High-performance internal circulation anaerobic granular sludge reactor for cattle slaughterhouse wastewater treatment and simultaneous biogas production.

This research investigates the efficacy of a high-performance pilot-scale Internal Circulation Anaerobic Reactor inoculated with Granular Sludge (ICAGSR) for treating cattle slaughterhouse wastewater while concurrently generating biogas. The primary objective is to assess the efficiency and performance of ICAGSR in terms of organic pollutant removal and biogas production using granular anaerobic sludge. The research methodology entails operating the ICAGSR system under ambient conditions and systematically varying key parameters, including different Hydraulic Retention Times (HRTs) (24, 12, and 8 h) and Organic Loading Rates (OLRs) (3.3, 6.14, and 12.83 kg COD/m³. d). The study focuses on evaluating pollutants' removal and biogas production rates. Results reveal that the ICAGSR system achieves exceptional removal efficiency for organic pollutants, with Chemical Oxygen Demand (COD) removal exceeding 74%, 67%, and 68% at HRTs of 24, 12, and 8 h, respectively. Furthermore, the system demonstrates stable and sustainable biogas production, maintaining average methane contents of 80%, 76%, and 72% throughout the experimental period. The successful operation of the ICAGSR system underscores its potential as a viable technology for treating cattle slaughterhouse wastewater and generating renewable biogas. In conclusion, this study contributes to wastewater treatment and renewable energy production by providing a comprehensive analysis of the ICAGSR system's hydrodynamic properties. The research enhances our understanding of the system's performance optimization under varying conditions, emphasizing the benefits of utilizing ICAGSR reactors with granular sludge as an effective and sustainable approach. Identifying current gaps, future research directions aim to further refine and broaden the application of ICAGSR technology in wastewater treatment and renewable energy initiatives.

Detection of thermotolerant coliforms and SARS-CoV-2 RNA in sewage and recreational waters in the Ecuadorian coast: A call for improving water quality regulation.

Wastewater surveillance represents an alternative approach to regulating contamination and the early detection of infectious agents and outbreaks of diseases of public health importance. This study evaluated domestic wastewater effects on recreational waters in estuarine and seawater bodies in Guayas and Santa Elena provinces in Ecuador, South America. Fecal indicator bacteria (thermotolerant coliforms) served as key indicators for evaluation. Physical, chemical, and microbiological quality markers following the Ecuadorian environmental quality standard and the discharge of effluents to the water resource were analyzed. Samples were collected from 44 coastal sites and 2 oxidation lagoons during the dry and rainy seasons of 2020 and 2021, respectively. SARS-CoV-2 RNA was detected in samples with higher E. coli concentrations using reverse transcription quantitative PCR to detect the genes N and ORF1ab. All samples analyzed for SARS-CoV-2 showed Ct ˂ 40 for at least one gene. Four samples showed at least 20 genome copies of gene N per reaction. These were at an artisanal fishing port, an estuarine area (Palmar), a recreational bay, and an oxidation lagoon. A moderate correlation was found between SARS-CoV-2 RNA, thermotolerant coliform and E. coli (p-value ≤ 0.0037), and a strong and positive correlation between thermotolerant coliform and E. coli. (p-value ≤ 0.00001), highlighting the utility of these established parameters as a proxy of the virus. Significant differences were found in the concentrations of thermotolerant coliforms between seasons (p-value = 0.016) and sites (p-value = 0.005). The highest levels of coliforms were found in the dry season (63000 MPN/100 mL) in Anconcito and during the rainy season (14000 MPN/100 mL) at Esterillo in Playas County. It is recommended that the decentralized autonomous governments of the surveyed provinces in Ecuador implement urgent corrective actions and establish medium-term mechanisms to minimize a potential contamination route. Additional parameters must be included in the monitoring, such as Enterococcus and intestinal parasites, due to their public health implications. In the oxidation lagoons, maintenance actions must be carried out, including the dissolution of sediments, an increase in water retention times, and in situ treatment of the sludge, to improve the system's performance.

Physicochemical parameters affecting the adhesion of ciprofloxacin-resistant Escherichia coli to activated sludge.

To investigate the physicochemical conditions necessary to stably remove antibiotic-resistant bacteria (ARB) via contact with activated sludge (AS), the adhesion of ciprofloxacin (CIP)-resistant and -susceptible Escherichia coli to AS was simulated by contact tests in the laboratory. The CIP-resistant E. coli and susceptible E. coli were removed by a 3 log smaller concentration by a 5 h contact test at maximum. Considering the hydraulic retention time of a reaction tank (∼5 h) and step-feeding operation, we considered the removal rate of E. coli in the current simulated contact test to be in agreement with the actual situation where 1-2 log concentrations of E. coli were reported to be removed from an AS reaction tank. With the increase in the AS concentration and/or dissolved oxygen, the removal rate of E. coli increased. The removal rate of CIP-resistant E. coli was greater than that of susceptible E. coli under all experimental conditions. Although the mechanism by which CIP-resistant E. coli preferably adhered to AS was not clearly understood in detail, finding optimum conditions under which bacteria, including ARB, were efficiently removed by the AS process may be possible.

Characterization and efficacy against carbapenem-resistant Acinetobacter baumannii of a novel Friunavirus phage from sewage.

Carbapenem-resistant Acinetobacter baumannii (CRAB) has become a new threat in recent years, owing to its rapidly increasing resistance to antibiotics and new effective therapies are needed to combat this pathogen. Phage therapy is considered to be the most promising alternative for treating CRAB infections. In this study, a novel phage, Ab_WF01, which can lyse clinical CRAB, was isolated and characterized from hospital sewage. The multiplicity of infection, morphology, one-step growth curve, stability, sensitivity, and lytic activity of the phage were also investigated. The genome of phage Ab_WF01 was 41, 317 bp in size with a GC content of 39.12% and encoded 51 open reading frames (ORFs). tRNA, virulence, and antibiotic resistance genes were not detected in the phage genome. Comparative genomic and phylogenetic analyses suggest that phage Ab_WF01 is a novel species of the genus Friunavirus, subfamily Beijerinckvirinae, and family Autographiviridae. The in vivo results showed that phage Ab_WF01 significantly increased the survival rate of CRAB-infected Galleria mellonella (from 0% to 70% at 48 h) and mice (from 0% to 60% for 7 days). Moreover, after day 3 post-infection, phage Ab_WF01 reduced inflammatory response, with strongly ameliorated histological damage and bacterial clearance in infected tissue organs (lungs, liver, and spleen) in mouse CRAB infection model. Taken together, these results show that phage Ab_WF01 holds great promise as a potential alternative agent with excellent stability for against CRAB infections.

Metagenomic analysis of sewage for surveillance of bacterial pathogens: A release experiment to determine sensitivity.

Accurate monitoring of gastro-enteric and other diseases in large populations poses a challenge for public health management. Sewage represents a larger population, is freely obtainable and non-subject to ethical approval. Metagenomic sequencing offers simultaneous, multiple-target analysis. However, no study has demonstrated the sensitivity of metagenomics for detecting bacteria in sewage. In this study, we spot-released 1013 colony-forming units (CFU) of Staphyloccus hyicus (non-pathogenetic strain 842J-88). The strain was flushed down a toilet into the sewer in the catchment area of a public wastewater treatment plant (WWTP), serving a population of 36,000 people. Raw sewage was continuously sampled at the WWTP's inlet over 30- and 60-minute intervals for a total period of seven hours. The experiment was conducted twice with one week in-between release days and under comparable weather conditions. For the metagenomics analyses, the pure single isolate of S. hyicus was sequenced, assembled and added to a large database of bacterial reference sequences. All sewage samples were analyzed by shotgun metagenome sequencing and mapped against the reference database. S. hyicus was identified in duplicate samples at both of two release days and these sequence fragment counts served as a proxy to estimate the minimum number of sick people or sensitivity required in order to observe at least one sick person at 95% probability. We found the sensitivity to be in the range 41-140 and 16-36 sick people at release days 1 and 2, respectively. The WWTP normally serves 36,000 people giving a normalized sensitivity in the range of one in 257 to 2,250 persons.

Yanghanlia caeni gen. nov., sp. nov., a novel taxon within the family Alcaligenaceae isolated from sludge of a pesticide-manufacturing factory.

A Gram-stain-negative bacterium, designated LG-2T, was isolated from sludge collected at a pesticide-manufacturing factory in Jiangsu Province, PR China. Cells of strain LG-2T were strictly aerobic, non-motile and spherical. Growth was observed at 15-42 °C (optimum, 30 °C), pH 6.0-9.0 (optimum, pH 7.0) and 0-3.0 % (w/v) NaCl (optimum, 1.0 %). LG-2T showed 95.5-96.9 % 16S rRNA sequence similarity to type strains in the genera Pusillimonas, Bordetella, Parapusillimonas, Candidimonas and Paracandidimonas of the family Alcaligenaceae. The phylogenomic tree indicated that strain LG-2T was clustered in the family Alcaligenaceae and formed a clade with Paracandidimonas soli IMT-305T, while the phylogenetic trees based on 16S rRNA gene sequences indicated that strain LG-2T formed a distinct clade within the family Alcaligenaceae. The average nucleotide identity, digital DNA-DNA hybridization and average amino acid identity values between LG-2T and its closely related type strains in the genera Pusillimonas, Bordetella, Parapusillimonas, Candidimonas and Paracandidimonas were 70.8-75.3, 18.9-23.7 and 59.6 %-69.3 %, respectively. The major cellular fatty acids were C16 : 0, C17 : 0 cyclo, summed feature 3 (C16 : 1 ω7c and/or C16 : 1 ω6c), summed feature 8 (C18 : 1 ω7c and/or C18 : 1 ω6c) and summed feature 2 (C12 : 0 aldehyde and/or unknown 10.928). The predominant menaquinone was Q-8. The polar lipid profile consisted of phosphatidylethanolamine, phosphatidylglycerol, two aminophospholipids, three aminolipids and nine unknown polar lipids. The genome size of strain LG-2T was 3.2 Mb and the DNA G+C content was 63.4 mol%. On the basis of the phenotypic, phylogenetic and genomic results from this study, strain LG-2T represents a novel species of a new genus in the family Alcaligenaceae, for which the name Yanghanlia caeni gen. nov., sp. nov. is proposed, with strain LG-2T (=KCTC 8084T= CCTCC AB 2023123T) as the type strain.

Comparative study of the removal of sulfate by UASB in light and dark environment.

At present, the application of sewage treatment technologies is restricted by high sulfate concentrations. In the present work, the sulfate removal was biologically treated using an upflow anaerobic sludge blanket (UASB) in the absence/presence of light. First, the start-up of UASB for the sulfate removal was studied in terms of COD degradation, sulfate removal, and effluent pH. Second, the impacts of different operation parameters (i.e., COD/SO42- ratio, temperature and illumination time) on the UASB performance were explored. Third, the properties of sludge derived from the UASB at different time were analyzed. Results show that after 28 days of start-up, the COD removal efficiencies in both the photoreactor and non-photoreactor could reach a range of 85-90% while such reactors could achieve > 90% of sulfate being removed. Besides, higher illumination time could facilitate the removal of pollutants in the photoreactor. To sum up, the present study can provide technical support for the clean removal of sulfate from wastewater using photoreactors.

Genomic analysis of multidrug-resistant Escherichia coli from Urban Environmental water sources in Accra, Ghana, Provides Insights into public health implications.

Wastewater discharge into the environment in resource-poor countries poses a threat to public health. Studies in this area within these countries are limited, and the use of high-throughput whole-genome sequencing technologies is lacking. Therefore, understanding of environmental impacts is inadequate. The present study investigated the antibiotic resistance profiles and diversity of beta-lactamases in Escherichia coli strains isolated from environmental water sources in Accra, Ghana. Microbiological analyses were conducted on wastewater samples from three hospitals, a sewage and wastewater treatment plant, and water samples from two urban surface water bodies. Confirmed isolates (N = 57) were selected for phenotypic antibiotic resistance profiles. Multi-drug-resistant isolates (n = 25) were genome sequenced using Illumina MiSeq sequencing technology and screened for sequence types, antibiotic resistance, virulence and beta-lactamase genes, and mobile genetic elements. Isolates were frequently resistant to ampicillin (63%), meropenem (47%), azithromycin (46%), and sulfamethoxazole-trimethoprim (42%). Twenty different sequence types (STs) were identified, including clinically relevant ones such as ST167 and ST21. Five isolates were assigned to novel STs: ST14531 (n = 2), ST14536, ST14537, and ST14538. The isolates belonged to phylogroups A (52%), B1 (44%), and B2 (4%) and carried ß-lactamase (TEM-1B, TEM-1C, CTX-M-15, and blaDHA-1) and carbapenemase (OXA-1, OXA-181) resistance genes. Dominant plasmid replicons included Col440I (10.2%) and IncFIB (AP001918) (6.8%). Polluted urban environments in Accra are reservoirs for antibiotic-resistant bacteria, posing a substantial public health risk. The findings underscore the need for targeted public health interventions to mitigate the spread of antibiotic-resistant bacteria and protect public health.

Microbiome structure and function in parallel full-scale aerobic granular sludge and activated sludge processes.

Aerobic granular sludge (AGS) and conventional activated sludge (CAS) are two different biological wastewater treatment processes. AGS consists of self-immobilised microorganisms that are transformed into spherical biofilms, whereas CAS has floccular sludge of lower density. In this study, we investigated the treatment performance and microbiome dynamics of two full-scale AGS reactors and a parallel CAS system at a municipal WWTP in Sweden. Both systems produced low effluent concentrations, with some fluctuations in phosphate and nitrate mainly due to variations in organic substrate availability. The microbial diversity was slightly higher in the AGS, with different dynamics in the microbiome over time. Seasonal periodicity was observed in both sludge types, with a larger shift in the CAS microbiome compared to the AGS. Groups important for reactor function, such as ammonia-oxidising bacteria (AOB), nitrite-oxidising bacteria (NOB), polyphosphate-accumulating organisms (PAOs) and glycogen-accumulating organisms (GAOs), followed similar trends in both systems, with higher relative abundances of PAOs and GAOs in the AGS. However, microbial composition and dynamics differed between the two systems at the genus level. For instance, among PAOs, Tetrasphaera was more prevalent in the AGS, while Dechloromonas was more common in the CAS. Among NOB, Ca. Nitrotoga had a higher relative abundance in the AGS, while Nitrospira was the main nitrifier in the CAS. Furthermore, network analysis revealed the clustering of the various genera within the guilds to modules with different temporal patterns, suggesting functional redundancy in both AGS and CAS. KEY POINTS: • Microbial community succession in parallel full-scale aerobic granular sludge (AGS) and conventional activated sludge (CAS) processes. • Higher periodicity in microbial community structure in CAS compared to in AGS. • Similar functional groups between AGS and CAS but different composition and dynamics at genus level.

Pre-exposure of Triclosan compromise tetracycline-derived antibiotic resistance in methanogenic digestion microbiome.

Triclosan (TCS) and tetracycline (TC) are commonly detected antibacterial agents in sewage and environment matrices. Nonetheless, the impact of sequential exposure to TCS and TC on the methanogenic digestion microbiome remains unknown. In this study, TCS was shown to reduce COD removal efficiency to 69.8%, but alleviated the inhibitive effect of consequent TC-amendment on the digestion microbiome. Interestingly, TCS pre-exposure resulted in abundance increase of acetotrophic Methanosaeta to 2.68%, being 2.91 folds higher than that without TCS amendment. Microbial network analyses showed that TCS pre-exposure caused microorganisms to establish a co-ecological relationship against TC disturbance. Further analyses of total antibiotic resistance genes (ARGs) showed the TCS-derived compromise of TC-induced ARGs enrichment in digestion microbiomes, e.g., 238.2% and 152.1% ARGs increase upon TC addition in digestion microbiomes without and with TCS pre-exposure, respectively. This study provides new insights into the impact of antibacterial agents on the methanogenic digestion microbiome.

Taxonomic characterization of Sphaerotilus microaerophilus sp. nov., a sheath-forming microaerophilic bacterium of activated sludge origin.

A microaerophilic Gram-stain-negative bacilliform bacterial strain, FB-5 T, was isolated from activated sludge in Yokohama, Japan, that exhibited filamentous growth and formed a microtube (sheath). Cells were motile using a single polar flagellum. The optimum growth temperature and pH were 30 °C and 7.5, respectively. Strain FB-5 T was catalase-negative. Peptides and amino acids were utilized as energy and carbon sources. Sugars and organic acids were not utilized. Vitamin B12 enhanced the growth of strain FB-5 T. Sulfur-dependent lithotrophic growth was possible. Major respiratory quinone was UQ-8. Major fatty acids were C16:1ω7 and C16:0. The genomic DNA G + C content was 69.16%. Phylogenetic analysis of the 16S rRNA gene suggested that strain FB-5 T belongs to the genus Sphaerotilus. The close relatives were S. natans subsup. sulfidivorans and S. natans subsup. natans with 98.0% and 97.8% similarity based on the 16S rRNA gene analysis, respectively. The genome size (6.06 Mbp) was larger than that (4.39-5.07 Mbp) of the Sphaerotilus strains. The AAI values against the related strains ranged from 71.0 to 72.5%. The range of ANI values was 81.7 - 82.5%. In addition to these distinguishable features of the genome, the core genome and dDDH analyses suggested that this strain is a novel member of the genus Sphaerotilus. Based on its physiological properties and genomic features, strain FB-5 T is considered as a novel species of the genus Sphaerotilus, for which the name S. microaerophilus sp. nov. is proposed. The type strain is FB-5 T (= JCM 35424 T = KACC 23146 T).

Sulfate-reducing bacteria decreases fractional pressure of H2 to accelerate short-chain fatty acids production from waste activated sludge fermentation assisted with zero-valent iron activated sulfite pretreatment.

The production of short-chain fatty acids (SCFAs) is constrained by substrate availability and the increased fractional pressure of H2 emitted by acidogenic/fermentative bacteria during anaerobic fermentation of waste activated sludge (WAS). This study introduced a novel approach employing zero-valent iron (ZVI)-activated sulfite pretreatment combined with H2-consuming sulfate-reducing bacteria (SRB) mediation to improve SCFAs, especially acetate production from WAS fermentation. Experimental results showed that the combined ZVI-activated sulfite and incomplete-oxidative SRB (io-SRB) process achieved a peak SCFAs production of 868.11 mg COD/L, with acetate accounting for 80.55 %, which was 7.90- and 2.18-fold higher than that obtained from raw WAS fermentation, respectively. This could be firstly attributed to the SO4- and OH generated by ZVI-activated sulfite, which significantly promoted WAS decomposition, e.g., soluble proteins and carbohydrates increased 14.3- and 10.8-fold, respectively, over those in raw WAS. The biodegradation of dissolved organic matter was subsequently enhanced by the synergistic interaction and H2 transfer between anaerobic fermentation bacteria (AFB) and io-SRB. The positive and negative correlations among AFB, nitrate-reducing bacteria (NRB) and the io-SRB consortia were revealed by molecular ecological network (MEN) and Mantel test. Moreover, the expression of functional genes was also improved, for instance, in relation to acetate formation, the relative abundances of phosphate acetyltransferase and acetate kinase was 0.002 % and 0.005 % higher than that in the control test, respectively. These findings emphasized the importance of sulfate radicals-based oxidation pretreatment and the collaborative relationships of multifunctional microbes on the value-added chemicals and energy recovery from sludge fermentation.

Removal of pharmaceutical compounds and toxicology study in wastewater using Malaysian fungal Ganoderma lucidum.

Elevated usage of pharmaceutical products leads to the accumulation of emerging contaminants in sewage. In the current work, Ganoderma lucidum (GL) was used to remove pharmaceutical compounds (PCs), proposed as a tertiary method in sewage treatment plants (STPs). The PCs consisted of a group of painkillers (ketoprofen, diclofenac, and dexamethasone), psychiatrists (carbamazepine, venlafaxine, and citalopram), beta-blockers (atenolol, metoprolol, and propranolol), and anti-hypertensives (losartan and valsartan). The performance of 800 mL of synthetic water, effluent STP, and hospital wastewater (HWW) was evaluated. Parameters, including treatment time, inoculum volume, and mechanical agitation speed, have been tested. The toxicity of the GL after treatment is being studied based on exposure levels to zebrafish embryos (ZFET) and the morphology of the GL has been observed via Field Emission Scanning Electron Microscopy (FESEM). The findings conclude that GL can reduce PCs from <10% to >90%. Diclofenac and valsartan are the highest (>90%) in the synthetic model, while citalopram and propranolol (>80%) are in the real wastewater. GL effectively removed pollutants in 48 h, 1% of the inoculum volume, and 50 rpm. The ZFET showed GL is non-toxic (LC50 is 209.95 mg/mL). In the morphology observation, pellets GL do not show major differences after treatment, showing potential to be used for a longer treatment time and to be re-useable in the system. GL offers advantages to removing PCs in water due to their non-specific extracellular enzymes that allow for the biodegradation of PCs and indicates a good potential in real-world applications as a favourable alternative treatment.

The growth and nutrient removal properties of heterotrophic microalgae Chlorella sorokiniana in simulated wastewater containing volatile fatty acids.

To reduce the high cost of organic carbon sources in waste resource utilization in the cultivation of microalgae, volatile fatty acids (VFAs) derived from activated sludge were used as the sole carbon source to culture Chlorella sorokiniana under the heterotrophic cultivation. The addition of VFAs in the heterotrophic condition enhanced the total nitrogen (TN) and phosphorus (TP) removal of C. sorokiniana, which proved the advantageous microalgae in using VFAs in the heterotrophic culture after screening in the previous study. To discover the possible mechanism of nitrogen and phosphorus adsorption in heterotrophic conditions by microalgae, the effect of different ratios of VFAs (acetic acid (AA): propionic acid (PA): butyric acid (BA)) on the nutrient removal and growth properties of C. sorokiniana was studied. In the 8:1:1 group, the highest efficiency (77.19%) of VFAs assimilation, the highest biomass (0.80 g L-1) and lipid content (31.35%) were achieved, with the highest TN and TP removal efficiencies of 97.44 % and 91.02 %, respectively. Moreover, an aerobic denitrifying bacterium, Pseudomonas, was determined to be the dominant genus under this heterotrophic condition. This suggested that besides nitrate uptake and utilization by C. sorokiniana under the heterotrophy, the conduct of the denitrification process was also the main reason for obtaining high nitrogen removal efficiency.

The ecology of the sewer systems: Microbial composition, function, assembly, and network in different spatial locations.

Microbial induced concrete corrosion (MICC) is the primary deterioration affecting global sewers. Disentangling ecological mechanisms in the sewer system is meaningful for implementing policies to protect sewer pipes using trenchless technology. It is necessary to understand microbial compositions, interaction networks, functions, alongside assembly processes in sewer microbial communities. In this study, sewer wastewater samples and microbial samples from the upper part (UP), middle part (MP) and bottom part (BP) of different pipes were collected for 16S rRNA gene amplicon analysis. It was found that BP harbored distinct microbial communities and the largest proportion of unique species (1141) compared to UP and MP. The community in BP tended to be more clustered. Furthermore, significant differences in microbial functions existed in different spatial locations, including the carbon cycle, nitrogen cycle and sulfur cycle. Active microbial sulfur cycling indicated the corrosion risk of MICC. Among the environmental factors, the oxidation‒reduction potential drove changes in BP, while sulfate managed changes in UP and BP. Stochasticity dominated community assembly in the sewer system. Additionally, the sewer microbial community exhibited numerous positive links. BP possessed a more complex, modular network with higher modularity. These deep insights into microbial ecology in the sewer system may guide engineering safety and disaster prevention in sewer infrastructure.

Methylosinus trichosporium OB3b bioaugmentation unleashes polyhydroxybutyrate-accumulating potential in waste-activated sludge.

BACKGROUND: Wastewater treatment plants contribute approximately 6% of anthropogenic methane emissions. Methanotrophs, capable of converting methane into polyhydroxybutyrate (PHB), offer a promising solution for utilizing methane as a carbon source, using activated sludge as a seed culture for PHB production. However, maintaining and enriching PHB-accumulating methanotrophic communities poses challenges. RESULTS: This study investigated the potential of Methylosinus trichosporium OB3b to bioaugment PHB-accumulating methanotrophic consortium within activated sludge to enhance PHB production. Waste-activated sludges with varying ratios of M. trichosporium OB3b (1:0, 1:1, 1:4, and 0:1) were cultivated. The results revealed substantial growth and methane consumption in waste-activated sludge with M. trichosporium OB3b-amended cultures, particularly in a 1:1 ratio. Enhanced PHB accumulation, reaching 37.1% in the same ratio culture, indicates the dominance of Type II methanotrophs. Quantification of methanotrophs by digital polymerase chain reaction showed gradual increases in Type II methanotrophs, correlating with increased PHB production. However, while initial bioaugmentation of M. trichosporium OB3b was observed, its presence decreased in subsequent cycles, indicating the dominance of other Type II methanotrophs. Microbial community analysis highlighted the successful enrichment of Type II methanotrophs-dominated cultures due to the addition of M. trichosporium OB3b, outcompeting Type I methanotrophs. Methylocystis and Methylophilus spp. were the most abundant in M. trichosporium OB3b-amended cultures. CONCLUSIONS: Bioaugmentation strategies, leveraging M. trichosporium OB3b could significantly enhance PHB production and foster the enrichment of PHB-accumulating methanotrophs in activated sludge. These findings contribute to integrating PHB production in wastewater treatment plants, providing a sustainable solution for resource recovery.

The risk of viable but non-culturable (VBNC) enterococci and antibiotic resistance transmission during simulated municipal sludge composting.

Sludge composting is a sludge resource utilization method that can reduce pollutants, such as pathogens. Enterococci are regarded as more reliable and conservative indicators of pathogen inactivation than fecal coliforms, which are typically used as indicators of fecal pollution. Non-spore pathogenic bacteria may enter a viable but non-culturable (VBNC) state during composting, leading to residual risk. The VBNC status of bacteria is related to their survival during composting. However, the survival mechanisms of enterococci during sludge composting remain unclear. Therefore, this study aimed to investigate the VBNC state of enterococci in different phases of simulated sludge composting and the fate of antibiotic resistance genes (ARGs) and mobile genetic elements (MGEs) during the composting process. This study is expected to provide a basis for subsequent exploration of possible methods to completely inactivate enterococci and reduce ARGs during sludge composting. Culturable enterococci were reduced in the thermophilic phase of sludge composting, but the proportion of VBNC subpopulation increased. It was reported for the first time that most VBNC enterococci were killed by extending the cooling phase of sludge compost, and by prolonging the cooling phase the types of ARG were reduced. However, there was a certain quantity (approximately 104/g dry weight) of culturable and VBNC enterococci in the compost products. In addition, MGEs and ARGs exist in both bacteria and compost products, leading to the risk of spreading antibiotic-resistant bacteria and antibiotic resistance when sludge compost products are used.

Unveiling long-term combined effect of salinity and Lead(II) on anammox activity and microbial community dynamics in saline wastewater treatment.

The study assessed the effect of salinity and lead (Pb(II)) on the anammox sludge for nitrogen removal from saline wastewater. Results showed decreased nitrogen removal and specific anammox activity (SAA) with elevated salinity and Pb(II). SAA reduced from 541.3 ± 4.3 mg N g-1 VSS d-1 at 0.5 mg/L Pb(II) to 436.0 ± 0.2 mg N g-1 VSS d-1 at 30 g/L NaCl, further to 303.6 ± 7.1 mg N g-1 VSS d-1 under 30 g/L NaCl + 0.5 mg/L Pb(II). Notably, the combined inhibition at salinity (15-20 g/L NaCl) and Pb(II) (0.3-0.4 mg/L) exhibited synergistic effect, while higher salinity and Pb(II) aligned with independent inhibition models. Combined inhibition decreased protein/polysaccharides ratio, indicating more severe negative effect on anammox aggregation capacity. Metagenomics confirmed decreased Candidatus Kuenenia, and enhanced denitrification under elevated salinity and Pb(II) conditions. This study offers insights into anammox operation for treating saline wastewater with heavy metals.

Assessing microbial stability and predicting biogas production in full-scale thermophilic dry methane fermentation of municipal solid waste.

Compared to typical anaerobic digestion processes, little is known about both sludge microbial compositions and biogas production models for full-scale dry methane fermentation treating municipal solid waste (MSW). The anaerobic sludge composed of one major hydrogenotrophic methanogen (Methanoculleus) and syntrophic acetate oxidizing bacteria (e.g., Caldicoprobacter), besides enrichment of MSW degraders such as Clostridia. The core population remained phylogenetically unchanged during the fermentation process, regardless of amounts of MSW supplied (∼35 ton/d) or biogas produced (∼12000 Nm3/d). Based on the correlations observed between feed amounts of MSW from 6 days in advance to the current day and biogas output (the strongest correlation: r = 0.77), the best multiple linear regression (MLR) model incorporating the temperature factor was developed with a good prediction for validation data (R2 = 0.975). The proposed simple MLR method with only data on the feedstock amounts will help decision-making processes to prevent low-efficient biogas production.

Performance and microbial community composition of full-scale high-rate cascade sludge digestion system via pie-shaped reactor configuration.

A full-scale high-rate cascade anaerobic digestion (CAD) system was evaluated for its ability to enhance enzymatic sludge hydrolysis. The system included a newly built digester, innovatively divided into three pie-shaped compartments (500 m3 each), followed by an existing, larger digester (1500 m3). The system treated a mixture of waste activated sludge and primary sludge, achieving a stable total chemical oxygen demand reduction efficiency (56.1 ± 6.8 %), and enhanced sludge hydrolytic enzyme activities at a 14.5-day total solids retention time (SRT). High-throughput sequencing data revealed a consistent microbial community across reactors, dominated by consortia that govern hydrolysis and acidogenesis. Despite relatively short SRTs in the initial reactors of the CAD system, acetoclastic methanogens belonging to Methanosaeta became the most abundant archaea. ‬‬‬‬‬‬‬‬‬‬‬‬‬ This study proves that the CAD system achieves stable sludge reduction, accelerates enzymatic hydrolysis at full-scale, and paves the way for its industrialization in municipal waste sewage sludge treatment.