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.

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.

Innovative DIY drinking water disinfection for underserved communities.

Waterborne pathogens threaten 2.2 billion people lacking access to safely managed drinking water services, causing over a million annual diarrheal deaths. Individuals without access to chlorine reagents or filtration devices often resort to do-it-yourself (DIY) methods, such as boiling or solar disinfection (SODIS). However, these methods are not simple to implement. In this study, we introduced an innovative and easily implemented disinfection approach. We discovered that immersing aluminum foil in various alkaline solutions produces alkali-treated aluminum foil (ATA foil) that effectively adsorbs Escherichia coli (E. coli), Salmonella, and Acinetobacter through the generated surface aluminum hydroxide. For example, a 25 cm2 ATA foil efficiently captures all 104E. coli DH5α strains in 100 mL water within 30 min. Using a saturated suspension of magnesium hydroxide, a type of fertilizer, as the alkaline solution, the properties of the saturated suspension eliminate the need for measuring reagents or changing solutions, making it easy for anyone to create ATA foil. ATA foils can be conveniently produced within mesh bags and placed in household water containers, reducing the risk of recontamination. Replacing the ATA foil with a foil improves the adsorption efficiency, and re-immersing the used foil in the production suspension restores its adsorption capacity. Consequently, ATA foil is an accessible and user-friendly alternative DIY method for underserved communities. Verification experiments covering variations in the water quality and climate are crucial for validating the efficacy of the foil. Fortunately, the ATA foil, with DIY characteristics similar to those of boiling and SODIS, is well-suited for testing under diverse global conditions, offering a promising solution for addressing waterborne pathogens worldwide.

Spread of antibiotic resistance genes in drinking water reservoirs: Insights from a deep metagenomic study using a curated database.

The exploration of antibiotic resistance genes (ARGs) in drinking water reservoirs is an emerging field. Using a curated database, we enhanced the ARG detection and conducted a comprehensive analysis using 2.2 Tb of deep metagenomic sequencing data to determine the distribution of ARGs across 16 drinking water reservoirs and associated environments. Our findings reveal a greater diversity of ARGs in sediments than in water, underscoring the importance of extensive background surveys. Crucial ARG carriers-specifically Acinetobacter, Pseudomonas, and Mycobacterium were identified in drinking water reservoirs. Extensive analysis of the data uncovered a considerable concern for drinking water safety, particularly in regions reliant on river sources. Mobile genetic elements have been found to contribute markedly to the propagation of ARGs. The results of this research suggest that the establishment of drinking water reservoirs for supplying raw water may be an effective strategy for alleviating the spread of water-mediated ARGs.

Influence of using synbiotics by various routes on Mandarah male chicks: intestinal bacterial counts, gut morphology and histological status.

This experiment investigated the influence of different synbiotic processing methods on the intestinal bacterial count, morphology and histological status of developed male Mandarah chicks. Two hundred and ten male Mandarah line chicks aged 1 d were randomized to receive one of 7 chicks. The method and dose for 1-time synbiotics administration to the day-old chicks were as follows: G1: chicks on basal diet received no treatment (control); G2: 0.25 mL synbiotics sprayed; G3: 0.50 mL synbiotics sprayed; G4: 0.25 mL of synbiotics are added to drinking water; G5: 0.50 mL of synbiotics are added to drinking water; G6: 0.25 mL of synbiotics dripped into the mouth; and G7: 0.50 mL of synbiotics dripped into mouth drops. Lactic acid bacteria(LAB) were significantly increased (P<0.0001) compared to the control group and other treated groups and had the maximum values after the use of synbiotics via drinking water (0.25 or 0.50 mL). Furthermore, when comparing the treated birds (G4, G5) with the control birds, the Escherichia coli concentration in the drinking water containing synbiotics was significantly lower. In addition, treated chickens at (G7) showed a higher duodenum, ileum villus height (VH), and VH. - Ileum crypt depth (CD) ratio compared to other groups. In addition, birds treated with 0.50 mL of synbiotics in drinking water (G5) performed better in duodenum, ileum, CD and VH. - CD ratio than the other groups. Meanwhile, intestinal tract length and visceral pH did not differ significantly between groups. It can be concluded that the use of 0.25 mL of synbiotics in drinking water can improve the overall health of birds.

Recent developments of (bio)-sensors for detection of main microbiological and non-biological pollutants in plastic bottled water samples: A critical review.

The importance of water in all biological processes is undeniable. Ensuring access to clean and safe drinking water is crucial for maintaining sustainable water resources. To elaborate, the consumption of water of inadequate quality can have a repercussion on human health. Furthermore, according to the instability of tap water quality, the consumption rate of bottled water is increasing every day at the global level. Although most people believe bottled water is safe, it can also be contaminated by microbiological or chemical pollution, which can increase the risk of disease. Over the last decades, several conventional analytical tools applied to analyze the contamination of bottled water. On the other hand, some limitations restrict their application in this field. Therefore, biosensors, as emerging analytical method, attract tremendous attention for detection both microbial and chemical contamination of bottled water. Biosensors enjoy several facilities including selectivity, affordability, and sensitivity. In this review, the developed biosensors for analyzing contamination of bottled water were highlighted, as along with working strategies, pros and cons of studies. Challenges and prospects were also examined.

Bacterial communities of planktonic bacteria and mature biofilm in service lines and premise plumbing of a Megacity: Composition, Diversity, and influencing factors.

Although simulated studies have provided valuable knowledge regarding the communities of planktonic bacteria and biofilms, the lack of systematic field studies have hampered the understanding of microbiology in real-world service lines and premise plumbing. In this study, the bacterial communities of water and biofilm were explored, with a special focus on the lifetime development of biofilm communities and their key influencing factors. The 16S rRNA gene sequencing results showed that both the planktonic bacteria and biofilm were dominated by Proteobacteria. Among the 15,084 observed amplicon sequence variants (ASVs), the 33 core ASVs covered 72.8 %, while the 12 shared core ASVs accounted for 62.2 % of the total sequences. Remarkably, it was found that the species richness and diversity of biofilm communities correlated with pipe age. The relative abundance of ASV2 (f_Sphingomonadaceae) was lower for pipe ages 40-50 years (7.9 %) than for pipe ages 10-20 years (59.3 %), while the relative abundance of ASV10 (f_Hyphomonadaceae) was higher for pipe ages 40-50 years (19.5 %) than its presence at pipe ages 20-30 years (1.9 %). The community of the premise plumbing biofilm had significantly higher species richness and diversity than that of the service line, while the steel-plastics composite pipe interior lined with polyethylene (S-PE) harbored significantly more diverse biofilm than the galvanized steel pipes (S-Zn). Interestingly, S-PE was enriched with ASV27 (g_Mycobacterium), while S-Zn pipes were enriched with ASV13 (g_Pseudomonas). Moreover, the network analysis showed that five rare ASVs, not core ASVs, were keystone members in biofilm communities, indicating the importance of rare members in the function and stability of biofilm communities. This manuscript provides novel insights into real-world service lines and premise plumbing microbiology, regarding lifetime dynamics (pipe age 10-50 years), and the influences of pipe types (premise plumbing vs. service line) and pipe materials (S-Zn vs. S-PE).

Impact of harmful algal bloom severity on bacterial communities in a full-scale biological filtration system for drinking water treatment.

The occurrence of harmful algal blooms (HABs) in freshwater environments has been expanded worldwide with growing frequency and severity. HABs can pose a threat to public water supplies, raising concerns about safety of treated water. Many studies have provided valuable information about the impacts of HABs and management strategies on the early-stage treatment processes (e.g., pre-oxidation and coagulation/flocculation) in conventional drinking water treatment plants (DWTPs). However, the potential effect of HAB-impacted water in the granular media filtration has not been well studied. Biologically-active filters (BAFs), which are used in drinking water treatment and rely largely on bacterial community interactions, have not been examined during HABs in full-scale DWTPs. In this study, we assessed the bacterial community structure of BAFs, functional profiles, assembly processes, and bio-interactions in the community during both severe and mild HABs. Our findings indicate that bacterial diversity in BAFs significantly decreases during severe HABs due to the predominance of bloom-associated bacteria (e.g., Spingopyxis, Porphyrobacter, and Sphingomonas). The excitation-emission matrix combined with parallel factor analysis (EEM-PARAFAC) confirmed that filter influent affected by the severe HAB contained a higher portion of protein-like substances than filter influent samples during a mild bloom. In addition, BAF community functions showed increases in metabolisms associated with intracellular algal organic matter (AOM), such as lipids and amino acids, during severe HABs. Further ecological process and network analyses revealed that severe HAB, accompanied by the abundance of bloom-associated taxa and increased nutrient availability, led to not only strong stochastic processes in the assembly process, but also a bacterial community with lower complexity in BAFs. Overall, this study provides deeper insights into BAF bacterial community structure, function, and assembly in response to HABs.

Domestic hot-water boilers harbour active thermophilic bacterial communities distinctly different from those in the cold-water supply.

Running cold and hot water in buildings is a widely established commodity. However, interests regarding hygiene and microbiological aspects had so far been focussed on cold water. Little attention has been given to the microbiology of domestic hot-water installations (DHWIs), except for aspects of pathogenic Legionella. World-wide, regulations consider hot (or warm) water as 'heated drinking water' that must comply (cold) drinking water (DW) standards. However, the few reports that exist indicate presence and growth of microbial flora in DHWIs, even when supplied with water with disinfectant residual. Using flow cytometric (FCM) total cell counting (TCC), FCM-fingerprinting, and 16S rRNA-gene-based metagenomic analysis, the characteristics and composition of bacterial communities in cold drinking water (DW) and hot water from associated boilers (operating at 50 - 60 °C) was studied in 14 selected inhouse DW installations located in Switzerland and Austria. A sampling strategy was applied that ensured access to the bulk water phase of both, supplied cold DW and produced hot boiler water. Generally, 1.3- to 8-fold enhanced TCCs were recorded in hot water compared to those in the supplied cold DW. FCM-fingerprints of cold and corresponding hot water from individual buildings indicated different composition of cold- and hot-water microbial floras. Also, hot waters from each of the boilers sampled had its own individual FCM-fingerprint. 16S rRNA-gene-based metagenomic analysis confirmed the marked differences in composition of microbiomes. E.g., in three neighbouring houses supplied from the same public network pipe each hot-water boiler contained its own thermophilic bacterial flora. Generally, bacterial diversity in cold DW was broad, that in hot water was restricted, with mostly thermophilic strains from the families Hydrogenophilaceae, Nitrosomonadaceae and Thermaceae dominating. Batch growth assays, consisting of cold DW heated up to 50 - 60 °C and inoculated with hot water, resulted in immediate cell growth with doubling times between 5 and 10 h. When cold DW was used as an inoculum no significant growth was observed. Even boilers supplied with UVC-treated cold DW contained an actively growing microbial flora, suggesting such hot-water systems as autonomously operating, thermophilic bioreactors. The generation of assimilable organic carbon from dissolved organic carbon due to heating appears to be the driver for growth of thermophilic microbial communities. Our report suggests that a man-made microbial ecosystem, very close to us all and of potential hygienic importance, may have been overlooked so far. Despite consumers having been exposed to microbial hot-water flora for a long time, with no major pathogens so far been associated specifically with hot-water usage (except for Legionella), the role of harmless thermophiles and their interaction with potential human pathogens able to grow at elevated temperatures in DHWIs remains to be investigated.

A Community Waterborne Salmonella Bovismorbificans Outbreak in Greece.

BACKGROUND: In August 2022, the Hellenic National Public Health Organisation was notified about a gastroenteritis outbreak in town A in Southern Greece. Investigations aimed to identify the source and implement control measures. METHODS: Case definition categories were used in a 1:3 case-control study. Cases and controls were interviewed about various exposures. Cases' stool samples were cultured on agar plates and characterised by serotyping, antimicrobial susceptibility testing and Pulse Field Gel Electrophoresis (PFGE). Environmental investigations included tap water sampling for microbiological and chemical analysis in town A and inspection of the water supply system. RESULTS: We identified 33 cases (median age: 17 years). Tap water consumption was the only significant risk factor for gastroenteritis (OR = 5.46, 95% CI = 1.02-53.95). Salmonella (S.) Bovismorbificans isolated from eight stool and one tap water samples had identical PFGE profiles. No resistant isolates were identified. Residual chlorine levels were lower than the acceptable limits before and during the outbreak. We advised consumption of bottled water and adherence to strict hand hygiene rules until tap water was declared suitable for drinking. CONCLUSIONS: Epidemiological and molecular data revealed a waterborne S. Bovismorbificans outbreak in town A. We recommend local water safety authorities to ensure that residual chlorine levels comply with the legislation towards water safety planning, to mitigate risks.

On-site filtration of large sample volumes improves the detection of opportunistic pathogens in drinking water distribution systems.

In this study, we compared conventional vacuum filtration of small volumes through disc membranes (effective sample volumes for potable water: 0.3-1.0 L) with filtration of high volumes using ultrafiltration (UF) modules (effective sample volumes for potable water: 10.6-84.5 L) for collecting bacterial biomass from raw, finished, and tap water at seven drinking water systems. Total bacteria, Legionella spp., Legionella pneumophila, Mycobacterium spp., and Mycobacterium avium complex in these samples were enumerated using both conventional quantitative PCR (qPCR) and viability qPCR (using propidium monoazide). In addition, PCR-amplified gene fragments were sequenced for microbial community analysis. The frequency of detection (FOD) of Legionella spp. in finished and tap water samples was much greater using UF modules (83% and 77%, respectively) than disc filters (24% and 33%, respectively). The FODs for Mycobacterium spp. in raw, finished, and tap water samples were also consistently greater using UF modules than disc filters. Furthermore, the number of observed operational taxonomic units and diversity index values for finished and tap water samples were often substantially greater when using UF modules as compared to disc filters. Conventional and viability qPCR yielded similar results, suggesting that membrane-compromised cells represented a minor fraction of total bacterial biomass. In conclusion, our research demonstrates that large-volume filtration using UF modules improved the detection of opportunistic pathogens at the low concentrations typically found in public drinking water systems and that the majority of bacteria in these systems appear to be viable in spite of disinfection with free chlorine and/or chloramine.IMPORTANCEOpportunistic pathogens, such as Legionella pneumophila, are a growing public health concern. In this study, we compared sample collection and enumeration methods on raw, finished, and tap water at seven water systems throughout the State of Minnesota, USA. The results showed that on-site filtration of large water volumes (i.e., 500-1,000 L) using ultrafiltration membrane modules improved the frequency of detection of relatively rare organisms, including opportunistic pathogens, compared to the common approach of filtering about 1 L using disc membranes. Furthermore, results from viability quantitative PCR (qPCR) with propidium monoazide were similar to conventional qPCR, suggesting that membrane-compromised cells represent an insignificant fraction of microorganisms. Results from these ultrafiltration membrane modules should lead to a better understanding of the microbial ecology of drinking water distribution systems and their potential to inoculate premise plumbing systems with opportunistic pathogens where conditions are more favorable for their growth.

Exposure to chlorinated drinking water alters the murine fecal microbiota.

An abundant body of scientific studies and regulatory guidelines substantiates antimicrobial efficacy of freshwater chlorination ensuring drinking water safety in large populations worldwide. In contrast to the purposeful use of chlorination ensuring antimicrobial safety of drinking water, only a limited body of research has addressed the molecular impact of chlorinated drinking water exposure on the gut microbiota. Here, for the first time, we have examined the differential effects of drinking water regimens stratified by chlorination agent [inorganic (HOCl) versus chloramine (TCIC)] on the C57BL/6J murine fecal microbiota. To this end, we exposed C57BL/6J mice to chlorinated drinking water regimens followed by fecal bacterial microbiota analysis at the end of the three-week feeding period employing 16S rRNA sequencing. α-diversity was strongly reduced when comparing chlorinated versus control drinking water groups and community dissimilarities (ß-diversity) were significant between groups even when comparing HOCl and TCIC. We detected significant differences in fecal bacterial composition as a function of drinking water chlorination observable at the phylum and genus levels. Differential abundance analysis of select amplicon sequence variants (ASVs) revealed changes as a function of chlorination exposure [up: Lactobacillus ASV1; Akkermansia muciniphila ASV7; Clostridium ss1 ASV10; down: Ileibacterium valens ASV5; Desulfovibrio ASV11; Lachnospiraceae UCG-006 ASV15]. Given the established complexity of murine and human gastrointestinal microbiota and their role in health and disease, the translational relevance of the chlorination-induced changes documented by us for the first time in the fecal murine microbiota remains to be explored.

Suboptimal Bacteriological Quality of Household Water in Municipal Ibadan, Nigeria.

Access to potable water is difficult for many African residents. This study evaluated the bacteriological quality of household water collected in the dry and wet seasons across five municipal local government areas (LGAs) in Ibadan, a large city in southwest Nigeria. A total of 447 water samples (dry season, n = 250; wet season, n = 197) were aseptically collected from a random sample of mapped households within Ibadan's five municipal LGAs. The pH values and total aerobic and coliform bacterial counts were measured, and samples were screened for Escherichia coli, Salmonella, Shigella, and Yersinia by standard phenotypic techniques and multiplex polymerase chain reaction. The most common source of water was well (53.2%), followed by borehole (34%). None of the households used municipal tap water. Cumulatively, aerobic (P = 0.0002) and coliform (P = 0.0001) counts as well as pH values (P = 0.0002) changed significantly between seasons, with increasing and decreasing counts depending on the LGA. Nonpotable water samples were found to be very common during the dry (86.8%) and wet (74.1%) seasons. Escherichia coli spp., as indicators of recent fecal contamination, were isolated from 115 (25.7%) of the household water sources. Thirty three Salmonella, four enteroaggregative E. coli, and four enterotoxigenic E. coli isolates but no Shigella or Yersinia isolates were identified. This study revealed the absence of treated tap water and the poor quality of alternative sources with detectable pathogens in municipal Ibadan. Addressing the city-wide lack of access to potable water is an essential priority for preventing a high prevalence of feco-orally transmitted infections.

Effect of booster disinfection on the prevalence of microbial antibiotic resistance and bacterial community in a simulated drinking water distribution system.

Booster disinfection was often applied to control the microorganism's growth in long-distance water supply systems. The effect of booster disinfection on the changing patterns of antibiotic resistance and bacterial community was investigated by a simulated water distribution system (SWDS). The results showed that the antibiotic resistance bacteria (ARB) and antibiotic resistance genes (ARGs) were initially removed after dosing disinfectants (chlorine and chloramine), but then increased with the increasing water age. However, the relative abundance of ARGs significantly increased after booster disinfection both in buck water and biofilm, then decreased along the pipeline. The pipe materials and disinfectant type also affected the antibiotic resistance. Chlorine was more efficient in controlling microbes and ARGs than chloramine. Compared with UPVC and PE pipes, SS pipes had the lowest total bacteria, ARB concentration, and ARB percentage, mainly due to higher disinfectant residuals and a smoother surface. The significant correlation (rs = 0.77, p < 0.001) of the 16S rRNA genes was observed between buck water and biofilm, while the correlations of targeted ARGs were found to be weak. Bray-Curtis similarity index indicated that booster disinfection significantly changed the biofilm bacterial community and the disinfectant type also had a marked impact on the bacterial community. At the genus level, the relative abundance of Pseudomonas, Sphingomonas, and Methylobacterium significantly increased after booster disinfection. Mycobacterium increased after chloramination while decreased after chlorination, indicating Mycobacterium might resist chloramine. Pseudomonas, Methylobacterium, and Phreatobacter were found to correlate well with the relative abundance of ARGs. These results highlighted antibiotic resistance shift and bacterial community alteration after booster disinfection, which may be helpful in controlling potential microbial risk in drinking water.

Microbial community of municipal drinking water in Hangzhou using metagenomic sequencing.

While traditional culture-dependent methods can effectively detect certain microorganisms, the comprehensive composition of the municipal drinking water (DW) microbiome, including bacteria, archaea, and viruses, remains unknown. Metagenomic sequencing has opened the door to accurately determine and analyze the entire microbial community of DW, providing a comprehensive understanding of DW species diversity, especially in the context of public health concerns during the COVID-19 era. In this study, we found that most of the culturable bacteria and some fecal indicator bacteria, such as Escherichia coli and Pseudomonas aeruginosa, were non-culturable using culture-dependent methods in all samples. However, metagenomic analysis showed that the predominant bacterial species in the DW samples belonged to the phyla Proteobacteria and Planctomycetes. Notably, the genus Methylobacterium was the most abundant in all water samples, followed by Sphingomonas, Gemmata, and Azospirilum. While low levels of virulence-associated factors, such as the Esx-5 type VII secretion system (T7SS) and DevR/S, were detected, only the erythromycin resistance gene erm(X), an rRNA methyltransferase, was identified at low abundance in one sample. Hosts corresponding to virulence and resistance genes were identified in some samples, including Mycobacterium spp. Archaeal DNA (Euryarchaeota, Crenarchaeota) was found in trace amounts in some DW samples. Viruses such as rotavirus, coxsackievirus, human enterovirus, and SARS-CoV-2 were negative in all DW samples using colloidal gold and real-time reverse transcription polymerase chain reaction (RT‒PCR) methods. However, DNA encoding a new order of reverse-transcribing viruses (Ortervirales) and Herpesvirales was found in some DW samples. The metabolic pathways of the entire microbial community involve cell‒cell communication and signal secretion, contributing to cooperation between different microbial populations in the water. This study provides insight into the microbial community and metabolic process of DW in Hangzhou, China, utilizing both culture-dependent methods and metagenomic sequencing combined with bioinformatics tools during the COVID-19 pandemic era.

Free-living lifestyle preferences drive the antibiotic resistance promotion during drinking water chlorination.

The risk associated with antibiotic resistance genes (ARGs) in size-fractionated bacterial community during drinking water chlorination remains unclear, and is of paramount importance for risk mitigation through process selection and optimization. This study employed metagenomic approaches to reveal the alterations of ARGs, their potential functions and hosts within the free-living and particle-associated fractions. The total relative abundance of ARGs, mobile genetic elements (MGEs), and virulence factor genes (VFGs) significantly increased in the free-living fraction after chlorination. The contribution of the free-living fraction to the ARG relative abundance rose from 16.40 ± 1.31 % to 93.62 ± 0.47 % after chlorination. Multidrug resistance genes (e.g. mexF and mexW) were major contributors, and their co-occurrence with MGEs in the free-living fraction was enhanced after chlorination. Considering multiple perspectives, including presence, mobility, and pathogenicity, chlorination led to a significant risk of the antibiotic resistome in the free-living fraction. Moreover, potential functions of ARGs, such as cell wall/membrane/envelope biogenesis, defense mechanisms, and transcription in the free-living fraction, were intensified following chlorination. Potential pathogens, including Pseudomonas aeruginosa, Pseudomonas alcaligenes, and Acinetobacter junii, were identified as the predominant hosts of multidrug resistance genes, with their increased abundances primarily contributing to the rise of the corresponding ARGs. Overall, alterations of hosts as well as enhancing mobility and biological functions could collectively aid the proliferation and spread of ARGs in the free-living fraction after chlorination. This study provides novel insights into antibiotic resistance evolution in size-fractionated bacteria community and offers a management strategy for microbiological safety in drinking water.

Incidence of co-resistance to antibiotics and chlorine in bacterial biofilm of hospital water systems: Insights into the risk of nosocomial infections.

The presence of biofilms in drinking water distribution systems (DWDS) in healthcare settings poses a considerable risk to the biological security of water, particularly when the biofilm bacteria demonstrate antimicrobial resistance characteristics. This study aimed to investigate the occurrence of antibiotic-resistant bacteria (ARB) in biofilms within DWDS of hospitals. The chlorine resistance of the isolated ARB was analyzed, and then chlorine-resistant bacteria (CRB) were identified using molecular methods. Additionally, the presence of several antibiotic resistance genes (ARGs) was monitored in the isolated ARB. Out of the 41 biofilm samples collected from hospitals, ARB were detected in 32 (78%) of the samples. A total of 109 colonies of ARB were isolated from DWDS of hospitals, with ß-lactam resistant bacteria, including ceftazidime-resistant and ampicillin-resistant bacteria, being the most frequently isolated ARB. Analyzing of ARGs revealed the highest detection of aac6, followed by sul1 gene. However, the ß-lactamase genes blaCTX-M and blaTEM were not identified in the ARB, suggesting the presence of other ß-lactamase genes not included in the tested panel. Exposure of ARB to free chlorine at a concentration of 0.5 mg/l showed that 64% of the isolates were CRB. However, increasing the chlorine concentration to 4 mg/l decreased the high fraction of ARB (91%). The domi||nant CRB identified were Sphingomonas, Brevundimonas, Stenotrophomonas, Bacillus and Staphylococcus with Bacillus exhibiting the highest frequency. The results highlight the potential risk of biofilm formation in the DWDS of hospitals, leading to the dissemination of ARB in hospital environments, which is a great concern for the health of hospitalized patients, especially vulnerable individuals. Surveillance of antimicrobial resistance in DWDS of hospitals can provide valuable insights for shaping antimicrobial use policies and practices that ensure their efficacy.

Rhodamine B functionalized silver nanoparticles paper discs as turn-on fluorescence sensor, coupled with a smartphone for the detection of microbial contamination in drinking water.

The precise detection of pathogenic microorganisms is crucial for the reduction of water-borne diseases. Herein, a filter-paper-based florescent chemosensor was fabricated for the detection of Escherichia coli and Staphylococcus aureus contamination exploiting protein-DNA interaction between the target and a specific probe. The sensing mechanism involved the self-assembly of Rhodamine B (RhB) on silver nanoparticles (AgNPs) surface that was labeled with a single-stranded DNA probe. This causes the fluorescence quenching of RhB by a distant-dependant process. The hybridization between pathogen-specific probe and bacterial surface protein causes the release of fluorescence of RhB, which was observed under UV light. For paper-based bio-surface preparation, the mixture comprising RhB-AgNP-ssDNA was drop-casted on filter paper discs. The conditions were optimized using isolated genomic DNA of the microbes. The method was applied for E.coli detection using an eae gene-based probe targeting intimin protein and S. aureus detection using tuf gene-based probe targeting EF-tuf protein on the microbe's surface. The chemosensor had a notable specificity and selectivity for E.coli, and S. aureus, with detection limits of 0.6 × 108 and 0.37 × 103 CFU/mL respectively. Moreover, the sensor was tested on real water samples, which presented excellent reproducibility of results (RSD ≤ 0.24%). Furthermore, the gradient change of fluorescence was captured by a smartphone, which allows direct detection of pathogens in a sensitive semi-quantitative way without the need for expensive instruments. The designed chemosensor can serve as a simple, inexpensive, and rapid method for the on-site detection of microbial contamination in drinking water.