RESUMO
BACKGROUND: Human milk is unquestionably beneficial for preterm infants. We investigated how the transition from tube to oral/breastfeeding impacts the preterm infants' oral and gut microbiome and metabolome. METHODS: We analyzed stool, saliva, and milk samples collected from a cohort of preterm infants enrolled in the MAP Study, a prospective observational trial. The microbiome and metabolome of the samples were analyzed from 4 longitudinal sample time points, 2 during tube feeds only and 2 after the initiation of oral/breastfeeding. RESULTS: We enrolled 11 mother-infant dyads (gestational age = 27.9 (23.4-32.2)) and analyzed a total of 39 stool, 44 saliva, and 43 milk samples over 4 timepoints. In saliva samples, there was a shift towards increased Streptococcus and decreased Staphylococcus after oral feeding/breastfeeding initiation (p < 0.05). Milk sample metabolites were strongly influenced by the route of feeding and milk type (p < 0.05) and represented the pathways of Vitamin E metabolism, Vitamin B12 metabolism, and Tryptophan metabolism. CONCLUSION: Our analysis demonstrated that the milk and preterm infant's saliva microbiome and metabolome changed over the course of the first four to 5 months of life, coinciding with the initiation of oral/breastfeeds. IMPACT: The microbiome and metabolome is altered in the infant's saliva but not their stool, and in mother's milk when feeds are transitioned from tube to oral/breastfeeding. We assessed the relationship between the gut and oral microbiome/metabolome with the milk microbiome/metabolome over a longitudinal period of time in preterm babies. Metabolites that changed in the infants saliva after the initiation of oral feeds have the potential to be used as biomarkers for disease risk.
RESUMO
AIMS: Gastrointestinal disease is a leading cause of morbidity in bottlenose dolphins (Tursiops truncatus) under managed care. Fecal microbiota transplantation (FMT) holds promise as a therapeutic tool to restore gut microbiota without antibiotic use. This prospective clinical study aimed to develop a screening protocol for FMT donors to ensure safety, determine an effective FMT administration protocol for managed dolphins, and evaluate the efficacy of FMTs in four recipient dolphins. METHODS AND RESULTS: Comprehensive health monitoring was performed on donor and recipient dolphins. Fecal samples were collected before, during, and after FMT therapy. Screening of donor and recipient fecal samples was accomplished by in-house and reference lab diagnostic tests. Shotgun metagenomics was used for sequencing. Following FMT treatment, all four recipient communities experienced engraftment of novel microbial species from donor communities. Engraftment coincided with resolution of clinical signs and a sustained increase in alpha diversity. CONCLUSION: The donor screening protocol proved to be safe in this study and no adverse effects were observed in four recipient dolphins. Treatment coincided with improvement in clinical signs.
Assuntos
Golfinho Nariz-de-Garrafa , Microbioma Gastrointestinal , Animais , Transplante de Microbiota Fecal/métodos , Estudos Prospectivos , Fezes , Resultado do TratamentoRESUMO
Reduced availability of agricultural water has spurred increased interest in using recycled irrigation water for U.S. food crop production. However, there are significant knowledge gaps concerning the microbiological quality of these water sources. To address these gaps, we used 16S rRNA gene and metagenomic sequencing to characterize taxonomic and functional variations (e.g., antimicrobial resistance) in bacterial communities across diverse recycled and surface water irrigation sources. We collected 1 L water samples (n = 410) between 2016 and 2018 from the Mid-Atlantic (12 sites) and Southwest (10 sites) U.S. Samples were filtered, and DNA was extracted. The V3-V4 regions of the 16S rRNA gene were then PCR amplified and sequenced. Metagenomic sequencing was also performed to characterize antibiotic, metal, and biocide resistance genes. Bacterial alpha and beta diversities were significantly different (p < 0.001) across water types and seasons. Pathogenic bacteria, such as Salmonella enterica, Staphylococcus aureus, and Aeromonas hydrophilia were observed across sample types. The most common antibiotic resistance genes identified coded against macrolides/lincosamides/streptogramins, aminoglycosides, rifampin and elfamycins, and their read counts fluctuated across seasons. We also observed multi-metal and multi-biocide resistance across all water types. To our knowledge, this is the most comprehensive longitudinal study to date of U.S. recycled water and surface water used for irrigation. Our findings improve understanding of the potential differences in the risk of exposure to bacterial pathogens and antibiotic resistance genes originating from diverse irrigation water sources across seasons and U.S. regions.
Assuntos
Antibacterianos , Desinfetantes , Estados Unidos , RNA Ribossômico 16S/genética , Antibacterianos/farmacologia , Estudos Longitudinais , Bactérias/genética , Resistência Microbiana a Medicamentos/genética , Água , Irrigação Agrícola , Águas Residuárias , Genes BacterianosRESUMO
The U.S. Food Safety Modernization Act (FSMA) Produce Safety Rule (PSR) requires that farmers generate a Microbial Water Quality Profile (MWQP) from 20 samples per agricultural water source, taken over 2-4 years and five annual samples thereafter. Farmers must use the MWQP to ascertain a geometric mean (GM) of ≤126 CFU/100 mL and statistical threshold value (STV) of ≤410 CFU/100 mL of generic Escherichia coli. Farmers are responsible for collecting samples and paying for testing, incurring a financial and time burden. To determine if testing frequency can be reduced without compromising accuracy, water samples (n = 279) were collected from twelve sites in the U.S. Mid-Atlantic region from 2016 to 2018 comprising tidal brackish river, non-tidal fresh river, pond, vegetable processing, and reclaimed water. The GM and STV were calculated for all sites and water types using all samples, and for multiple sub-samples of <20 from each site and water type. A Monte Carlo simulation was used to determine the proportion of sub-sample sizes that yielded the same determination as the entire sample size of PSR standard compliance. Four sites, two pond and two reclaimed water sites, complied with PSR GM and STV requirements when using the entire sample set. When a water source's calculated GM and STV using the entire sample set hovered close to the PSR thresholds, sub-sample sizes approached the recommended 20 samples to reach a congruent compliance determination. However, 99% agreement was obtained with a sub-sample of five when the absolute difference between the GM and STV from total samples and the PSR thresholds was ≥2.6 and 4.5 log CFU/100 mL E. coli, respectively. These findings suggest that under certain conditions the MWQP may be generated with well below 20 samples, reducing the economic burden on farmers while still maintaining a representative MWQP.
Assuntos
Irrigação Agrícola , Qualidade da Água , Escherichia coli , Inocuidade dos Alimentos , Microbiologia da ÁguaRESUMO
As climate change continues to stress freshwater resources, we have a pressing need to identify alternative (nontraditional) sources of microbially safe water for irrigation of fresh produce. This study is part of the center CONSERVE, which aims to facilitate the adoption of adequate agricultural water sources. A 26-month longitudinal study was conducted at 11 sites to assess the prevalence of bacteria indicating water quality, fecal contamination, and crop contamination risk (Escherichia coli, total coliforms [TC], Enterococcus, and Aeromonas). Sites included nontidal freshwater rivers/creeks (NF), a tidal brackish river (TB), irrigation ponds (PW), and reclaimed water sites (RW). Water samples were filtered for bacterial quantification. E. coli, TC, enterococci (â¼86%, 98%, and 90% positive, respectively; n = 333), and Aeromonas (â¼98% positive; n = 133) were widespread in water samples tested. Highest E. coli counts were in rivers, TC counts in TB, and enterococci in rivers and ponds (P < 0.001 in all cases) compared to other water types. Aeromonas counts were consistent across sites. Seasonal dynamics were detected in NF and PW samples only. E. coli counts were higher in the vegetable crop-growing (May-October) than nongrowing (November-April) season in all water types (P < 0.05). Only one RW and both PW sites met the U.S. Food Safety Modernization Act water standards. However, implementation of recommended mitigation measures of allowing time for microbial die-off between irrigation and harvest would bring all other sites into compliance within 2 days. This study provides comprehensive microbial data on alternative irrigation water and serves as an important resource for food safety planning and policy setting.IMPORTANCE Increasing demands for fresh fruit and vegetables, a variable climate affecting agricultural water availability, and microbial food safety goals are pressing the need to identify new, safe, alternative sources of irrigation water. Our study generated microbial data collected over a 2-year period from potential sources of irrigation (rivers, ponds, and reclaimed water sites). Pond water was found to comply with Food Safety Modernization Act (FSMA) microbial standards for irrigation of fruit and vegetables. Bacterial counts in reclaimed water, a resource that is not universally allowed on fresh produce in the United States, generally met microbial standards or needed minimal mitigation. We detected the most seasonality and the highest microbial loads in river water, which emerged as the water type that would require the most mitigation to be compliant with established FSMA standards. This data set represents one of the most comprehensive, longitudinal analyses of alternative irrigation water sources in the United States.
Assuntos
Aeromonas/isolamento & purificação , Irrigação Agrícola , Enterococcus/isolamento & purificação , Escherichia coli/isolamento & purificação , Lagoas/microbiologia , Rios/microbiologia , Irrigação Agrícola/métodos , Delaware , Estudos Longitudinais , Maryland , Microbiologia da ÁguaRESUMO
A quenching agent is commonly added to chlorinated, reclaimed water during sample collection to prevent chlorine-mediated die-off of viable microbiota. However, the effect of quenching on downstream 16S rRNA-based bacterial community analyses is unclear. We conducted a side-by-side comparison of 16S rRNA sequencing data from reclaimed water samples quenched with sodium thiosulfate and non-quenched samples. Our data showed that 16â¯S rRNA processing and sequencing methods, and resulting bacterial profiles, were not negatively impacted by quenching.
Assuntos
Microbiota , Tiossulfatos , Microbiologia da Água , Biodiversidade , Sequenciamento de Nucleotídeos em Larga Escala , Microbiota/efeitos dos fármacos , Microbiota/genética , Mid-Atlantic Region , RNA Ribossômico 16S/genética , Tiossulfatos/química , Tiossulfatos/farmacologia , Água/químicaRESUMO
Agricultural water withdrawals account for the largest proportion of global freshwater use. Increasing municipal water demands and droughts are straining agricultural water supplies. Therefore, alternative solutions to agricultural water crises are urgently needed, including the use of nontraditional water sources such as advanced treated wastewater or reclaimed water, brackish water, return flows, and effluent from produce processing facilities. However, it is critical to ensure that such usage does not compromise soil, crop, and public health. Here, we characterized five different nontraditional water types (nâ¯=â¯357 samples) for the presence of pharmaceuticals, herbicides, and disinfectants using ultra-high-pressure liquid chromatography tandem mass spectrometry based method (UPLC-MS/MS). We then evaluated whether the levels of these contaminants were influenced by season. The highest level of herbicides (atrazine) was detected in untreated pond water (median concentration 135.9â¯ng/L). Reclaimed water had the highest levels of antibiotics and stimulants including azithromycin (215â¯ng/L), sulfamethoxazole (232.1â¯ng/L), and caffeine (89.4â¯ng/L). Produce processing plant water also tended to have high levels of atrazine (102.7â¯ng/L) and ciprofloxacin (80.1â¯ng/L). In addition, we observed seasonal variability across water types, with the highest atrazine concentrations observed during summer months, while the highest median azithromycin concentrations were observed in reclaimed water during the winter season. Further studies are needed to evaluate if economically feasible on-farm water treatment technologies can effectively remove such contaminants from nontraditional irrigation water sources.
Assuntos
Desinfetantes/análise , Herbicidas/análise , Preparações Farmacêuticas , Poluentes Químicos da Água/análise , Cromatografia Líquida , Espectrometria de Massas em Tandem , Águas Residuárias , ÁguaRESUMO
The microbiome of the built environment comprises bacterial, archaeal, fungal, and viral communities associated with human-made structures. Even though most of these microbes are benign, antibiotic-resistant pathogens can colonize and emerge indoors, creating infection risk through surface transmission or inhalation. Several studies have catalogued the microbial composition and ecology in different built environment types. These have informed in vitro studies that seek to replicate the physicochemical features that promote pathogenic survival and transmission, ultimately facilitating the development and validation of intervention techniques used to reduce pathogen accumulation. Such interventions include using Bacillus-based cleaning products on surfaces or integrating bacilli into printable materials. Though this work is in its infancy, early research suggests the potential to use microbial biocontrol to reduce hospital- and home-acquired multidrug-resistant infections. Although these techniques hold promise, there is an urgent need to better understand the microbial ecology of built environments and to determine how these biocontrol solutions alter species interactions. This review covers our current understanding of microbial ecology of the built environment and proposes strategies to translate that knowledge into effective biocontrol of antibiotic-resistant pathogens.
Assuntos
Bacillus , Microbiota , Humanos , Bactérias/genética , Antibacterianos , Ambiente ConstruídoRESUMO
BACKGROUND: Microorganisms such as coliform-forming bacteria are commonly used to assess freshwater quality for drinking and recreational use. However, such organisms do not exist in isolation; they exist within the context of dynamic, interactive microbial communities which vary through space and time. Elucidating spatiotemporal microbial dynamics is imperative for discriminating robust community changes from ephemeral ecological trends, and for improving our overall understanding of the relationship between microbial communities and ecosystem health. We conducted a seven-year (2013-2019) microbial time-series investigation in the Chicago Area Waterways (CAWS): an urban river system which, in 2016, experienced substantial upgrades to disinfection processes at two wastewater reclamation plants (WRPs) that discharge into the CAWS and improved stormwater capture, to improve river water quality and reduce flooding. Using culture-independent and culture-dependent approaches, we compared CAWS microbial ecology before and after the intervention. RESULTS: Examinations of time-resolved beta distances between WRP-adjacent sites showed that community similarity measures were often consistent with the spatial orientation of site locations to one another and to the WRP outfalls. Fecal coliform results suggested that upgrades reduced coliform-associated bacteria in the effluent and the downstream river community. However, examinations of whole community changes through time suggest that the upgrades did little to affect overall riverine community dynamics, which instead were overwhelmingly driven by yearly patterns consistent with seasonality. CONCLUSIONS: This study presents a systematic effort to combine 16S rRNA gene amplicon sequencing with traditional culture-based methods to evaluate the influence of treatment innovations and systems upgrades on the microbiome of the Chicago Area Waterway System, representing the longest and most comprehensive characterization of the microbiome of an urban waterway yet attempted. We found that the systems upgrades were successful in improving specific water quality measures immediately downstream of wastewater outflows. Additionally, we found that the implementation of the water quality improvement measures to the river system did not disrupt the overall dynamics of the downstream microbial community, which remained heavily influenced by seasonal trends. Such results emphasize the dynamic nature of microbiomes in open environmental systems such as the CAWS, but also suggest that the seasonal oscillations remain consistent even when perturbed.
RESUMO
Extreme weather events induced by climate change have potential to impact water quality and have received increasing attention from surface water source management perspectives. However, it remains unclear how such phenomenon may influence concentration of emerging contaminants (ECs) in surface water that are vital source of irrigation. In the present study, we investigated the impact of high precipitation and ambient temperature on the distribution of ECs in surface water samples (N = 250) from Mid-Atlantic region, collected between 2016 and 2018. We analyzed the water samples using a liquid chromatography tandem mass spectrometry (LC-MS/MS) based method. We then investigated how the detection frequencies and concentrations of ten emerging contaminants were influenced by high precipitation and temperature events in the previous day or 7 days prior to the sampling events using a generalized additive model (GAM). We observed that heavy rainfalls occurring within 24 h before sampling increased the concentration/likelihood of detection of the ECs in surface waters, likely due to surface runoffs, remobilization from soil/sediment and sewage overflows. The impact of high precipitation during previous seven days varied across chemicals. Likewise, the detection frequency and concentration of most analytes increased with increasing temperature, in previous day of sampling event, likely due to enhanced solubility in water. Long-term high temperature events appeared to decrease the detection of the most tested ECs probably due to enhanced degradation. However, the potential risk of unknown degradation products cannot be ignored. Our results indicate potential decline of water quality after extreme weather events which may have implications for water source management under changing climate.
RESUMO
BACKGROUND: Determining the role of fomites in the transmission of SARS-CoV-2 is essential in the hospital setting and will likely be important outside of medical facilities as governments around the world make plans to ease COVID-19 public health restrictions and attempt to safely reopen economies. Expanding COVID-19 testing to include environmental surfaces would ideally be performed with inexpensive swabs that could be transported safely without concern of being a source of new infections. However, CDC-approved clinical-grade sampling supplies and techniques using a synthetic swab are expensive, potentially expose laboratory workers to viable virus and prohibit analysis of the microbiome due to the presence of antibiotics in viral transport media (VTM). To this end, we performed a series of experiments comparing the diagnostic yield using five consumer-grade swabs (including plastic and wood shafts and various head materials including cotton, synthetic, and foam) and one clinical-grade swab for inhibition to RNA. For three of these swabs, we evaluated performance to detect SARS-CoV-2 in twenty intensive care unit (ICU) hospital rooms of patients including COVID-19+ patients. All swabs were placed in 95% ethanol and further evaluated in terms of RNase activity. SARS-CoV-2 was measured both directly from the swab and from the swab eluent. RESULTS: Compared to samples collected in VTM, 95% ethanol demonstrated significant inhibition properties against RNases. When extracting directly from the swab head as opposed to the eluent, RNA recovery was approximately 2-4× higher from all six swab types tested as compared to the clinical standard of testing the eluent from a CDC-approved synthetic (SYN) swab. The limit of detection (LoD) of SARS-CoV-2 from floor samples collected using the consumer-grade plastic (CGp) or research-grade plastic The Microsetta Initiative (TMI) swabs was similar or better than the SYN swab, further suggesting that swab type does not impact RNA recovery as measured by the abundance of SARS-CoV-2. The LoD for TMI was between 0 and 362.5 viral particles, while SYN and CGp were both between 725 and 1450 particles. Lastly microbiome analyses (16S rRNA gene sequencing) of paired samples (nasal and floor from same patient room) collected using different swab types in triplicate indicated that microbial communities were not impacted by swab type, but instead driven by the patient and sample type. CONCLUSIONS: Compared to using a clinical-grade synthetic swab, detection of SARS-CoV-2 from environmental samples collected from ICU rooms of patients with COVID was similar using consumer-grade swabs, stored in 95% ethanol. The yield was best from the swab head rather than the eluent and the low level of RNase activity and lack of antibiotics in these samples makes it possible to perform concomitant microbiome analyses. Video abstract.
Assuntos
Teste de Ácido Nucleico para COVID-19/instrumentação , Teste de Ácido Nucleico para COVID-19/métodos , Microbiota , RNA Viral/análise , SARS-CoV-2/isolamento & purificação , Manejo de Espécimes/métodos , Transporte Biológico , Etanol/química , Estudos de Viabilidade , Humanos , Unidades de Terapia Intensiva , Limite de Detecção , RNA Ribossômico 16S/genética , RNA Viral/genética , Ribonucleases/metabolismoRESUMO
BACKGROUND: SARS-CoV-2 is an RNA virus responsible for the coronavirus disease 2019 (COVID-19) pandemic. Viruses exist in complex microbial environments, and recent studies have revealed both synergistic and antagonistic effects of specific bacterial taxa on viral prevalence and infectivity. We set out to test whether specific bacterial communities predict SARS-CoV-2 occurrence in a hospital setting. METHODS: We collected 972 samples from hospitalized patients with COVID-19, their health care providers, and hospital surfaces before, during, and after admission. We screened for SARS-CoV-2 using RT-qPCR, characterized microbial communities using 16S rRNA gene amplicon sequencing, and used these bacterial profiles to classify SARS-CoV-2 RNA detection with a random forest model. RESULTS: Sixteen percent of surfaces from COVID-19 patient rooms had detectable SARS-CoV-2 RNA, although infectivity was not assessed. The highest prevalence was in floor samples next to patient beds (39%) and directly outside their rooms (29%). Although bed rail samples more closely resembled the patient microbiome compared to floor samples, SARS-CoV-2 RNA was detected less often in bed rail samples (11%). SARS-CoV-2 positive samples had higher bacterial phylogenetic diversity in both human and surface samples and higher biomass in floor samples. 16S microbial community profiles enabled high classifier accuracy for SARS-CoV-2 status in not only nares, but also forehead, stool, and floor samples. Across these distinct microbial profiles, a single amplicon sequence variant from the genus Rothia strongly predicted SARS-CoV-2 presence across sample types, with greater prevalence in positive surface and human samples, even when compared to samples from patients in other intensive care units prior to the COVID-19 pandemic. CONCLUSIONS: These results contextualize the vast diversity of microbial niches where SARS-CoV-2 RNA is detected and identify specific bacterial taxa that associate with the viral RNA prevalence both in the host and hospital environment. Video Abstract.
Assuntos
COVID-19 , SARS-CoV-2 , Hospitais , Humanos , Pandemias , Filogenia , RNA Ribossômico 16S/genética , RNA Viral/genéticaRESUMO
Understanding weather-related drivers of crop plant-microbiome relationships is important for food security and food safety in the face of a changing climate. Cucumber and tomato are commercially important commodities that are susceptible to plant disease and have been implicated in foodborne disease outbreaks. To investigate the influence of precipitation on plant-associated microbiomes, epiphytically associated bacterial communities of cucumber and tomato samples were profiled by 16 S rRNA gene sequencing (V1-V3) in the days surrounding two rain events over a 17-day period. Following rain, α (within-sample) diversity measured on cucumber and tomato fruit surfaces, but not tomato leaf surfaces, increased significantly and remained elevated for several days. Bacterial ß (between-sample) diversity on cucumber and tomato fruit responded to precipitation. In the cucumber fruit surface (carpoplane), notable shifts in the families Xanthomonadaceae, Oxalobacteriaceae, Sphingobacteriaceae and Comamonadaceae were detected following precipitation. In the tomato carpoplane, shifts were detected in the families Enterobacteriaceae and Xanthomonadaceae following the first rain event, and in the Pseudomonadaceae and Oxalobacteriaceae following the second rain event. Few taxonomic shifts were detected in the tomato leaf surface (phylloplane). Exploring rain-induced shifts in plant microbiomes is highly relevant to crop protection, food safety and agroecology, and can aid in devising ways to enhance crop resilience to stresses and climate fluctuations.
Assuntos
Bactérias/patogenicidade , Cucumis sativus/microbiologia , Frutas/microbiologia , Microbiota/fisiologia , Chuva/microbiologia , Solanum lycopersicum/microbiologia , Bactérias/genética , Inocuidade dos Alimentos , Doenças Transmitidas por Alimentos/microbiologia , Microbiota/genética , Doenças das Plantas/microbiologia , RNA Ribossômico 16S/genéticaRESUMO
Background: Determining the role of fomites in the transmission of SARS-CoV-2 is essential in the hospital setting and will likely be important outside of medical facilities as governments around the world make plans to ease COVID-19 public health restrictions and attempt to safely reopen economies. Expanding COVID-19 testing to include environmental surfaces would ideally be performed with inexpensive swabs that could be transported safely without concern of being a source of new infections. However, CDC-approved clinical-grade sampling supplies and techniques using a synthetic swab are expensive, potentially expose laboratory workers to viable virus and prohibit analysis of the microbiome due to the presence of antibiotics in viral transport media (VTM). To this end, we performed a series of experiments comparing the diagnostic yield using five consumer-grade swabs (including plastic and wood shafts and various head materials including cotton, synthetic, and foam) and one clinical grade swab for inhibition to RNA. For three of these swabs, we evaluated performance to detect SARS-CoV-2 in twenty intensive care unit (ICU) hospital rooms of patients including COVID-19+ patients. All swabs were placed in 95% ethanol and further evaluated in terms of RNase activity. SARS-CoV-2 was measured both directly from the swab and from the swab eluent. Results: Compared to samples collected in VTM, 95% ethanol demonstrated significant inhibition properties against RNases. When extracting directly from the swab head as opposed to the eluent, RNA recovery was approximately 2-4x higher from all six swab types tested as compared to the clinical standard of testing the eluent from a CDC-approved synthetic (SYN) swab. The limit of detection (LoD) of SARSSARS-CoV-2 from floor samples collected using the consumer-grade plastic (CGp) or research-grade plastic The Microsetta Initiative (TMI) swabs was similar or better than the SYN swab, further suggesting that swab type does not impact RNA recovery as measured by the abundance of SARSSARS-CoV-2. The LoD for TMI was between 0-362.5 viral particles while SYN and CGp were both between 725-1450 particles. Lastly microbiome analyses (16S rRNA gene sequencing) of paired samples (nasal and floor from same patient-room) collected using different swab types in triplicate indicated that microbial communities were not impacted by swab type, but instead driven by the patient and sample type. Conclusions: Compared to using a clinical-grade synthetic swab, detection of SARS-CoV-2 from environmental samples collected from ICU rooms of patients with COVID was similar using consumer grade swabs, stored in 95% ethanol. The yield was best from the swab head rather than the eluent and the low level of RNase activity and lack of antibiotics in these samples makes it possible to perform concomitant microbiome analyses.
RESUMO
Background Determining the role of fomites in the transmission of SARS-CoV-2 is essential in the hospital setting and will likely be important outside of medical facilities as governments around the world make plans to ease COVID-19 public health restrictions and attempt to safely reopen economies. Expanding COVID-19 testing to include environmental surfaces would ideally be performed with inexpensive swabs that could be transported safely without concern of being a source of new infections. However, CDC-approved clinical-grade sampling supplies and techniques using a synthetic swab are expensive, potentially expose laboratory workers to viable virus and prohibit analysis of the microbiome due to the presence of antibiotics in viral transport media (VTM). To this end, we performed a series of experiments comparing the diagnostic yield using five consumer-grade swabs (including plastic and wood shafts and various head materials including cotton, synthetic, and foam) and one clinical grade swab for inhibition to RNA. For three of these swabs, we evaluated performance to detect SARS-CoV-2 in twenty intensive care unit (ICU) hospital rooms of patients with 16 COVID-19+. All swabs were placed in 95% ethanol and further evaluated in terms of RNase activity. SARS-CoV-2 was measured both directly from the swab and from the swab eluent. Results Compared to samples collected in VTM, 95% ethanol demonstrated significant inhibition properties against RNases. When extracting directly from the swab head as opposed to the eluent, RNA recovery was approximately 2-4x higher from all six swab types tested as compared to the clinical standard of testing the eluent from a CDC-approved synthetic swab. The limit of detection (LoD) of SARs-CoV-2 from floor samples collected using the CGp or TMI swabs was similar or better than the CDC standard, further suggesting that swab type does not impact RNA recovery as measured by SARs-CoV-2. The LoD for TMI was between 0-362.5 viral particles while SYN and CGp were both between 725-1450 particles. Lastly microbiome analyses (16S rRNA) of paired samples (e.g., environment to host) collected using different swab types in triplicate indicated that microbial communities were not impacted by swab type but instead driven by the patient and sample type (floor or nasal). Conclusions Compared to using a clinical-grade synthetic swab, detection of SARS-CoV-2 from environmental samples collected from ICU rooms of patients with COVID was similar using consumer grade swabs, stored in 95% ethanol. The yield was best from the swab head rather than the eluent and the low level of RNase activity in these samples makes it possible to perform concomitant microbiome analysis. Keywords: COVID-19, SARS-CoV-2, RT-qPCR, swab, global health.
RESUMO
Mangrove ecosystems provide important ecological benefits and ecosystem services, including carbon storage and coastline stabilization, but they also suffer great anthropogenic pressures. Microorganisms associated with mangrove sediments and the rhizosphere play key roles in this ecosystem and make essential contributions to its productivity and carbon budget. Understanding this nexus and moving from descriptive studies of microbial taxonomy to hypothesis-driven field and lab studies will facilitate a mechanistic understanding of mangrove ecosystem interaction webs and open opportunities for microorganism-mediated approaches to mangrove protection and rehabilitation. Such an effort calls for a multidisciplinary and collaborative approach, involving chemists, ecologists, evolutionary biologists, microbiologists, oceanographers, plant scientists, conservation biologists, and stakeholders, and it requires standardized methods to support reproducible experiments. Here, we outline the Mangrove Microbiome Initiative, which is focused around three urgent priorities and three approaches for advancing mangrove microbiome research.
RESUMO
Irrigation water contaminated with Salmonella enterica and Listeria monocytogenes may provide a route of contamination of raw or minimally processed fruits and vegetables. While previous work has surveyed specific and singular types of agricultural irrigation water for bacterial pathogens, few studies have simultaneously surveyed different water sources repeatedly over an extended period of time. This study quantified S. enterica and L. monocytogenes levels (MPN/L) at 6 sites, including river waters: tidal freshwater river (MA04, n = 34), non-tidal freshwater river, (MA05, n = 32), one reclaimed water holding pond (MA06, n = 25), two pond water sites (MA10, n = 35; MA11, n = 34), and one produce wash water site (MA12, n = 10) from September 2016-October 2018. Overall, 50% (84/168) and 31% (53/170) of sampling events recovered S. enterica and L. monocytogenes, respectively. Results showed that river waters supported significantly (p < 0.05) greater levels of S. enterica than pond or reclaimed waters. The non-tidal river water sites (MA05) with the lowest water temperature supported significantly greater level of L. monocytogenes compared to all other sites; L. monocytogenes levels were also lower in winter and spring compared to summer seasons. Filtering 10 L of water through a modified Moore swab (MMS) was 43.5 (Odds ratio, p < 0.001) and 25.5 (p < 0.001) times more likely to recover S. enterica than filtering 1 L and 0.1 L, respectively; filtering 10 L was 4.8 (p < 0.05) and 3.9 (p < 0.05) times more likely to recover L. monocytogenes than 1L and 0.1 L, respectively. Work presented here shows that S. enterica and L. monocytogenes levels are higher in river waters compared to pond or reclaimed waters in the Mid-Atlantic region of the U.S., and quantitatively shows that analyzing 10 L water is more likely recover pathogens than smaller samples of environmental waters.
Assuntos
Irrigação Agrícola/métodos , Água Doce/microbiologia , Listeria monocytogenes/isolamento & purificação , Salmonella enterica/isolamento & purificação , Estações do Ano , Microbiologia da Água , Mid-Atlantic Region , Prevalência , Estados UnidosRESUMO
Synergistic effects of bacteria on viral stability and transmission are widely documented but remain unclear in the context of SARS-CoV-2. We collected 972 samples from hospitalized ICU patients with coronavirus disease 2019 (COVID-19), their health care providers, and hospital surfaces before, during, and after admission. We screened for SARS-CoV-2 using RT-qPCR, characterized microbial communities using 16S rRNA gene amplicon sequencing, and contextualized the massive microbial diversity in this dataset in a meta-analysis of over 20,000 samples. Sixteen percent of surfaces from COVID-19 patient rooms were positive, with the highest prevalence in floor samples next to patient beds (39%) and directly outside their rooms (29%). Although bed rail samples increasingly resembled the patient microbiome throughout their stay, SARS-CoV-2 was less frequently detected there (11%). Despite surface contamination in almost all patient rooms, no health care workers providing COVID-19 patient care contracted the disease. SARS-CoV-2 positive samples had higher bacterial phylogenetic diversity across human and surface samples, and higher biomass in floor samples. 16S microbial community profiles allowed for high classifier accuracy for SARS-CoV-2 status in not only nares, but also forehead, stool and floor samples. Across these distinct microbial profiles, a single amplicon sequence variant from the genus Rothia was highly predictive of SARS-CoV-2 across sample types, and had higher prevalence in positive surface and human samples, even when comparing to samples from patients in another intensive care unit prior to the COVID-19 pandemic. These results suggest that bacterial communities contribute to viral prevalence both in the host and hospital environment.
RESUMO
The impact of microbially contaminated irrigation water on risks to produce safety and public health is a complex issue that is not well understood. This study tracked fecal indicators, pathogenic bacteria, and total bacterial communities from a creek water irrigation source to irrigated produce to assess the impact of irrigation events on soil and produce-associated microbiota. Kale and radishes were drip-irrigated using Mid-Atlantic creek water in October 2017. Plant and soil samples were collected immediately before and after irrigation, and for 3 consecutive days thereafter. All samples (nâ¯=â¯134), including irrigation water, were tested for generic Escherichia coli and total coliforms (TC) using standard membrane filtration or direct plating, and for Salmonella enterica and Listeria monocytogenes by selective enrichment. DNA extracted from all samples was PCR-amplified for the V3-V4 region of the 16S rRNA gene for bacterial community profiling. In soil, TC levels were significantly higher immediately and 3â¯days post-irrigation compared to pre-irrigation (pâ¯<â¯0.01). E. coli levels in soil increased after irrigation, but the difference was not significant (pâ¯=â¯0.31), and die-off was not observed. No E. coli were detected on kale leaves. TC increased over the study period on radish roots (pâ¯<â¯0.01) but not kale leaves (pâ¯=â¯0.43). Although target pathogens were detected in irrigation water, S. enterica was detected from only one post-irrigation kale sample and L. monocytogenes was not detected in the field. The 16S rRNA gene sequencing data revealed differences in bacterial community structure and composition across sample types and showed that radish soil and root surface bacterial communities were more strongly influenced by irrigation compared to kale samples. This study provides insights into the impact of irrigation water on fresh produce microbiota, revealing that, although irrigation did influence crop-associated microbiota (especially below ground) in the field, bacterial pathogens were not likely transferred to the crop.
Assuntos
Bactérias/isolamento & purificação , Brassica/microbiologia , Fezes/microbiologia , Microbiologia de Alimentos , Raphanus/microbiologia , Microbiologia do Solo , Microbiologia da Água , Irrigação Agrícola , Brassica/crescimento & desenvolvimento , Produtos Agrícolas , Escherichia coli/isolamento & purificação , Maryland , Microbiota , Raphanus/crescimento & desenvolvimentoRESUMO
The Mid-Atlantic Microbiome Meet-up (M3) organization brings together academic, government, and industry groups to share ideas and develop best practices for microbiome research. In January of 2018, M3 held its fourth meeting, which focused on recent advances in biodefense, specifically those relating to infectious disease, and the use of metagenomic methods for pathogen detection. Presentations highlighted the utility of next-generation sequencing technologies for identifying and tracking microbial community members across space and time. However, they also stressed the current limitations of genomic approaches for biodefense, including insufficient sensitivity to detect low-abundance pathogens and the inability to quantify viable organisms. Participants discussed ways in which the community can improve software usability and shared new computational tools for metagenomic processing, assembly, annotation, and visualization. Looking to the future, they identified the need for better bioinformatics toolkits for longitudinal analyses, improved sample processing approaches for characterizing viruses and fungi, and more consistent maintenance of database resources. Finally, they addressed the necessity of improving data standards to incentivize data sharing. Here, we summarize the presentations and discussions from the meeting, identifying the areas where microbiome analyses have improved our ability to detect and manage biological threats and infectious disease, as well as gaps of knowledge in the field that require future funding and focus.