RESUMO
Wastewater-based monitoring for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) at the individual building level could be an efficient, passive means of early detection of new cases in congregate living settings, but this approach has not been validated. Preliminary samples were collected from a hospital and a local municipal wastewater treatment plant. Molecular diagnostic methods were compared side by side to assess feasibility, performance, and sensitivity. Refined sample collection and processing protocols were then used to monitor two occupied dormitory complexes (n = 105 and 66) over 8 weeks. Wastewater results were validated using known case counts from external clinical testing of building occupants. Results confirm that ultracentrifugation from a 24-h composite collection had a sensitivity of 96.2% and a specificity of 100%. However, the method could not distinguish new infectious cases from persistent convalescent shedding of SARS-CoV-2 RNA. If the detection of convalescent shedding is considered a false positive, then the sensitivity is 100% and specificity drops to 45%. It was determined that the proposed approach constitutes a highly sensitive wastewater surveillance method for detecting SARS-CoV-2, but it could not distinguish new infectious cases from persistent convalescent shedding. Future work must focus on approaches to distinguish new infections from convalescent shedding to fully realize the potential of building wastewater as a surveillance tool for congregate living. IMPORTANCE Some of the most severe outbreaks of COVID-19 have taken place in places where persons live together, such as nursing homes. Wastewater testing from individual buildings could be used for frequent pooled surveillance of virus from all occupants, including those who are contagious, with or without symptoms. This work provides a sensitive practical method for detecting infected individuals, as validated in two building complexes housing occupants who underwent frequent clinical testing performed by external entities. Although this sensitive method could be deployed now for pooled surveillance as an early warning system to limit outbreaks, the study shows that the approach will require further refinement to differentiate contagious, newly infected individuals from persons who have persistent viral fragments shedding in their stool outside the contagious period.
Assuntos
COVID-19/epidemiologia , Instituições Residenciais , SARS-CoV-2/isolamento & purificação , Águas Residuárias/virologia , COVID-19/diagnóstico , Humanos , Técnicas de Diagnóstico Molecular , Reprodutibilidade dos Testes , SARS-CoV-2/genética , Vigilância Epidemiológica Baseada em Águas ResiduáriasRESUMO
Liquid wastes (LW) disposed in hospital handwashing sinks may affect colonization of sink P-traps by carbapenemase-producing Klebsiella pneumoniae (CPKP), causing CPKP dispersal into the patient care environment. This study aimed to determine the effect of LW on biofilm formation and CPKP colonization in a P-Trap model (PTM). PTMs containing polymicrobial biofilms grown in autoclaved municipal tap water (ATW) supplemented with 5% dextrose in water (D5W), nutritional shake (Shake), sugar-based soft drink (Soda), or ATW were inoculated with K. pneumoniae ST258 KPC+ (ST258) or K. pneumoniae CAV1016 (CAV1016) and sampled after 7, 14, and 21 d. Biofilm bio-volume, mean thickness, and heterotrophic plate counts were significantly reduced and roughness coefficient significantly increased by Soda compared with D5W, Shake, or ATW. CPKP were significantly reduced by Soda but significantly amplified by D5W (ST258; CAV1016, 7 d) and Shake (ST258) suggesting that reducing LW disposal in sinks may reduce CPKP dispersal into patient care environments.
Assuntos
Infecções por Klebsiella , Klebsiella pneumoniae , Antibacterianos/uso terapêutico , Proteínas de Bactérias , Biofilmes , Humanos , Nutrientes , beta-LactamasesRESUMO
Hospital wastewater is an increasingly recognized reservoir for resistant Gram-negative organisms. Factors involved in establishment and persistence of Klebsiella pneumoniae carbapenemase-producing organisms (KPCOs) in hospital wastewater plumbing are unclear. This study was conducted at a hospital with endemic KPCOs linked to wastewater reservoirs and robust patient perirectal screening for silent KPCO carriage. Over 5 months, both rooms occupied and rooms not occupied by KPCO-positive patients were sampled at three wastewater sites within each room (sink drain, sink P-trap, and toilet or hopper). Risk factors for KPCO positivity were assessed using logistic regression. Whole-genome sequencing (WGS) identified environmental seeding by KPCO-positive patients. A total of 219/475 (46%) room sampling events were KPCO positive in at least one wastewater site. KPCO-positive patient exposure was associated with increased risk of environmental positivity for the room and toilet/hopper. Previous positivity and intensive care unit room type were consistently associated with increased risk. Tube feeds were associated with increased risk for the drain, while exposure to patients with Clostridioides difficile was associated with decreased risk. Urinary catheter exposure was associated with increased risk of P-trap positivity. P-trap heaters reduced risk of P-trap and sink drain positivity. WGS identified genomically linked environmental seeding in 6 of 99 room occupations by 40 KPCO-positive patients. In conclusion, KPCO-positive patients seed the environment in at least 6% of opportunities; once positive for KPCOs, wastewater sites are at greater risk of being positive subsequently. Increased nutrient exposure, e.g., due to tube food disposal down sinks, may increase risk; frequent flushing may be protective.IMPORTANCEKlebsiella pneumoniae carbapenemase-producing organisms (KPCOs) are bacteria that are resistant to most antibiotics and thus are challenging to treat when they cause infections in patients. These organisms can be acquired by patients who are hospitalized for other reasons, complicating their hospital stay and even leading to death. Hospital wastewater sites, such as sink drains and toilets, have played a role in many reported outbreaks over the past decade. The significance of our research is in identifying risk factors for environmental positivity for KPCOs, which will facilitate further work to prevent transmission of these organisms to patients from the hospital environment.
Assuntos
Proteínas de Bactérias/análise , Infecções por Klebsiella/epidemiologia , Klebsiella pneumoniae/isolamento & purificação , Águas Residuárias/microbiologia , beta-Lactamases/análise , Hospitais , Humanos , Infecções por Klebsiella/microbiologia , Virginia/epidemiologia , Águas Residuárias/análiseRESUMO
Several emerging pathogens have arisen as a result of selection pressures exerted by modern health care. Klebsiella quasipneumoniae was recently defined as a new species, yet its prevalence, niche, and propensity to acquire antimicrobial resistance genes are not fully described. We have been tracking inter- and intraspecies transmission of the Klebsiella pneumoniae carbapenemase (KPC) gene, blaKPC, between bacteria isolated from a single institution. We applied a combination of Illumina and PacBio whole-genome sequencing to identify and compare K. quasipneumoniae from patients and the hospital environment over 10- and 5-year periods, respectively. There were 32 blaKPC-positive K. quasipneumoniae isolates, all of which were identified as K. pneumoniae in the clinical microbiology laboratory, from 8 patients and 11 sink drains, with evidence for seven separate blaKPC plasmid acquisitions. Analysis of a single subclade of K. quasipneumoniae subsp. quasipneumoniae (n = 23 isolates) from three patients and six rooms demonstrated seeding of a sink by a patient, subsequent persistence of the strain in the hospital environment, and then possible transmission to another patient. Longitudinal analysis of this strain demonstrated the acquisition of two unique blaKPC plasmids and then subsequent within-strain genetic rearrangement through transposition and homologous recombination. Our analysis highlights the apparent molecular propensity of K. quasipneumoniae to persist in the environment as well as acquire carbapenemase plasmids from other species and enabled an assessment of the genetic rearrangements which may facilitate horizontal transmission of carbapenemases.
Assuntos
Klebsiella/enzimologia , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Carbapenêmicos/farmacologia , Resistência a Múltiplos Medicamentos/genética , Hospitais , Humanos , Klebsiella/efeitos dos fármacos , Testes de Sensibilidade Microbiana , Plasmídeos/genética , beta-Lactamases/genética , beta-Lactamases/metabolismoRESUMO
With multidrug-resistant (MDR) Enterobacterales on the rise, a nontoxic antimicrobial agent with a unique mechanism of action such as fosfomycin seems attractive. However, establishing accurate fosfomycin susceptibility testing for non-Escherichia coli isolates in a clinical microbiology laboratory remains problematic. We evaluated fosfomycin susceptibility by multiple methods with 96 KPC-producing clinical isolates of multiple strains and species collected at a single center between 2008 and 2016. In addition, we assessed the presence of fosfomycin resistance genes from whole-genome sequencing (WGS) data using NCBI's AMRFinder and custom HMM search. Susceptibility testing was performed using a glucose-6-phosphate-supplemented fosfomycin Etest and Kirby-Bauer disk diffusion (DD) assays, and the results were compared to those obtained by agar dilution. Clinical Laboratory and Standards Institute (CLSI) breakpoints for E. coli were applied for interpretation. Overall, 63% (60/96) of isolates were susceptible by Etest, 70% (67/96) by DD, and 88% (84/96) by agar dilution. fosA was detected in 80% (70/88) of previously sequenced isolates, with species-specific associations and alleles, and fosA-positive isolates were associated with higher MIC distributions. Disk potentiation testing was performed using sodium phosphonoformate to inhibit fosA and showed significant increases in the zone diameter of DD testing for isolates that were fosA positive compared to those that were fosA negative. The addition of sodium phosphonoformate (PPF) corrected 10/14 (71%) major errors in categorical agreement with agar dilution. Our results indicate that fosA influences the inaccuracy of susceptibility testing by methods readily available in a clinical laboratory compared to agar dilution. Further research is needed to determine the impact of fosA on clinical outcomes.
Assuntos
Antibacterianos/farmacologia , Proteínas de Bactérias/genética , Farmacorresistência Bacteriana , Fosfomicina/farmacologia , Infecções por Klebsiella/microbiologia , Klebsiella pneumoniae/efeitos dos fármacos , Klebsiella pneumoniae/genética , beta-Lactamases/genética , Proteínas de Bactérias/biossíntese , Genoma Bacteriano , Humanos , Klebsiella pneumoniae/isolamento & purificação , Testes de Sensibilidade Microbiana , Sequenciamento Completo do Genoma , beta-Lactamases/biossínteseRESUMO
An alarming rise in hospital outbreaks implicating hand-washing sinks has led to widespread acknowledgment that sinks are a major reservoir of antibiotic-resistant pathogens in patient care areas. An earlier study using green fluorescent protein (GFP)-expressing Escherichia coli (GFP-E. coli) as a model organism demonstrated dispersal from drain biofilms in contaminated sinks. The present study further characterizes the dispersal of microorganisms from contaminated sinks. Replicate hand-washing sinks were inoculated with GFP-E. coli, and dispersion was measured using qualitative (settle plates) and quantitative (air sampling) methods. Dispersal caused by faucet water was captured with settle plates and air sampling methods when bacteria were present on the drain. In contrast, no dispersal was captured without or in between faucet events, amending an earlier theory that bacteria aerosolize from the P-trap and disperse. Numbers of dispersed GFP-E. coli cells diminished substantially within 30 minutes after faucet usage, suggesting that the organisms were associated with larger droplet-sized particles that are not suspended in the air for long periods.IMPORTANCE Among the possible environmental reservoirs in a patient care environment, sink drains are increasingly recognized as a potential reservoir to hospitalized patients of multidrug-resistant health care-associated pathogens. With increasing antimicrobial resistance limiting therapeutic options for patients, a better understanding of how pathogens disseminate from sink drains is urgently needed. Once this knowledge gap has decreased, interventions can be engineered to decrease or eliminate transmission from hospital sink drains to patients. The current study further defines the mechanisms of transmission for bacteria that colonize sink drains.
Assuntos
Microbiologia do Ar , Escherichia coli/fisiologia , Desinfecção das Mãos , Hospitais , Água/química , Aerossóis/análise , Infecção Hospitalar/microbiologia , Infecção Hospitalar/prevenção & controle , Contaminação de Equipamentos , Escherichia coli/isolamento & purificação , Proteínas de Fluorescência Verde/análise , HumanosRESUMO
Background: The increasing prevalence of nosocomial carbapenemase-producing Enterobacteriaceae is a concern. However, the role of the environment in multispecies outbreaks remains poorly understood. There is increasing recognition that hospital wastewater plumbing may play a role. Methods: Covers were installed on all hoppers (a "toilet-like" waste disposal system) in adult intensive care units (ICUs) of a university hospital; additionally in the surgical ICU, sink trap heating and vibration devices were also installed. Patient acquisitions of Klebsiella pneumoniae carbapenemase-producing organisms (KPCOs) for patients who were admitted to an intervention unit were compared for 18-month preintervention and intervention periods. Results: Sixty hopper covers and 23 sink trap devices were installed. Fifty-six new multispecies KPCO acquisitions occurred preintervention compared to 30 during the intervention. Decreases for all KPCO acquisitions (odds ratio [OR], 0.51; 95% confidence interval [CI], 0.31-0.81; P = .003) and KPCO-positive clinical cultures (OR, 0.29; 95% CI, 0.17-0.48; P < .001) per admission in patients exposed to an intervention unit were observed. The incidence rate ratio was 0.51-fold (95% CI, 0.43-0.61) lower for all KPCO acquisitions during the intervention. The effect of the sink trap devices alone could not be determined, although the proportion of sink drain cultures positive for KPCO decreased (12/15 [80%] sites sampled preintervention vs 40/840 [5%] sampled during the intervention; P = .001). Conclusions: An intervention targeting wastewater plumbing fixtures, by installation of hopper covers, demonstrated a decrease in patient KPCO acquisitions. Considering wastewater reservoirs in nosocomial transmission of multispecies carbapenemase-producing Enterobacteriaceae may be critical.
Assuntos
Controle de Infecções/métodos , Unidades de Terapia Intensiva , Infecções por Klebsiella/prevenção & controle , Klebsiella pneumoniae/isolamento & purificação , Águas Residuárias/microbiologia , Proteínas de Bactérias/metabolismo , Aparelho Sanitário/microbiologia , Enterobacteriáceas Resistentes a Carbapenêmicos/isolamento & purificação , Portador Sadio/microbiologia , Infecção Hospitalar , Surtos de Doenças/prevenção & controle , Hospitais Universitários , Humanos , Controle de Infecções/instrumentação , Klebsiella pneumoniae/enzimologia , Estudos Prospectivos , Engenharia Sanitária/métodos , beta-Lactamases/metabolismoRESUMO
Over the last 20 years there have been 32 reports of carbapenem-resistant organisms in the hospital water environment, with half of these occurring since 2010. The majority of these reports have described associated clinical outbreaks in the intensive care setting, affecting the critically ill and the immunocompromised. Drains, sinks, and faucets were most frequently colonized, and Pseudomonas aeruginosa the predominant organism. Imipenemase (IMP), Klebsiella pneumoniae carbapenemase (KPC), and Verona integron-encoded metallo-ß-lactamase (VIM) were the most common carbapenemases found. Molecular typing was performed in almost all studies, with pulse field gel electrophoresis being most commonly used. Seventy-two percent of studies reported controlling outbreaks, of which just more than one-third eliminated the organism from the water environment. A combination of interventions seems to be most successful, including reinforcement of general infection control measures, alongside chemical disinfection. The most appropriate disinfection method remains unclear, however, and it is likely that replacement of colonized water reservoirs may be required for long-term clearance.
Assuntos
Carbapenêmicos/farmacologia , Infecção Hospitalar/microbiologia , Reservatórios de Doenças/microbiologia , Farmacorresistência Bacteriana , Hospitais , Microbiologia da Água , Abastecimento de Água , Antibacterianos/uso terapêutico , Proteínas de Bactérias/biossíntese , Enterobacteriáceas Resistentes a Carbapenêmicos/efeitos dos fármacos , Enterobacteriáceas Resistentes a Carbapenêmicos/enzimologia , Enterobacteriáceas Resistentes a Carbapenêmicos/genética , Infecção Hospitalar/epidemiologia , Surtos de Doenças/prevenção & controle , Desinfecção , Eletroforese em Gel de Campo Pulsado , Equipamentos e Provisões Hospitalares/microbiologia , Humanos , Infecções por Klebsiella/tratamento farmacológico , Infecções por Klebsiella/epidemiologia , Infecções por Klebsiella/microbiologia , Klebsiella pneumoniae/efeitos dos fármacos , Klebsiella pneumoniae/enzimologia , Klebsiella pneumoniae/genética , Testes de Sensibilidade Microbiana , Tipagem Molecular , Infecções por Pseudomonas/tratamento farmacológico , Infecções por Pseudomonas/epidemiologia , Infecções por Pseudomonas/microbiologia , Pseudomonas aeruginosa/efeitos dos fármacos , Pseudomonas aeruginosa/enzimologia , Pseudomonas aeruginosa/genética , beta-Lactamases/biossínteseRESUMO
There have been an increasing number of reports implicating Gammaproteobacteria as often carrying genes of drug resistance from colonized sink traps to vulnerable hospitalized patients. However, the mechanism of transmission from the wastewater of the sink P-trap to patients remains poorly understood. Herein we report the use of a designated hand-washing sink lab gallery to model dispersion of green fluorescent protein (GFP)-expressing Escherichia coli from sink wastewater to the surrounding environment. We found no dispersion of GFP-expressing E. coli directly from the P-trap to the sink basin or surrounding countertop with coincident water flow from a faucet. However, when the GFP-expressing E. coli cells were allowed to mature in the P-trap under conditions similar to those in a hospital environment, a GFP-expressing E. coli-containing putative biofilm extended upward over 7 days to reach the strainer. This subsequently resulted in droplet dispersion to the surrounding areas (<30 in.) during faucet operation. We also demonstrated that P-trap colonization could occur by retrograde transmission along a common pipe. We postulate that the organisms mobilize up to the strainer from the P-trap, resulting in droplet dispersion rather than dispersion directly from the P-trap. This work helps to further define the mode of transmission of bacteria from a P-trap reservoir to a vulnerable hospitalized patient.IMPORTANCE Many recent reports demonstrate that sink drain pipes become colonized with highly consequential multidrug-resistant bacteria, which then results in hospital-acquired infections. However, the mechanism of dispersal of bacteria from the sink to patients has not been fully elucidated. Through establishment of a unique sink gallery, this work found that a staged mode of transmission involving biofilm growth from the lower pipe to the sink strainer and subsequent splatter to the bowl and surrounding area occurs rather than splatter directly from the water in the lower pipe. We have also demonstrated that bacterial transmission can occur via connections in wastewater plumbing to neighboring sinks. This work helps to more clearly define the mechanism and risk of transmission from a wastewater source to hospitalized patients in a world with increasingly antibiotic-resistant bacteria that can thrive in wastewater environments and cause infections in vulnerable patients.
Assuntos
Contaminação de Equipamentos , Infecções por Escherichia coli/transmissão , Escherichia coli/isolamento & purificação , Proteínas de Fluorescência Verde/análise , Desinfecção das Mãos , Águas Residuárias/microbiologia , Biofilmes , Infecção Hospitalar/microbiologia , Infecção Hospitalar/prevenção & controle , Reservatórios de Doenças/microbiologia , Farmacorresistência Bacteriana , Escherichia coli/química , Infecções por Escherichia coli/microbiologia , Hospitalização , Humanos , Pacientes InternadosRESUMO
A laboratory scale semi-batch fed anaerobic ammonia oxidation (ANAMMOX) reactor was operated in the lab under two different feeding operations. In the first scenario, termed as phase I, the reactor was seeded and operated with NO(2) -N added externally with the filtrate to the reactor in the ratio needed for the successful ANAMMOX. A second reactor was also initiated shortly after the start-up of the ANAMMOX to accomplish partial nitrification (nitritation reactor) to generate NO(2) -N. In phase II, the operation of the ANAMMOX reactor was switched to the mode in which case the partially nitrified effluent from the nitritation reactor was fed to the ANAMMOX reactor. In both phases, real filtrate from a local wastewater treatment plant was used as the feed. The ANAMMOX reactor sustained a loading rate (average 0.33 ± 0.03 with a max of 0.4 g N (L day)(-1) ) which is comparable with many other fed-batch reactors in the literature. Consistent total N removal (average of 82 ± 4%) could be sustained in the ANAMMOX reactor during both phases. The nitritation reactor also consistently enabled a NO(2) -N to NH(3) -N ratio of 1.2:1 which was needed for the successful operation of the ANAMMOX reactor in phase II. Sequence analysis and FISH showed that Kuenenia stuttgartiensis dominated the enriched ANAMMOX community along with several unidentified, but seemingly enriched, potential ANAMMOX strains. Microbial ecology analysis for nitritation reactor showed the dominance of Nitrosomonas europaea. In summary, this manuscript provides important information on the start-up and operation of anammox reactor with detailed investigation on microbial ecology in this reactor.
Assuntos
Bactérias Anaeróbias/metabolismo , Reatores Biológicos , Fungos/metabolismo , Anaerobiose , Bactérias Anaeróbias/genética , Sequência de Bases , Primers do DNA , DNA Bacteriano/isolamento & purificação , Filtração , Fungos/genética , Hibridização in Situ Fluorescente , Oxirredução , Filogenia , Reação em Cadeia da PolimeraseRESUMO
Antibiotic-resistant bacteria originating from hospitals are ultimately discharged to municipal wastewater treatment plants (WWTP), which may serve as important reservoirs for the spread of antibiotic resistant genes. This study traced and quantified the presence of a rare but clinically relevant antimicrobial resistance gene; Klebsiella pneumoniae carbapenamase (KPC)-and the viable organisms (KPCO) which carried this gene in hospital, non-hospital wastewater discharges, various compartments within a municipal WWTP, receiving water and sediment samples. High concentration of the gene, blaKPC harbored in viable and multispecies KPCO was detected in the hospital wastewater and in the forepart stages of the WWTP, but was not detected in the final effluent following UV disinfection. KPCO were not detected in multiple non-hospital sources of wastewater discharges tested. The treatment train used in the sampled WWTP was found to help remove and reduce KPCO load. Using whole-genome sequencing, a KPC-producing Klebsiella oxytoca strain identical to strains seen in the patients and hospital environment was isolated from the downstream receiving water on one sampling event. KPCO were also found to persist in the biosolids throughout the WWTP, but were not detected in the processed compost-products made from WWTP-biosolids. This study systematically demonstrates dissemination of KPCO from hospital point source to environment via municipal WWTP. Understanding hospitals as the origin and source of spread of some of the most clinically urgent antimicrobial-resistant organisms may help direct interventions that target rate at which antibiotic resistant bacteria evolve and spread via enhancement of wastewater treatment and mitigation of dissemination at source.
RESUMO
Congregate living poses one of the highest risk situations for the transmission of respiratory viruses including SARS-CoV-2. University dormitories exemplify such high-risk settings. We demonstrate the value of using building-level SARS-CoV-2 wastewater surveillance as an early warning system to inform when prevalence testing of all building occupants is warranted. Coordinated daily testing of composite wastewater samples and clinical testing in dormitories was used to prompt the screening of otherwise unrecognized infected occupants. We overlay the detection patterns in the context of regular scheduled occupant testing to validate a wastewater detection model. The trend of wastewater positivity largely aligned well with the clinical positivity and epidemiology of dormitory occupants. However, the predictive ability of wastewater-surveillance to detect new positive cases is hampered by convalescent shedding in recovered/noncontagious individuals as they return to the building. Building-level pooled wastewater-surveillance and forecasting is most productive for predicting new cases in low-prevalence instances at the community level. For higher-education facilities and other congregate living settings to remain in operation during a pandemic, a thorough surveillance-based decision-making system is vital. Building-level wastewater monitoring on a daily basis paired with regular testing of individual dormitory occupants is an effective and efficient approach for mitigating outbreaks on university campuses.
RESUMO
BACKGROUND: Sink drains in healthcare facilities may provide an environment for antimicrobial-resistant microorganisms, including carbapenemase-producing Klebsiella pneumoniae (CPKP). METHODS: We investigated the colonization of a biofilm consortia by CPKP in a model system simulating a sink-drain P-trap. Centers for Disease Control (CDC) biofilm reactors (CBRs) were inoculated with microbial consortia originally recovered from 2 P-traps collected from separate patient rooms (designated rooms A and B) in a hospital. Biofilms were grown on stainless steel (SS) or polyvinyl chloride (PVC) coupons in autoclaved municipal drinking water (ATW) for 7 or 28 days. RESULTS: Microbial communities in model systems (designated CBR-A or CBR-B) were less diverse than communities in respective P-traps A and B, and they were primarily composed of ß and γ Proteobacteria, as determined using 16S rRNA community analysis. Following biofilm development CBRs were inoculated with either K. pneumoniae ST45 (ie, strain CAV1016) or K. pneumoniae ST258 KPC+ (ie, strain 258), and samples were collected over 21 days. Under most conditions tested (CBR-A: SS, 7-day biofilm; CBR-A: PVC, 28-day biofilm; CBR-B: SS, 7-day and 28-day biofilm; CBR-B: PVC, 28-day biofilm) significantly higher numbers of CAV1016 were observed compared to 258. CAV1016 showed no significant difference in quantity or persistence based on biofilm age (7 days vs 28 days) or substratum type (SS vs PVC). However, counts of 258 were significantly higher on 28-day biofilms and on SS. CONCLUSIONS: These results suggest that CPKP persistence in P-trap biofilms may be strain specific or may be related to the type of P-trap material or age of the biofilm.
Assuntos
Enterobacteriáceas Resistentes a Carbapenêmicos , Klebsiella pneumoniae , Biofilmes , Carbapenêmicos/farmacologia , Humanos , Klebsiella pneumoniae/genética , RNA Ribossômico 16SRESUMO
Antimicrobial resistance has been recognized as a threat to human health. The role of hospital sinks acting as a reservoir for some of the most concerning antibiotic resistant organisms, carbapenemase producing Enterobacterales (CPE) is evident but not well understood. Strategies to prevent establishment, interventions to eliminate these reservoirs and factors which drive persistence of CPE are not well established. We use a uniquely designed sink lab to transplant CPE colonized hospital sink plumbing with an aim to understand CPE dynamics in a controlled setting, notably exploiting both molecular and culture techniques. After ex situ installation the CPE population in the sink plumbing drop from previously detectable to undetectable levels. The addition of nutrients is followed by a quick rebound in CPE detection in the sinks after as many as 37 days. We did not however detect a significant shift in microbial community structure or the overall resistance gene carriage in longitudinal samples from a subset of these transplanted sinks using whole shotgun metagenomic sequencing. Comparing nutrient types in a benchtop culture study model, protein rich nutrients appear to be the most supportive for CPE growth and biofilm formation ability. The role of nutrients exposure is determining factor for maintaining a high bioburden of CPE in the sink drains and P-traps. Therefore, limiting nutrient disposal into sinks has reasonable potential with regard to decreasing the CPE wastewater burden, especially in hospitals seeking to control an environmental reservoir.
Assuntos
Klebsiella pneumoniae , beta-Lactamases , Proteínas de Bactérias , Humanos , NutrientesRESUMO
OBJECTIVE: Describe the epidemiological and molecular characteristics of an outbreak of Klebsiella pneumoniae carbapenemase (KPC)-producing organisms and the novel use of a cohorting unit for its control. DESIGN: Observational study. SETTING: A 566-room academic teaching facility in Milwaukee, Wisconsin. PATIENTS: Solid-organ transplant recipients. METHODS: Infection control bundles were used throughout the time of observation. All KPC cases were intermittently housed in a cohorting unit with dedicated nurses and nursing aids. The rooms used in the cohorting unit had anterooms where clean supplies and linens were placed. Spread of KPC-producing organisms was determined using rectal surveillance cultures on admission and weekly thereafter among all consecutive patients admitted to the involved units. KPC-positive strains underwent pulsed-field gel electrophoresis and whole-genome sequencing. RESULTS: A total of 8 KPC cases (5 identified by surveillance) were identified from April 2016 to April 2017. After the index patient, 3 patients acquired KPC-producing organisms despite implementation of an infection control bundle. This prompted the use of a cohorting unit, which immediately halted transmission, and the single remaining KPC case was transferred out of the cohorting unit. However, additional KPC cases were identified within 2 months. Once the cohorting unit was reopened, no additional KPC cases occurred. The KPC-positive species identified during this outbreak included Klebsiella pneumoniae, Enterobacter cloacae complex, and Escherichia coli. blaKPC was identified on at least 2 plasmid backbones. CONCLUSIONS: A complex KPC outbreak involving both clonal and plasmid-mediated dissemination was controlled using weekly surveillances and a cohorting unit.
Assuntos
Infecção Hospitalar/prevenção & controle , Surtos de Doenças/prevenção & controle , Controle de Infecções/métodos , Infecções por Klebsiella/prevenção & controle , Idoso , Proteínas de Bactérias/genética , Infecção Hospitalar/epidemiologia , DNA Bacteriano/genética , Eletroforese em Gel de Campo Pulsado , Feminino , Humanos , Infecções por Klebsiella/epidemiologia , Klebsiella pneumoniae/enzimologia , Klebsiella pneumoniae/genética , Masculino , Pessoa de Meia-Idade , Epidemiologia Molecular , Pacotes de Assistência ao Paciente , Wisconsin/epidemiologia , beta-Lactamases/genéticaRESUMO
Bacillus coagulans strain IIT-BT S1 isolated from anaerobically digested activated sewage sludge was investigated for its ability to produce H(2) from glucose-based medium under the influence of different environmental parameters. At mid-exponential phase of cell growth, H(2) production initiated and reached maximum production rate in the stationary phase. The maximal H(2) yield (2.28 mol H(2)/molglucose) was recorded at an initial glucose concentration of 2% (w/v), pH 6.5, temperature 37 degrees C, inoculum volume of 10% (v/v) and inoculum age of 14 h. Cell growth rate and rate of hydrogen production decreased when glucose concentration was elevated above 2% w/v, indicating substrate inhibition. The ability of the organism to utilize various carbon sources for H(2) fermentation was also determined.
Assuntos
Anaerobiose , Bacillus/metabolismo , Hidrogênio/metabolismo , Esgotos/microbiologia , Bacillus/genética , Bacillus/crescimento & desenvolvimento , Bacillus/isolamento & purificação , DNA Ribossômico/genética , Glucose/metabolismo , Concentração de Íons de Hidrogênio , Filogenia , RNA Ribossômico 16S/genéticaRESUMO
Around 150 lipase producing bacterial isolates were screened from the local soils enriched with oil. Citrobacrer freundii IIT-BT L139, an isolated microbial strain, produced lipase that had high activity (8.8 U/ml) at pH 9.0 and 40 degrees C. The 16S rDNA phylogenetic studies showed that Citrobacter freundii belongs to the family Enterobacteriaceae and later confirmed by the microbial identification. Suitable C and N sources for lipase production were deduced to be starch and peptone-urea, respectively. In a controlled fermenter (1 L), the lipase activity was found to increase by 36% (12 Uml(-1)). The variation of lipase activity, pH and dissolved oxygen (DO) during growth of the organism in the controlled batch fermenter were monitored. The rheological characteristics of the fermentation broth indicated that it behaved like a Newtonian fluid throughout the fermentation. The fermentation time was comparatively short (60 h). The lipase was also found to be substantially resistant to common detergents. This lipase was, thus, characterized as alkaline, thermostable and solvent stable, which was essentially desirable in pharmaceutical, detergent and other industrial applications or production.
Assuntos
Citrobacter freundii/enzimologia , Lipase/biossíntese , DNA Ribossômico/química , Estabilidade Enzimática , Fermentação , Concentração de Íons de Hidrogênio , Lipase/química , Metabolismo dos Lipídeos , Oxigênio/metabolismo , Filogenia , Reologia , Solventes/química , Tensoativos/química , Temperatura , Fatores de TempoRESUMO
Solids reduction in activated sludge processes (ASP) at source using process manipulation has been researched widely over the last two-decades. However, the absence of nutrient removal component, lack of understanding on the organic carbon, and limited information on key microbial community in solids minimizing ASP preclude the widespread acceptance of sludge minimizing processes. In this manuscript, we report simultaneous solids reduction through anaerobiosis along with nitrogen and phosphorus removals. The manuscript also reports carbon mass balance using stable isotope of carbon, microbial ecology of nitrifiers and polyphosphate accumulating organisms (PAOs). Two laboratory scale reactors were operated in anaerobic-aerobic-anoxic (A(2)O) mode. One reactor was run in the standard mode (hereafter called the control-SBR) simulating conventional A(2)O type of activated sludge process and the second reactor was run in the sludge minimizing mode (called the modified-SBR). Unlike other research efforts where the sludge minimizing reactor was maintained at nearly infinite solids retention time (SRT). To sustain the efficient nutrient removal, the modified-SBR in this research was operated at a very small solids yield rather than at infinite SRT. Both reactors showed consistent NH3-N, phosphorus and COD removals over a period of 263 days. Both reactors also showed active denitrification during the anoxic phase even if there was no organic carbon source available during this phase, suggesting the presence of denitrifying PAOs (DNPAOs). The observed solids yield in the modified-SBR was 60% less than the observed solids yield in the control-SBR. Specific oxygen uptake rate (SOUR) for the modified-SBR was almost 44% more than the control-SBR under identical feeding conditions, but was nearly the same for both reactors under fasting conditions. The modified-SBR showed greater diversity of ammonia oxidizing bacteria and PAOs compared to the control-SBR. The diversity of PAOs in the modified-SBR was even more interesting in which case novel clades of Candidatus Accumulibacter phosphatis (CAP), an uncultured but widely found PAOs, were found.
Assuntos
Betaproteobacteria/metabolismo , Reatores Biológicos , Carbono/metabolismo , Oxigênio/metabolismo , Esgotos/química , Amônia/metabolismo , Betaproteobacteria/genética , DNA Bacteriano/genética , Dados de Sequência Molecular , Oxirredução , Filogenia , Reação em Cadeia da Polimerase , Polimorfismo de Fragmento de Restrição , Polifosfatos/metabolismo , RNA Ribossômico 16S/genética , Análise de Sequência de DNARESUMO
This research demonstrates the first ever application of lytic bacteriophage (virus) mediated biocontrol of biomass bulking in the activated sludge process using Haliscomenobacter hydrossis as a model filamentous bacterium. Bacteriophages are viruses that specifically infect bacteria only. The lytic phage specifically infecting H. hydrossis was isolated from the mixed liquor of a local wastewater treatment plant. The isolated bacteriophage belongs to the Myoviridae family with a contractile tail (length-126 nm; diameter-18 nm) and icosahedral head (diameter-81 nm). Titer of the isolated phage with H. hydrossis was calculated to be 5.2 ± 0.3 × 10(5) PFU/mL and burst size was found to be 105 ± 7 PFU/infected cell. The phage was considerably stable after exposure to high temperature (42 °C) and pH between 5 and 8, emphasizing that it can withstand the seasonal/operational fluctuations under real-time applications. Phage to host (bacteria) ratio for the optimal infection was found to be 1:1000 with â¼54% host death. The isolated phage showed no cross infectivity with other bacteria most commonly found in activated sludge systems, thus validating its suitability for biocontrol of filamentous bulking caused by H. hydrossis. Following the phage application, successful reduction in sludge volume index (SVI) from 155 to 105 was achieved, indicating improved biomass settling. The application of phage did not affect nutrient removal efficiency of the biomass, suggesting no collateral damage. Similar to phage therapy in medical applications, phage-mediated biocontrol holds a great potentiality for large-scale applications as economic agent in the mitigation of several water, wastewater and environmental problems. Present study in this direction is a novel effort.
Assuntos
Bacteroidetes/virologia , Biomassa , Myoviridae/genética , Controle Biológico de Vetores/métodos , Esgotos/microbiologia , Bacteriólise , Nefelometria e TurbidimetriaRESUMO
Two full-scale trickling filter/solids contact (TF/SC) basin plants, each successfully performing nitrification, were sampled throughout various seasons over a period of one year. Concentrations of ammonia, nitrate and nitrite were measured at various sampling locations along the treatment train. DNA was also extracted from mixed liquor in the solids contact basins. These DNA samples were subjected to terminal restriction fragment length polymorphism (TRFLP) in order to profile the ammonia oxidizing bacteria and nitrite oxidizing bacteria communities. In both plants, there was a prevalence of Nitrosomonas europaea among the ammonia oxidizing bacteria (AOBs). However, during the summer months, there was increased diversity of Nitrosomonas species. Likewise, Nitrospira spp. was the dominant nitrite oxidizing bacteria (NOBs) in both plants regardless of season. Yet there was an increased presence of Nitrobacter among the NOBs in the summer months. These results add an important understanding of the ecology and dynamics in nitrifying population in full-scale TF/SC wastewater treatment plants.