action of earthworm mucus remain unclear. This study investigated the mechanisms of fecal coliform reduction related to earthworm mucus during FVWs vermicomposting by comparing treatments with and without earthworms. The results show that the secretion of earthworm mucus decreased by 13.93 % during the startup phase, but significantly (P < 0.001) increased by 57.80 % during the degradation phase. Compared to the control without earthworms, vermicomposting led to a significant (P < 0.05) 1.22 -fold increase in the population of active bacteria, with a strong positive correlation between mucus characteristics and dominant bacterial phyla. As the dominant fecal coliforms, Escherichia coli and Klebsiella pneumoniae significantly (P < 0.05) declined by 86.20 % and 93.38 %, respectively, in the vermi-reactor relative to the control. Bacterial dispersal limitation served as a key factor constraining the elimination of E. coli (r = 0.73, P < 0.01) and K. pneumoniae (r = 0.77, P < 0.001) during vermicomposting. This study suggests that earthworm mucus increases the active bacterial abundance and cooperation by weakening the bacterial dispersal limitation, thus intensifying competition and antagonism between fecal coliforms and other bacteria.

Selective Elucidation of Living Microbial Communities in Fermented Grains of Chinese Baijiu: Development of a Technique Integrating Propidium Monoazide Probe Pretreatment and Amplicon Sequencing.

The fermentation process of Chinese Baijiu's fermented grains involves the intricate succession and metabolism of microbial communities, collectively shaping the Baijiu's quality. Understanding the composition and succession of these living microbial communities within fermented grains is crucial for comprehending fermentation and flavor formation mechanisms. However, conducting high-throughput analysis of living microbial communities within the complex microbial system of fermented grains poses significant challenges. Thus, this study addressed this challenge by devising a high-throughput analysis framework using light-flavor Baijiu as a model. This framework combined propidium monoazide (PMA) pretreatment technology with amplicon sequencing techniques. Optimal PMA treatment parameters, including a concentration of 50 µM and incubation in darkness for 5 min followed by an exposure incubation period of 5 min, were identified. Utilizing this protocol, viable microorganism biomass ranging from 8.71 × 106 to 1.47 × 108 copies/µL was successfully detected in fermented grain samples. Subsequent amplicon sequencing analysis revealed distinct microbial community structures between untreated and PMA-treated groups, with notable differences in relative abundance compositions, particularly in dominant species such as Lactobacillus, Bacillus, Pediococcus, Saccharomycopsis, Issatchenkia and Pichia, as identified by LEfSe analysis. The results of this study confirmed the efficacy of PMA-amplicon sequencing technology for analyzing living microbial communities in fermented grains and furnished a methodological framework for investigating living microbial communities in diverse traditional fermented foods. This technical framework holds considerable significance for advancing our understanding of the fermentation mechanisms intrinsic to traditional fermented foods.

Bioturbation analysis of microbial communities and flavor metabolism in a high-yielding cellulase Bacillus subtilis biofortified Daqu.

In this study, a fortified Daqu (FF Daqu) was prepared using high cellulase-producing Bacillus subtilis, and the effects of in situ fortification on the physicochemical properties, flavor, active microbial community and metabolism of Daqu were analyzed. The saccharification power, liquefaction power, and cellulase activity of the FF Daqu were significantly increased compared with that of the traditional Daqu (CT Daqu). The overall differences in flavor components and their contents were not significant, but the higher alcohols were lower in FF Daqu. The relative abundance of dominant active species in FF Daqu was 85.08% of the total active microbiota higher than 63.42% in CT Daqu, and the biomarkers were Paecilomyces variotii and Aspergillus cristatus, respectively. The enzymes related to starch and sucrose metabolic pathways were up-regulated and expressed in FF Daqu. In the laboratory level simulation of baijiu brewing, the yield of baijiu was increased by 3.36% using FF Daqu.

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

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

Bioprospecting for electrochemically active perchlorate-reducing microorganisms.

This study evaluated the electrochemical capacity of four perchlorate-reducing microorganisms (PRMs) isolated from an Altiplanic Andean watershed naturally pressured with perchlorate. Three-electrode electrochemical cells were used to test the electrochemical activity of the obtained isolates. Electrochemical evaluation (i.e., cyclic voltammetry, electrochemical impedance spectroscopy, chronoamperometry) revealed that two isolates identified as Dechloromonas sp. CS-1 and Clostridioides sp. CS-2 are electrochemically active PRMs. Bacterial isolates exhibiting cathodic peaks at -651 mV and -303 mV (vs. Ag/AgCl) for CS-1 and CS-2, respectively. Electrotrophic perchlorate removal was demonstrated by a 6-days chronoamperometry with removal rates of 27 and 17 mg L-1 day-1 and cathodic efficiencies of 93% and 45%, for CS-1 and CS-2, respectively. Chemical and electrochemical results suggest two different mechanisms of electrotrophic perchlorate removal, a complete eight-electron bio-reduction (i.e., perchlorate to chloride) for CS-1 and a partial two-electron bio-reduction (i.e., perchlorate to chlorate) for CS-2. The observed differences could be linked to their enzymatic differences, as in their membrane compositions. Thus, the results of this work increase the limited number of known electrotrophic microorganisms and expand the application of bioelectrochemical systems to develop new perchlorate treatment and remediation technologies.

Winogradsky Bioelectrochemical System as a Novel Strategy to Enrich Electrochemically Active Microorganisms from Arsenic-Rich Sediments.

Bioelectrochemical systems (BESs) have been extensively studied for treatment and remediation. However, BESs have the potential to be used for the enrichment of microorganisms that could replace their natural electron donor or acceptor for an electrode. In this study, Winogradsky BES columns with As-rich sediments extracted from an Andean watershed were used as a strategy to enrich lithotrophic electrochemically active microorganisms (EAMs) on electrodes (i.e., cathodes). After 15 months, Winogradsky BESs registered power densities up to 650 µWcm-2. Scanning electron microscopy and linear sweep voltammetry confirmed microbial growth and electrochemical activity on cathodes. Pyrosequencing evidenced differences in bacterial composition between sediments from the field and cathodic biofilms. Six EAMs from genera Herbaspirillum, Ancylobacter, Rhodococcus, Methylobacterium, Sphingomonas, and Pseudomonas were isolated from cathodes using a lithoautotrophic As oxidizers culture medium. These results suggest that the tested Winogradsky BES columns result in an enrichment of electrochemically active As-oxidizing microorganisms. A bioelectrochemical boost of centenarian enrichment approaches, such as the Winogradsky column, represents a promising strategy for prospecting new EAMs linked with the biogeochemical cycles of different metals and metalloids.

Nanomaterials Facilitating Microbial Extracellular Electron Transfer at Interfaces.

Electrochemically active bacteria can transport their metabolically generated electrons to anodes, or accept electrons from cathodes to synthesize high-value chemicals and fuels, via a process known as extracellular electron transfer (EET). Harnessing of this microbial EET process has led to the development of microbial bio-electrochemical systems (BESs), which can achieve the interconversion of electrical and chemical energy and enable electricity generation, hydrogen production, electrosynthesis, wastewater treatment, desalination, water and soil remediation, and sensing. Here, the focus is on the current understanding of the microbial EET process occurring at both the bacteria-electrode interface and the biotic interface, as well as some attempts to improve the EET by using various nanomaterials. The behavior of nanomaterials in different EET routes and their influence on the performance of BESs are described. The inherent mechanisms will guide rational design of EET-related materials and lead to a better understanding of EET mechanisms.

[Growth Features of Water Supply Pipeline Biofilms Based on Active Microorganisms].

Biofilm in the water distribution system is composed of various active microorganisms, microbial residues, and organic and inorganic impurities. The growth of pipeline biofilms causes several drinking water hygiene problems, such as the colonization of opportunistic pathogens, corrosion of pipelines, and deterioration of water quality. To investigate the biofilm formation process and the distribution characteristics of live microorganisms attached to drinking water pipe walls, heterotrophic plate count (HPC), flow cytometry (FCM), and high-throughput 16S rRNA sequencing were analyzed to examine the growth characteristics and diversity of active microorganisms on the wall of three typical building water supply pipeline materials:polyvinyl chloride (PVC), polypropylene random (PPR), and stainless steel (STS). The results showed that after 73 d, the biomass on the PVC pipe wall was firstly up to peak. The relationships between the maximum biomass and mature biofilm biomass per unit area among the three pipelines were both PVC > PPR > STS. Regarding the composition and structure of the biofilm active bacterial community on the three different pipelines, the predominant phylum on PVC was Nitrospirae, while Verrucomicrobia was predominant on PPR and STS. The species richness and diversity of biofilm on STS were smallest, and thus the community structure was the simplest. Compared to STS, there were more Cyanobacteria and Actinomycetes on PVC and PPR-hence, odor is more likely to be an issue. The results demonstrated that pipe material can affect the community structure of the attached biofilm.

Microbial fuel cells as an alternative energy source: current status.

Microbial fuel cell (MFC) technology is an emerging area for alternative renewable energy generation and it offers additional opportunities for environmental bioremediation. Recent scientific studies have focused on MFC reactor design as well as reactor operations to increase energy output. The advancement in alternative MFC models and their performance in recent years reflect the interests of scientific community to exploit this technology for wider practical applications and environmental benefit. This is reflected in the diversity of the substrates available for use in MFCs at an economically viable level. This review provides an overview of the commonly used MFC designs and materials along with the basic operating parameters that have been developed in recent years. Still, many limitations and challenges exist for MFC development that needs to be further addressed to make them economically feasible for general use. These include continued improvements in fuel cell design and efficiency as well scale-up with economically practical applications tailored to local needs.

Pyrosequencing Reveals a Core Community of Anodic Bacterial Biofilms in Bioelectrochemical Systems from China.

Bioelectrochemical systems (BESs) are promising technologies for energy and product recovery coupled with wastewater treatment, and the core microbial community in electrochemically active biofilm in BESs remains controversy. In the present study, 7 anodic communities from 6 bioelectrochemical systems in 4 labs in southeast, north and south-central of China are explored by 454 pyrosequencing. A total of 251,225 effective sequences are obtained for 7 electrochemically active biofilm samples at 3% cutoff level. While Alpha-, Beta-, and Gamma-proteobacteria are the most abundant classes (averaging 16.0-17.7%), Bacteroidia and Clostridia are the two sub-dominant and commonly shared classes. Six commonly shared genera i.e., Azospira, Azospirillum, Acinetobacter, Bacteroides, Geobacter, Pseudomonas, and Rhodopseudomonas dominate the electrochemically active communities and are defined as core genera. A total of 25 OTUs with average relative abundance >0.5% were selected and designated as core OTUs, and some species relating to these OTUs have been reported electrochemically active. Furthermore, cyclic voltammetry and chronoamperometry tests show that two strains from Acinetobacter guillouiae and Stappia indica, bacteria relate to two core OTUs, are electrochemically active. Using randomly selected bioelectrochemical systems, the study has presented extremely diverse bacterial communities in anodic biofilms, though, we still can suggest some potentially microbes for investigating the electrochemical mechanisms in bioelectrochemical systems.