A fluorescent sensor array based on antibiotic-stabilized metal nanoclusters for the multiplex detection of bacteria.

To address the need for facile, rapid detection of pathogens in water supplies, a fluorescent sensing array platform based on antibiotic-stabilized metal nanoclusters was developed for the multiplex detection of pathogens. Using five common antibiotics, eight different nanoclusters (NCs) were synthesized including ampicillin stabilized copper NCs, cefepime stabilized gold and copper NCs, kanamycin stabilized gold and copper NCs, lysozyme stabilized gold NCs, and vancomycin stabilized gold/silver and copper NCs. Based on the different interaction of each NC with the bacteria strains, unique patterns were generated. Various machine learning algorithms were employed for pattern discernment, among which the artificial neural networks proved to have the highest performance, with an accuracy of 100%. The developed prediction model performed well on an independent test dataset and on real samples gathered from drinking water, tap water and the Anzali Lagoon water, with prediction accuracy of 96.88% and 95.14%, respectively. This work demonstrates how generic antibiotics can be implemented for NC synthesis and used as recognition elements for pathogen detection. Furthermore, it displays how merging machine learning techniques can elevate sensitivity of analytical devices.

Microbiological assessment of ready-to-eat foods and drinking water sources as a potential vehicle of bacterial pathogens in northern India.

Foodborne illnesses caused by the consumption of contaminated foods have frequent occurrences in developing countries. The incorporation of contaminated water in food processes, preparation, and serving is directly linked to several gastrointestinal infections. Keeping in view, this study was conducted to assess the microbial quality of both drinking water sources and commonly consumed fresh ready-to-eat (RTE) foods in the region. The drinking water samples from water sources and consumer points, as well as food samples from canteens, cafes, hotels, and restaurants, were collected for the microbiological analysis. Fifty-five percent (n = 286) of water samples were found to be positive for total coliforms with MPN counts ranging from 3 to 2600 (100 ml) -1. E. coli was detected in nearly 30% of the total water samples. Overall, 65% tap water samples were found unsatisfactory, followed by submersible (53%), filter (40%), and WTP (30%) sources. Furthermore, the examination of RTE foods (n = 80) found that 60% were of unsatisfactory microbial quality with high aerobic plate counts. The salads were the most contaminated category with highest mean APC 8.3 log CFU/g followed by pani puri, chats, and chutneys. Presence of coliforms and common enteropathogens was observed in both water and food samples. The detected isolates from the samples were identified as Enterobacter spp., Klebsiella spp., Pseudomonas aeruginosa, Salmonella spp., Shigella spp., and Staphylococcus spp. Based on these findings, microbiological quality was found compromised and this may pose hazard to public health. This exploratory study in the Punjab region also suggests that poor microbiological quality of water sources can be an important source of contamination for fresh uncooked RTE foods, thus transferring pathogens to the food chain. Therefore, only safe potable drinking water post-treatment should be used at all stages.

Genetic Diversity of Legionella pneumophila Isolates from Artificial Water Sources in Brazil.

Legionella pneumophila (Lp) is a Gram-negative bacterium found in natural and artificial aquatic environments and inhalation of contaminated aerosols can cause severe pneumonia known as Legionnaires' Disease (LD). In Brazil there is hardly any information about this pathogen, so we studied the genetic variation of forty Legionella spp. isolates obtained from hotels, malls, laboratories, retail centers, and companies after culturing in BCYE medium. These isolates were collected from various sources in nine Brazilian states. Molecular identification of the samples was carried out using Sequence-Based Typing (SBT), which consists of sequencing and analysis of seven genes (flaA, pilE, asd, mip, mompS, proA, and neuA) to define a Sequence Type (ST). Eleven STs were identified among 34/40 isolates, of which eight have been previously described (ST1, ST80, ST152, ST242, ST664, ST1185, ST1464, ST1642) and three were new STs (ST2960, ST2962, and ST2963), the former identified in five different cooling towers in the city of São Paulo. The ST1 that is widely distributed in many countries was also the most prevalent in this study. In addition, other STs that we observed have also been associated with legionellosis in other countries, reinforcing the potential of these isolates to cause LD in Brazil. Unfortunately, no human isolates could be characterized until presently, but our observations strongly suggest the need of surveillance implementation system and control measures of Legionella spp. in Brazil, including the use of more sensitive genotyping procedures besides ST.

Synthesis of recovery patterns in microbial communities across environments.

BACKGROUND: Disturbances alter the diversity and composition of microbial communities. Yet a generalized empirical assessment of microbiome responses to disturbance across different environments is needed to understand the factors driving microbiome recovery, and the role of the environment in driving these patterns. RESULTS: To this end, we combined null models with Bayesian generalized linear models to examine 86 time series of disturbed mammalian, aquatic, and soil microbiomes up to 50 days following disturbance. Overall, disturbances had the strongest effect on mammalian microbiomes, which lost taxa and later recovered their richness, but not their composition. In contrast, following disturbance, aquatic microbiomes tended away from their pre-disturbance composition over time. Surprisingly, across all environments, we found no evidence of increased compositional dispersion (i.e., variance) following disturbance, in contrast to the expectations of the Anna Karenina Principle. CONCLUSIONS: This is the first study to systematically compare secondary successional dynamics across disturbed microbiomes, using a consistent temporal scale and modeling approach. Our findings show that the recovery of microbiomes is environment-specific, and helps to reconcile existing, environment-specific research into a unified perspective. Video Abstract.

Development, confirmation, and application of a seeded Escherichia coli process control organism to validate Salmonella enterica serovar Typhi environmental surveillance methods.

Salmonella enterica serovar Typhi (S. Typhi) is the causative agent of Typhoid fever. Blood culture is the gold standard for clinical diagnosis, but this is often difficult to employ in resource limited settings. Environmental surveillance of waste-impacted waters is a promising supplement to clinical surveillance, however validating methods is challenging in regions where S. Typhi concentrations are low. To evaluate existing S. Typhi environmental surveillance methods, a novel process control organism (PCO) was created as a biosafe surrogate. Using a previous described qPCR assay, a modified PCR amplicon for the staG gene was cloned into E. coli. We developed a target region that was recognized by the Typhoid primers in addition to a non-coding internal probe sequence. A multiplex qPCR reaction was developed that differentiates between the typhoid and control targets, with no cross-reactivity or inhibition of the two probes. The PCO was shown to mimic S. Typhi in lab-based experiments with concentration methods using primary wastewater: filter cartridge, recirculating Moore swabs, membrane filtration, and differential centrifugation. Across all methods, the PCO seeded at 10 CFU/mL and 100 CFU/mL was detected in 100% of replicates. The PCO is detected at similar quantification cycle (Cq) values across all methods at 10 CFU/mL (Average = 32.4, STDEV = 1.62). The PCO was also seeded into wastewater at collection sites in Vellore (India) and Blantyre (Malawi) where S. Typhi is endemic. All methods tested in both countries were positive for the seeded PCO. The PCO is an effective way to validate performance of environmental surveillance methods targeting S. Typhi in surface water.

Shiga Toxin-Producing Escherichia coli O157:H7 Illness Outbreak Associated with Untreated, Pressurized, Municipal Irrigation Water - Utah, 2023.

During July-September 2023, an outbreak of Shiga toxin-producing Escherichia coli O157:H7 illness among children in city A, Utah, caused 13 confirmed illnesses; seven patients were hospitalized, including two with hemolytic uremic syndrome. Local, state, and federal public health partners investigating the outbreak linked the illnesses to untreated, pressurized, municipal irrigation water (UPMIW) exposure in city A; 12 of 13 ill children reported playing in or drinking UPMIW. Clinical isolates were genetically highly related to one another and to environmental isolates from multiple locations within city A's UPMIW system. Microbial source tracking, a method to indicate possible contamination sources, identified birds and ruminants as potential sources of fecal contamination of UPMIW. Public health and city A officials issued multiple press releases regarding the outbreak reminding residents that UPMIW is not intended for drinking or recreation. Public education and UPMIW management and operations interventions, including assessing and mitigating potential contamination sources, covering UPMIW sources and reservoirs, indicating UPMIW lines and spigots with a designated color, and providing conspicuous signage to communicate risk and intended use might help prevent future UPMIW-associated illnesses.

Detection and modeling of Staphylococcus aureus and fecal bacteria in Hawaiian coastal waters and sands.

Microbial pollution of recreational waters leads to millions of skin, respiratory, and gastrointestinal illnesses globally. Fecal indicator bacteria (FIB) are monitored to assess recreational waters but may not reflect the presence of Staphylococcus aureus, a global leader in bacterial fatalities. Since many community-acquired S. aureus skin infections are associated with high recreational water usage, this study measured and modeled S. aureus, methicillin-resistant S. aureus (MRSA), and FIB (Enterococcus spp., Clostridium perfringens) concentrations in seawater and sand at six beaches in Hilo, Hawai'i, USA, over 37 sample dates from July 2016 to February 2019 using culturing techniques. Generalized linear models predicted bacterial concentrations with physicochemical and environmental data. Beach visitors were also surveyed on their preferred activities. S. aureus and FIB concentrations were roughly 6-78 times higher at beaches with freshwater discharge than at those without. Seawater concentrations of Enterococcus spp. were positively associated with MRSA but not S. aureus. Elevated S. aureus was associated with lower tidal heights, higher freshwater discharge, onsite sewage disposal system density, and turbidity. Regular monitoring of beaches with freshwater input, utilizing real-time water quality measurements with robust modeling techniques, and raising awareness among recreational water users may mitigate exposure to S. aureus, MRSA, and FIB. PRACTITIONER POINTS: Staphylococcus aureus and fecal bacteria concentrations were higher in seawater and sand at beaches with freshwater discharge. In seawater, Enterococcus spp. positively correlated with MRSA, but not S. aureus. Freshwater discharge, OSDS density, water turbidity, and tides significantly predicted bacterial concentrations in seawater and sand. Predictive bacterial models based upon physicochemical and environmental data developed in this study are readily available for user-friendly application.

Metagenomics datasets of water and sediments from eutrophication-impacted artificial lakes in South Africa.

We present metagenomes of 16 samples of water and sediment from two lakes, collected from eutrophic and non-eutrophic areas, including pooled samples enriched with phosphate and nitrate. Additionally, we assembled 167 bacterial metagenome-assembled genomes (MAGs). These MAGs were de-replicated into 83 unique genomes representing different species found in the lakes. All the MAGs exhibited >70% completeness and <10% contamination, with 79 MAGs being classified as 'nearly complete' (completeness >90%), while 54 falling within 80-90% range and 34 between 75-80% complete. The most abundant MAGs identified across all samples were Proteobacteria (n = 80), Firmicutes_A (n = 35), Firmicutes (n = 13), and Bacteriodota (n = 22). Other groups included Desulfobacteria_I (n = 2), Verrucomicrobiota (n = 4), Campylobacterota (n = 4) and Actinobacteriota (n = 6). Importantly, phylogenomic analysis identified that approximately 50.3% of the MAGs could not be classified to known species, suggesting the presence of potentially new and unknown bacteria in these lakes, warranting further in-depth investigation. This study provides valuable new dataset on the diverse and often unique microbial communities living in polluted lakes, useful in developing effective strategies to manage pollution.

Bacterial community profiles within the water samples of leptospirosis outbreak areas.

Background: Leptospirosis is a water-related zoonotic disease. The disease is primarily transmitted from animals to humans through pathogenic Leptospira bacteria in contaminated water and soil. Rivers have a critical role in Leptospira transmissions, while co-infection potentials with other waterborne bacteria might increase the severity and death risk of the disease. Methods: The water samples evaluated in this study were collected from four recreational forest rivers, Sungai Congkak, Sungai Lopo, Hulu Perdik, and Gunung Nuang. The samples were subjected to next-generation sequencing (NGS) for the 16S rRNA and in-depth metagenomic analysis of the bacterial communities. Results: The water samples recorded various bacterial diversity. The samples from the Hulu Perdik and Sungai Lopo downstream sampling sites had a more significant diversity, followed by Sungai Congkak. Conversely, the upstream samples from Gunung Nuang exhibited the lowest bacterial diversity. Proteobacteria, Firmicutes, and Acidobacteria were the dominant phyla detected in downstream areas. Potential pathogenic bacteria belonging to the genera Burkholderiales and Serratia were also identified, raising concerns about co-infection possibilities. Nevertheless, Leptospira pathogenic bacteria were absent from all sites, which is attributable to its limited persistence. The bacteria might also be washed to other locations, contributing to the reduced environmental bacterial load. Conclusion: The present study established the presence of pathogenic bacteria in the river ecosystems assessed. The findings offer valuable insights for designing strategies for preventing pathogenic bacteria environmental contamination and managing leptospirosis co-infections with other human diseases. Furthermore, closely monitoring water sample compositions with diverse approaches, including sentinel programs, wastewater-based epidemiology, and clinical surveillance, enables disease transmission and outbreak early detections. The data also provides valuable information for suitable treatments and long-term strategies for combating infectious diseases.

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.

Standard addition method for rapid, cultivation-independent quantification of Legionella pneumophila cells by qPCR in biotrickling filters.

Cultivation-independent molecular biological methods are essential to rapidly quantify pathogens like Legionella pneumophila (L. pneumophila) which is important to control aerosol-generating engineered water systems. A standard addition method was established to quantify L. pneumophila in the very complex matrix of process water and air of exhaust air purification systems in animal husbandry. Therefore, cryopreserved standards of viable L. pneumophila were spiked in air and water samples to calibrate the total bioanalytical process which includes cell lysis, DNA extraction, and qPCR. A standard addition algorithm was employed for qPCR to determine the initial concentration of L. pneumophila. In mineral water, the recovery rate of this approach (73%-134% within the concentration range of 100-5000 Legionella per mL) was in good agreement with numbers obtained from conventional genomic unit (GU) calibration with DNA standards. In air samples of biotrickling filters, in contrast, the conventional DNA standard approach resulted in a significant overestimation of up to 729%, whereas our standard addition gave a more realistic recovery of 131%. With this proof-of-principle study, we were able to show that the molecular biology-based standard addition approach is a suitable method to determine realistic concentrations of L. pneumophila in air and process water samples of biotrickling filter systems. Moreover, this quantification strategy is generally a promising method to quantify pathogens in challenging samples containing a complex microbiota and the classical GU approach used for qPCR leads to unreliable results.

Plastiome: Plastisphere-enriched mobile resistome in aquatic environments.

Aquatic microplastics (MPs) act as reservoirs for microbial communities, fostering the formation of a mobile resistome encompassing diverse antibiotic (ARGs) and biocide/metal resistance genes (BMRGs), and mobile genetic elements (MGEs). This collective genetic repertoire, referred to as the "plastiome," can potentially perpetuate environmental antimicrobial resistance (AMR). Our study examining two Japanese rivers near Tokyo revealed that waterborne MPs are primarily composed of polyethylene and polypropylene fibers and sheets of diverse origin. Clinically important genera like Exiguobacterium and Eubacterium were notably enriched on MPs. Metagenomic analysis uncovered a 3.46-fold higher enrichment of ARGs on MPs than those in water, with multidrug resistance genes (MDRGs) and BMRGs prevailing, particularly within MPs. Specific ARG and BMRG subtypes linked to resistance to vancomycin, beta-lactams, biocides, arsenic, and mercury showed selective enrichment on MPs. Network analysis revealed intense associations between host genera with ARGs, BMRGs, and MGEs on MPs, emphasizing their role in coselection. In contrast, river water exhibited weaker associations. This study underscores the complex interactions shaping the mobile plastiome in aquatic environments and emphasizes the global imperative for research to comprehend and effectively control AMR within the One Health framework.

Site-specific risk-based threshold (RBT) concentrations for sewage-associated markers in estuarine swimming waters.

This study establishes site-specific risk-based threshold (RBT) concentrations for sewage-associated markers, including Bacteroides HF183 (HF183), Lachnospiraceae Lachno3 (Lachno3), cross-assembly phage (CrAssphage), and pepper mild mottle virus (PMMoV), utilizing quantitative microbial risk assessment (QMRA) for recreational estuarine waters (EW). The QMRA model calculates a RBT concentration corresponding to a selected target illness risk for ingestion of EW contaminated with untreated sewage. RBT concentrations were estimated considering site-specific decay rates and concentrations of markers and reference pathogen (human norovirus; HNoV), aiding in the identification of high-risk days during the swimming season. Results indicated varying RBT concentrations for fresh (Day 0) and aged (Days 1 to 10) sewage contamination scenarios over 10 days. HF183 exhibited the highest RBT concentration (26,600 gene copis (GC)/100 mL) initially but decreased rapidly with aging (2570 to 3120 GC/100 mL on Day 10) depending on the decay rates, while Lachno3 and CrAssphage remained relatively stable. PMMoV, despite lower initial RBT (3920 GC/100 mL), exhibited increased RBT (4700 to 6440 GC/100 mL) with aging due to its slower decay rate compared to HNoV. Sensitivity analysis revealed HNoV concentrations as the most influential parameter. Comparison of marker concentrations in estuarine locations with RBT concentrations showed instances of marker exceedance, suggesting days of potential higher risks. The observed discrepancies between bacterial and viral marker concentrations in EW highlight the need for optimized sample concentration method and simultaneous measurement of multiple markers for enhanced risk predictions. Future research will explore the utility of multiple markers in risk management. Overall, this study contributes to better understanding human health risks in recreational waters, aiding regulators, and water quality managers in effective decision-making for risk prioritization and mitigation strategies.

Phenotypic and Genotypic Methods for the Identification and Quantification of Cyanobacteria in Lake Water.

Early monitoring of Microcystis, a cyanobacterium that produces microcystin, is paramount in order to confirm the presence of Microcystis spp. Both phenotypic and genotypic methods have been used. The phenotypic methods provide the presence of the microcystis but do not confirm its species type and toxin produced. Additionally, phenotypic methods cannot differentiate toxigenic from non-toxigenic Microcystis. Therefore, the current protocol also describes genetic methods based on PCR to detect toxigenic Microcystis spp. based on microcystin synthetase E (mcy E) gene and 16-23S RNA genes for species-specific identification, which can effectively comprehend distinct lineages and discrimination of potential complexity of microcystin populations. The presence of these microcystin toxins in blood, in most cases, indicates contamination of drinking water by cyanobacteria. The methods presented herein are used to identify microcystin toxins in drinking water and blood.

Agricultural and urban practices are correlated to changes in the resistome of riverine systems.

The objective of this study was to comprehensively characterise the resistome, the collective set of antimicrobial resistance genes in a given environment, of two rivers, from their source to discharge into the sea, as these flow through areas of different land use. Our findings reveal significant differences in the riverine resistome composition in areas of different land uses, with increased abundance and diversity of AMR in downstream agricultural and urban locations, with the resistome in urban areas more similar to the resistome in wastewater. The changes in resistome were accompanied by changes in microbial communities, with a reduction in microbial diversity in downstream agricultural and urban affected areas, driven mostly by increased relative abundance in the phyla, Bacteroidetes and Proteobacteria. These results provide insight into how pollution associated with agricultural and urban activities affects microbial communities and influences AMR in aquatic water bodies. These results add valuable insights to form effective strategies for mitigating and preserving aquatic ecosystems. Overall, our study highlights the critical role of the environment in the development and dissemination of AMR and underscores the importance of adopting a One Health approach to address this global public health threat.

Using a combination of quantitative culture, molecular, and infrastructure data to rank potential sources of fecal contamination in Town Creek Estuary, North Carolina.

Estuarine water quality is declining worldwide due to increased tourism, coastal development, and a changing climate. Although well-established methods are in place to monitor water quality, municipalities struggle to use the data to prioritize infrastructure for monitoring and repair and to determine sources of contamination when they occur. The objective of this study was to assess water quality and prioritize sources of contamination within Town Creek Estuary (TCE), Beaufort, North Carolina, by combining culture, molecular, and geographic information systems (GIS) data into a novel contamination source ranking system. Water samples were collected from TCE at ten locations on eight sampling dates in Fall 2021 (n = 80). Microbiological water quality was assessed using US Environmental Protection Agency (U.S. EPA) approved culture-based methods for fecal indicator bacteria (FIB), including analysis of total coliforms (TC), Escherichia coli (EC), and Enterococcus spp. (ENT). The quantitative microbial source tracking (qMST) human-associated fecal marker, HF183, was quantified using droplet digital PCR (ddPCR). This information was combined with environmental data and GIS information detailing proximal sewer, septic, and stormwater infrastructure to determine potential sources of fecal contamination in the estuary. Results indicated FIB concentrations were significantly and positively correlated with precipitation and increased throughout the estuary following rainfall events (p < 0.01). Sampling sites with FIB concentrations above the U.S. EPA threshold also had the highest percentages of aged, less durable piping materials. Using a novel ranking system combining concentrations of FIB, HF183, and sewer infrastructure data at each site, we found that the two sites nearest the most aged sewage infrastructure and stormwater outflows were found to have the highest levels of measurable fecal contamination. This case study supports the inclusion of both traditional water quality measurements and local infrastructure data to support the current need for municipalities to identify, prioritize, and remediate failing infrastructure.

Occurrence of coliforms and biofilm-forming bacteria in raw, treated, and distributed water from two waterwork systems in Osun State, Southwestern Nigeria.

This study assessed the bacteriological quality of raw, treated, and distributed water from Ede-Erinle and Opa reservoirs in Osun State, Nigeria. This was to determine the potability of water from these waterwork stations. Eighteen sampling points were established across the two reservoir networks for this study. Samples were collected bi-monthly for two annual cycles. Serial dilution and pour plate methods were employed for the enumeration of bacterial load. Total heterotrophic bacteria count (THBC) and total coliform bacteria count (TCBC) were enumerated on nutrient and MacConkey agar at 37 °C, respectively. Bacterial isolates were characterized using biochemical identification methods with reference to Bergey's Manual of Determinative Bacteriology. Bacterial isolates and biofilm formation were further identified molecularly through the PCR method using specific universal primers. Mean values of THBC and TCBC in distributed water from Ede-Erinle (9.61 × 104 ± 1.50 × 104 CFU/mL; 69.56 ± 26.81 CFU/mL) and Opa waterworks (9.58 × 104 ± 2.55 × 104 CFU/mL; 142.94 ± 44.41 CFU/mL) exceeded permissible limits for drinking water. Paenibacillus lautus, Bacillus pseudomycoides, Pseudomonas aeruginosa, and Pseudomonas stutzeri showed biofilm-forming capacity. The study concluded that the presence of coliforms and biofilm-forming bacteria in distributed water implies that the water is unfit for consumption without further treatment.

Occurrence of sulfate-reducing bacteria in well water: identification of anaerobic sulfidogenic bacterial enrichment cultures.

Bacteriological studies of well water mainly focus on aerobic and facultative aerobic coliform bacteria. However, the presence of obligate anaerobic bacteria in well water, especially sulfate-reducing bacteria (SRB), possible causative agents of some diseases, is often ignored. In this study, the presence of SRB and coexisting anaerobic bacteria with SRB in sulfate-reducing enrichment cultures obtained from 10 well water samples in Istanbul was investigated. A nested polymerase chain reaction-denaturing gradient gel electrophoresis strategy was performed to characterize the bacterial community structure of the enrichments. The most probable number method was used to determine SRB number. Out of 10, SRB growth was observed in only one (10%) enrichment culture and the SRB number was low (<10 cells/mL). Community members were identified as Desulfolutivibrio sulfodismutans and Anaerosinus sp. The results show that SRB coexist with Anaerosinus sp., and this may indicate poor water quality, posing a risk to public health. Furthermore, Anaerosinus sp., found in the human intestinal tract, may be used as an alternative anaerobic fecal indicator. It is worth noting that the detection of bacteria using molecular analyzes following enrichment culture techniques can bring new perspectives to determine the possible origin and presence of alternative microbial indicators in aquatic environments.

Is redox zonation an appropriate method for determining the stage of natural remediation in deep contaminated groundwater?

Groundwater contamination resulting from petroleum development poses a significant threat to drinking water sources, especially in developing countries. In situ natural remediation methods, including microbiological processes, have gained popularity for the reduction of groundwater contaminants. However, assessing the stage of remediation in deep contaminated groundwater is challenging and costly due to the complexity of diverse geological conditions and unknown initial concentrations of contaminants. This research proposes that redox zonation may be a more convenient and comprehensive indicator than the concentration of contaminants for determining the stage of natural remediation in deep groundwater. The combination of sequencing microbial composition using the high-throughput 16S rRNA gene and function predicted by FAPROTAX is a useful approach to determining the redox conditions of different contaminated groundwater. The sulfate-reducing environment, represented by Desulfobacteraceae, Peptococcaceae, Desulfovibrionaceae, and Desulfohalobiaceae could be used as characteristic early stages of remediation for produced water contamination in wells with high concentrations of SO42-, benzene, and salinity. The nitrate-reducing environment, enriched with microorganisms related to denitrification, sulfur-oxidizing, and methanophilic microorganisms could be indicative of the mid stages of in situ bioremediation. The oxygen reduction environment, enriched with oligotrophic and pathogenic Sphingomonadaceae, Caulobacteraceae, Syntrophaceae, Legionellales, Moraxellaceae, and Coxiellaceae, could be indicative of the late stages of remediation. This comprehensive approach could provide valuable insights into the process of natural remediation and facilitate improved environmental management in areas of deep contaminated groundwater.