Last data update: May 20, 2024. (Total: 46824 publications since 2009)
Records 1-23 (of 23 Records) |
Query Trace: Mercante A [original query] |
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Detecting Mpox cases through wastewater surveillance - United States, August 2022-May 2023
Adams C , Kirby AE , Bias M , Riser A , Wong KK , Mercante JW , Reese H . MMWR Morb Mortal Wkly Rep 2024 73 (2) 37-43 In October 2022, CDC's National Wastewater Surveillance System began routine testing of U.S. wastewater for Monkeypox virus. Wastewater surveillance sensitivity, positive predictive value (PPV), and negative predictive value (NPV) for Monkeypox virus were evaluated by comparing wastewater detections (Monkeypox virus detected versus not detected) to numbers of persons with mpox in a county who were shedding virus. Case ascertainment was assumed to be complete, and persons with mpox were assumed to shed virus for 25 days after symptom onset. A total of 281 cases and 3,492 wastewater samples from 89 sites in 26 counties were included in the analysis. Wastewater surveillance in a single week, from samples representing thousands to millions of persons, had a sensitivity of 32% for detecting one or more persons shedding Monkeypox virus, 49% for detecting five or more persons shedding virus, and 77% for detecting 15 or more persons shedding virus. Weekly PPV and NPV for detecting persons shedding Monkeypox virus in a county were 62% and 80%, respectively. An absence of detections in counties with wastewater surveillance signified a high probability that a large number of cases were not present. Results can help to guide the public health response to Monkeypox virus wastewater detections. A single, isolated detection likely warrants a limited public health response. An absence of detections, in combination with no reported cases, can give public health officials greater confidence that no cases are present. Wastewater surveillance can serve as a useful complement to case surveillance for guiding the public health response to an mpox outbreak. |
Validation of novel Mycobacterium tuberculosis isoniazid resistance mutations not detectable by common molecular tests (preprint)
Kandler JL , Mercante AD , Dalton TL , Ezewudo MN , Cowan LS , Burns SP , Metchock B , Cegielski P , Posey JE . bioRxiv 2018 322750 Resistance to the first-line anti-tuberculosis (TB) drug, isoniazid (INH), is widespread, and the mechanism of resistance is unknown in approximately 15% of INH-resistant (INH-R) strains. To improve molecular detection of INH-R TB, we used whole genome sequencing (WGS) to analyze 52 phenotypically INH-R Mycobacterium tuberculosis complex (MTBC) clinical isolates that lacked the common katG S315T or inhA promoter mutations. Approximately 94% (49/52) of strains had mutations at known INH-associated loci that were likely to confer INH resistance. All such mutations would be detectable by sequencing more DNA adjacent to existing target regions. Use of WGS minimized the chances of missing infrequent INH resistance mutations outside commonly targeted hotspots. We used recombineering to generate 12 observed clinical katG mutations in the pansusceptible H37Rv reference strain and determined their impact on INH resistance. Our functional genetic experiments have confirmed the role of seven suspected INH resistance mutations and discovered five novel INH resistance mutations. All recombineered katG mutations conferred resistance to INH at a minimum inhibitory concentration of ≥0.25 μg/mL and should be added to the list of INH resistance determinants targeted by molecular diagnostic assays. We conclude that WGS is a superior method for detection of INH-R MTBC compared to current targeted molecular testing methods and could provide earlier diagnosis of drug-resistant TB. |
Agritourism and Kidding Season: A Large Outbreak of Human Shiga Toxin-Producing Escherichia coli O157 (STEC O157) Infections Linked to a Goat Dairy Farm-Connecticut, 2016.
Nichols MC , Gacek P , Phan Q , Gambino-Shirley KJ , Gollarza LM , Schroeder MN , Mercante A , Mullins J , Blackstock A , Laughlin ME , Olson SM , Pizzo E , Nguyen TN , Mank L , Holmes-Talbot K , McNutt A , Noel D , Muyombwe A , Razeq JH , Lis MJ , Sherman B , Kasacek W , Whitlock L , Strockbine N , Martin H , Vidyaprakash E , McCormack P , Cartter M . Front Vet Sci 2021 8 744055 The objective of this study was to determine sources of Shiga toxin-producing Escherichia coli O157 (STEC O157) infection among visitors to Farm X and develop public health recommendations. A case-control study was conducted. Case-patients were defined as the first ill child (aged <18 years) in the household with laboratory-confirmed STEC O157, or physician-diagnosed hemolytic uremic syndrome with laboratory confirmation by serology, who visited Farm X in the 10 days prior to illness. Controls were selected from Farm X visitors aged <18 years, without symptoms during the same time period as case-patients. Environment and animal fecal samples collected from Farm X were cultured; isolates from Farm X were compared with patient isolates using whole genome sequencing (WGS). Case-patients were more likely than controls to have sat on hay bales at the doe barn (adjusted odds ratio: 4.55; 95% confidence interval: 1.41-16.13). No handwashing stations were available; limited hand sanitizer was provided. Overall, 37% (29 of 78) of animal and environmental samples collected were positive for STEC; of these, 62% (18 of 29) yielded STEC O157 highly related by WGS to patient isolates. STEC O157 environmental contamination and fecal shedding by goats at Farm X was extensive. Farms should provide handwashing stations with soap, running water, and disposable towels. Access to animal areas, including animal pens and enclosures, should be limited for young children who are at risk for severe outcomes from STEC O157 infection. National recommendations should be adopted to reduce disease transmission. |
Using Wastewater Surveillance Data to Support the COVID-19 Response - United States, 2020-2021.
Kirby AE , Walters MS , Jennings WC , Fugitt R , LaCross N , Mattioli M , Marsh ZA , Roberts VA , Mercante JW , Yoder J , Hill VR . MMWR Morb Mortal Wkly Rep 2021 70 (36) 1242-1244 Wastewater surveillance, the measurement of pathogen levels in wastewater, is used to evaluate community-level infection trends, augment traditional surveillance that leverages clinical tests and services (e.g., case reporting), and monitor public health interventions (1). Approximately 40% of persons infected with SARS-CoV-2, the virus that causes COVID-19, shed virus RNA in their stool (2); therefore, community-level trends in SARS-CoV-2 infections, both symptomatic and asymptomatic (2) can be tracked through wastewater testing (3-6). CDC launched the National Wastewater Surveillance System (NWSS) in September 2020 to coordinate wastewater surveillance programs implemented by state, tribal, local, and territorial health departments to support the COVID-19 pandemic response. In the United States, wastewater surveillance was not previously implemented at the national level. As of August 2021, NWSS includes 37 states, four cities, and two territories. This report summarizes NWSS activities and describes innovative applications of wastewater surveillance data by two states, which have included generating alerts to local jurisdictions, allocating mobile testing resources, evaluating irregularities in traditional surveillance, refining health messaging, and forecasting clinical resource needs. NWSS complements traditional surveillance and enables health departments to intervene earlier with focused support in communities experiencing increasing concentrations of SARS-CoV-2 in wastewater. The ability to conduct wastewater surveillance is not affected by access to health care or the clinical testing capacity in the community. Robust, sustainable implementation of wastewater surveillance requires public health capacity for wastewater testing, analysis, and interpretation. Partnerships between wastewater utilities and public health departments are needed to leverage wastewater surveillance data for the COVID-19 response for rapid assessment of emerging threats and preparedness for future pandemics. |
Legionellosis cluster associated with working at a racetrack facility in West Virginia, 2018
Rispens JR , Hast M , Edens C , Ritter T , Mercante JW , Siegel M , Martin SB , Thomasson E , Barskey AE . J Environ Health 2021 83 (6) 14-19 In October 2018, the Centers for Disease Control and Prevention was notified of a cluster of Legionnaires' disease cases in workers at a racetrack facility. The objective of the resulting investigation was to determine the extent of the outbreak and identify potential sources of exposure to halt transmission. Case-finding and interviews were conducted among symptomatic racetrack workers who were known to be at the facility within 14 days prior to symptom onset. An environmental assessment of the facility and surrounding area was conducted for sources of potential Legionella exposure. In total, 17 legionellosis cases were identified. The environmental assessment revealed a poorly maintained hot tub in the jockey locker room as the most likely source. Further investigation identified deficiencies in the facility's ventilation systems, which suggested a transmission mechanism for workers who never entered the locker room floor. Considering indirect exposure routes via air handling systems can be useful for source identification and case-finding in legionellosis outbreaks. |
Knowledge and Practices Regarding Safe Household Cleaning and Disinfection for COVID-19 Prevention - United States, May 2020.
Gharpure R , Hunter CM , Schnall AH , Barrett CE , Kirby AE , Kunz J , Berling K , Mercante JW , Murphy JL , Garcia-Williams AG . MMWR Morb Mortal Wkly Rep 2020 69 (23) 705-709 A recent report described a sharp increase in calls to poison centers related to exposures to cleaners and disinfectants since the onset of the coronavirus disease 2019 (COVID-19) pandemic (1). However, data describing cleaning and disinfection practices within household settings in the United States are limited, particularly concerning those practices intended to prevent transmission of SARS-CoV-2, the virus that causes COVID-19. To provide contextual and behavioral insight into the reported increase in poison center calls and to inform timely and relevant prevention strategies, an opt-in Internet panel survey of 502 U.S. adults was conducted in May 2020 to characterize knowledge and practices regarding household cleaning and disinfection during the COVID-19 pandemic. Knowledge gaps were identified in several areas, including safe preparation of cleaning and disinfectant solutions, use of recommended personal protective equipment when using cleaners and disinfectants, and safe storage of hand sanitizers, cleaners, and disinfectants. Thirty-nine percent of respondents reported engaging in nonrecommended high-risk practices with the intent of preventing SARS-CoV-2 transmission, such as washing food products with bleach, applying household cleaning or disinfectant products to bare skin, and intentionally inhaling or ingesting these products. Respondents who engaged in high-risk practices more frequently reported an adverse health effect that they believed was a result of using cleaners or disinfectants than did those who did not report engaging in these practices. Public messaging should continue to emphasize evidence-based, safe practices such as hand hygiene and recommended cleaning and disinfection of high-touch surfaces to prevent transmission of SARS-CoV-2 in household settings (2). Messaging should also emphasize avoidance of high-risk practices such as unsafe preparation of cleaning and disinfectant solutions, use of bleach on food products, application of household cleaning and disinfectant products to skin, and inhalation or ingestion of cleaners and disinfectants. |
Lessons from the reestablishment of Public Health Laboratory activities in Puerto Rico after Hurricane Maria
Hardy MC , Stinnett RC , Kines KJ , Rivera-Nazario DM , Lowe DE , Mercante AM , Gonzalez Jimenez N , Cuevas Ruiz RI , Rivera Arbolay HI , Gonzalez Pena RL , Toro M , Trujillo AA , Pappas CL , Llewellyn AC , Candal F , Burgos Garay M , Gomez GA , Concepcion Acevedo J , Ansbro M , Moura H , Shaw MW , Muehlenbachs A , Romanoff LC , Sunshine BJ , Rose DA , Patel A , Shapiro CN , Luna-Pinto SC , Pillai SK , O'Neill E . Nat Commun 2019 10 (1) 2720 Public Health Laboratories (PHLs) in Puerto Rico did not escape the devastation caused by Hurricane Maria. We implemented a quality management system (QMS) approach to systematically reestablish laboratory testing, after evaluating structural and functional damage. PHLs were inoperable immediately after the storm. Our QMS-based approach began in October 2017, ended in May 2018, and resulted in the reestablishment of 92% of baseline laboratory testing capacity. Here, we share lessons learned from the historic recovery of the largest United States' jurisdiction to lose its PHL capacity, and provide broadly applicable tools for other jurisdictions to enhance preparedness for public health emergencies. |
Legionnaires disease at a hotel in Missouri, 2015: The importance of environmental health expertise in understanding water systems
Ahmed SS , Hunter CM , Mercante JW , Garrison LE , Turabelidze G , Kunz J , Cooley LA . J Environ Health 2019 81 (7) 8-13 During a Legionnaires disease outbreak at a Missouri hotel in 2015, the Centers for Disease Control and Prevention assisted state and local health departments to identify possible sources and transmission factors and to recommend improvements to water management. We performed an environmental assessment to understand the hotels water systems and identify areas of risk for Legionella amplifi cation and transmission. We obtained samples from the pool, spa, and potable water systems for Legionella culture. In the potable water system, we noted temperatures ideal for Legionella amplifi cation and areas of water stagnation. Additionally, we found inadequate documentation of pool and spa disinfection and maintenance. Of 40 water samples, Legionella pneumophila serogroup 1 that matched the sequence type of one available clinical isolate was recovered from five sink and shower fixtures. A comprehensive environmental assessment proved crucial to identifying maintenance issues in the hotels water systems and underscored the need for a water management program to reduce Legionnaires disease risk. |
Genomic heterogeneity differentiates clinical and environmental subgroups of Legionella pneumophila sequence type 1.
Mercante JW , Caravas JA , Ishaq MK , Kozak-Muiznieks NA , Raphael BH , Winchell JM . PLoS One 2018 13 (10) e0206110 Legionella spp. are the cause of a severe bacterial pneumonia known as Legionnaires' disease (LD). In some cases, current genetic subtyping methods cannot resolve LD outbreaks caused by common, potentially endemic L. pneumophila (Lp) sequence types (ST), which complicates laboratory investigations and environmental source attribution. In the United States (US), ST1 is the most prevalent clinical and environmental Lp sequence type. In order to characterize the ST1 population, we sequenced 289 outbreak and non-outbreak associated clinical and environmental ST1 and ST1-variant Lp strains from the US and, together with international isolate sequences, explored their genetic and geographic diversity. The ST1 population was highly conserved at the nucleotide level; 98% of core nucleotide positions were invariant and environmental isolates unassociated with human disease (n = 99) contained ~65% more nucleotide diversity compared to clinical-sporadic (n = 139) or outbreak-associated (n = 28) ST1 subgroups. The accessory pangenome of environmental isolates was also ~30-60% larger than other subgroups and was enriched for transposition and conjugative transfer-associated elements. Up to ~10% of US ST1 genetic variation could be explained by geographic origin, but considerable genetic conservation existed among strains isolated from geographically distant states and from different decades. These findings provide new insight into the ST1 population structure and establish a foundation for interpreting genetic relationships among ST1 strains; these data may also inform future analyses for improved outbreak investigations. |
Validation of novel Mycobacterium tuberculosis isoniazid resistance mutations not detectable by common molecular tests.
Kandler JL , Mercante AD , Dalton TL , Ezewudo MN , Cowan LS , Burns SP , Metchock B , Cegielski P , Posey JE . Antimicrob Agents Chemother 2018 62 (10) Resistance to the first-line anti-tuberculosis (TB) drug, isoniazid (INH), is widespread, and the mechanism of resistance is unknown in approximately 15% of INH-resistant (INH-R) strains. To improve molecular detection of INH-R TB, we used whole genome sequencing (WGS) to analyze 52 phenotypically INH-R Mycobacterium tuberculosis complex (MTBC) clinical isolates that lacked the common katG S315T or inhA promoter mutations. Approximately 94% (49/52) of strains had mutations at known INH-associated loci that were likely to confer INH resistance. All such mutations would be detectable by sequencing more DNA adjacent to existing target regions. Use of WGS minimized the chances of missing infrequent INH resistance mutations outside commonly targeted hotspots. We used recombineering to generate 12 observed clinical katG mutations in the pansusceptible H37Rv reference strain and determined their impact on INH resistance. Our functional genetic experiments have confirmed the role of seven suspected INH resistance mutations and discovered five novel INH resistance mutations. All recombineered katG mutations conferred resistance to INH at a minimum inhibitory concentration of >/=0.25 mug/mL and should be added to the list of INH resistance determinants targeted by molecular diagnostic assays. We conclude that WGS is a useful tool for detecting uncommon INH resistance mutations that would otherwise be missed by current targeted molecular testing methods, and suggest that its use (or use of expanded conventional or NGS-based targeted sequencing) may provide earlier diagnosis of INH-R TB. |
Initial public health laboratory response after Hurricane Maria - Puerto Rico, 2017
Concepcion-Acevedo J , Patel A , Luna-Pinto C , Pena RG , Cuevas Ruiz RI , Arbolay HR , Toro M , Deseda C , De Jesus VR , Ribot E , Gonzalez JQ , Rao G , De Leon Salazar A , Ansbro M , White BB , Hardy MC , Georgi JC , Stinnett R , Mercante AM , Lowe D , Martin H , Starks A , Metchock B , Johnston S , Dalton T , Joglar O , Stafford C , Youngblood M , Klein K , Lindstrom S , Berman L , Galloway R , Schafer IJ , Walke H , Stoddard R , Connelly R , McCaffery E , Rowlinson MC , Soroka S , Tranquillo DT , Gaynor A , Mangal C , Wroblewski K , Muehlenbachs A , Salerno RM , Lozier M , Sunshine B , Shapiro C , Rose D , Funk R , Pillai SK , O'Neill E . MMWR Morb Mortal Wkly Rep 2018 67 (11) 333-336 Hurricane Maria made landfall in Puerto Rico on September 20, 2017, causing major damage to infrastructure and severely limiting access to potable water, electric power, transportation, and communications. Public services that were affected included operations of the Puerto Rico Department of Health (PRDOH), which provides critical laboratory testing and surveillance for diseases and other health hazards. PRDOH requested assistance from CDC for the restoration of laboratory infrastructure, surveillance capacity, and diagnostic testing for selected priority diseases, including influenza, rabies, leptospirosis, salmonellosis, and tuberculosis. PRDOH, CDC, and the Association of Public Health Laboratories (APHL) collaborated to conduct rapid needs assessments and, with assistance from the CDC Foundation, implement a temporary transport system for shipping samples from Puerto Rico to the continental United States for surveillance and diagnostic and confirmatory testing. This report describes the initial laboratory emergency response and engagement efforts among federal, state, and nongovernmental partners to reestablish public health laboratory services severely affected by Hurricane Maria. The implementation of a sample transport system allowed Puerto Rico to reinitiate priority infectious disease surveillance and laboratory testing for patient and public health interventions, while awaiting the rebuilding and reinstatement of PRDOH laboratory services. |
Comparative genome analysis reveals a complex population structure of Legionella pneumophila subspecies.
Kozak-Muiznieks NA , Morrison SS , Mercante JW , Ishaq MK , Johnson T , Caravas J , Lucas CE , Brown E , Raphael BH , Winchell JM . Infect Genet Evol 2018 59 172-185 The majority of Legionnaires' disease (LD) cases are caused by Legionella pneumophila, a genetically heterogeneous species composed of at least 17 serogroups. Previously, it was demonstrated that L. pneumophila consists of three subspecies: pneumophila, fraseri and pascullei. During an LD outbreak investigation in 2012, we detected that representatives of both subspecies fraseri and pascullei colonized the same water system and that the outbreak-causing strain was a new member of the least represented subspecies pascullei. We used partial sequence based typing consensus patterns to mine an international database for additional representatives of fraseri and pascullei subspecies. As a result, we identified 46 sequence types (STs) belonging to subspecies fraseri and two STs belonging to subspecies pascullei. Moreover, a recent retrospective whole genome sequencing analysis of isolates from New York State LD clusters revealed the presence of a fourth L. pneumophila subspecies that we have termed raphaeli. This subspecies consists of 15 STs. Comparative analysis was conducted using the genomes of multiple members of all four L. pneumophila subspecies. Whereas each subspecies forms a distinct phylogenetic clade within the L. pneumophila species, they share more average nucleotide identity with each other than with other Legionella species. Unique genes for each subspecies were identified and could be used for rapid subspecies detection. Improved taxonomic classification of L. pneumophila strains may help identify environmental niches and virulence attributes associated with these genetically distinct subspecies. |
Legionnaires' Disease Outbreak Caused by Endemic Strain of Legionella pneumophila, New York, New York, USA, 2015.
Lapierre P , Nazarian E , Zhu Y , Wroblewski D , Saylors A , Passaretti T , Hughes S , Tran A , Lin Y , Kornblum J , Morrison SS , Mercante JW , Fitzhenry R , Weiss D , Raphael BH , Varma JK , Zucker HA , Rakeman JL , Musser KA . Emerg Infect Dis 2017 23 (11) 1784-1791 During the summer of 2015, New York, New York, USA, had one of the largest and deadliest outbreaks of Legionnaires' disease in the history of the United States. A total of 138 cases and 16 deaths were linked to a single cooling tower in the South Bronx. Analysis of environmental samples and clinical isolates showed that sporadic cases of legionellosis before, during, and after the outbreak could be traced to a slowly evolving, single-ancestor strain. Detection of an ostensibly virulent Legionella strain endemic to the Bronx community suggests potential risk for future cases of legionellosis in the area. The genetic homogeneity of the Legionella population in this area might complicate investigations and interpretations of future outbreaks of Legionnaires' disease. |
Characterization of Legionella Species from Watersheds in British Columbia, Canada.
Peabody MA , Caravas JA , Morrison SS , Mercante JW , Prystajecky NA , Raphael BH , Brinkman FSL . mSphere 2017 2 (4) Legionella spp. present in some human-made water systems can cause Legionnaires' disease in susceptible individuals. Although legionellae have been isolated from the natural environment, variations in the organism's abundance over time and its relationship to aquatic microbiota are poorly understood. Here, we investigated the presence and diversity of legionellae through 16S rRNA gene amplicon and metagenomic sequencing of DNA from isolates collected from seven sites in three watersheds with varied land uses over a period of 1 year. Legionella spp. were found in all watersheds and sampling sites, comprising up to 2.1% of the bacterial community composition. The relative abundance of Legionella tended to be higher in pristine sites than in sites affected by agricultural activity. The relative abundance levels of Amoebozoa, some of which are natural hosts of legionellae, were similarly higher in pristine sites. Compared to other bacterial genera detected, Legionella had both the highest richness and highest alpha diversity. Our findings indicate that a highly diverse population of legionellae may be found in a variety of natural aquatic sources. Further characterization of these diverse natural populations of Legionella will help inform prevention and control efforts aimed at reducing the risk of Legionella colonization of built environments, which could ultimately decrease the risk of human disease. IMPORTANCE Many species of Legionella can cause Legionnaires' disease, a significant cause of bacterial pneumonia. Legionella in human-made water systems such as cooling towers and building plumbing systems are the primary sources of Legionnaires' disease outbreaks. In this temporal study of natural aquatic environments, Legionella relative abundance was shown to vary in watersheds associated with different land uses. Analysis of the Legionella sequences detected at these sites revealed highly diverse populations that included potentially novel Legionella species. These findings have important implications for understanding the ecology of Legionella and control measures for this pathogen that are aimed at reducing human disease. |
Complete Genome Sequences of Legionella pneumophila subsp. fraseri Strains Detroit-1 and Dallas 1E.
Raphael BH , Kozak-Muiznieks NA , Morrison SS , Mercante JW , Winchell JM . Genome Announc 2017 5 (5) We report here the complete genome sequences of two of the earliest known strains of Legionella pneumophila subsp. fraseri Detroit-1 is serogroup 1 and was isolated from a lung biopsy specimen in 1977. Dallas 1E is serogroup 5 and was isolated in 1978 from a cooling tower. |
Notes from the field: Outbreak of Escherichia coli O157 infections associated with goat dairy farm visits - Connecticut, 2016
Laughlin M , Gambino-Shirley K , Gacek P , Phan Q , Stevenson L , Mercante A , Mullins J , Burnworth L , Blackstock A , Razeq JH , Cartter M , Nichols M . MMWR Morb Mortal Wkly Rep 2016 65 (5051) 1453-1454 On March 24, 2016, the Connecticut Department of Public Health (DPH) identified a cluster of seven culture-confirmed Shiga toxin–producing Escherichia coli (STEC) infections in patients ranging in age from 2 to 25 years. All seven patients reported bloody diarrhea; four were evaluated in an emergency department, three were hospitalized, and two developed hemolytic uremic syndrome (HUS). Six of the seven patients reported visiting the same goat dairy farm in southeastern Connecticut during the week preceding illness onset. An investigation was initiated by DPH, the Connecticut Department of Agriculture, CDC, and the local health district to determine the magnitude of the outbreak, identify risk factors and potential sources of infection, and develop recommendations to prevent further illnesses. A total of 50 confirmed cases of E. coli infection were associated with the outbreak, including 47 with an epidemiologic link to the goat farm. | PulseNet, the national molecular subtyping laboratory network for foodborne disease surveillance, and pulsed-field gel electrophoresis (PFGE) were used to identify the outbreak strains as STEC O157. A confirmed case was defined as 1) laboratory-confirmed E. coli O157 infection with the PFGE-identified outbreak strain or 2) physician-diagnosed HUS during March–April 2016 in a person with an epidemiologic link to the goat farm. Ill farm visitors were encouraged to contact DPH through a public statement released on March 28, 2016, and were interviewed about their visit. Environmental samples were collected at the farm and cultured, and the resulting bacterial isolates were compared with patient isolates using PFGE and whole genome sequencing. Genetic relatedness of isolates was determined by high-quality single nucleotide polymorphism analysis. The 2013 Compendium of Measures to Prevent Disease Associated with Animals in Public Settings was used to evaluate practices at the farm (1). |
Genomic Analysis Reveals Novel Diversity among the 1976 Philadelphia Legionnaires' Disease Outbreak Isolates and Additional ST36 Strains.
Mercante JW , Morrison SS , Desai HP , Raphael BH , Winchell JM . PLoS One 2016 11 (9) e0164074 Legionella pneumophila was first recognized as a cause of severe and potentially fatal pneumonia during a large-scale outbreak of Legionnaires' disease (LD) at a Pennsylvania veterans' convention in Philadelphia, 1976. The ensuing investigation and recovery of four clinical isolates launched the fields of Legionella epidemiology and scientific research. Only one of the original isolates, "Philadelphia-1", has been widely distributed or extensively studied. Here we describe the whole-genome sequencing (WGS), complete assembly, and comparative analysis of all Philadelphia LD strains recovered from that investigation, along with L. pneumophila isolates sharing the Philadelphia sequence type (ST36). Analyses revealed that the 1976 outbreak was due to multiple serogroup 1 strains within the same genetic lineage, differentiated by an actively mobilized, self-replicating episome that is shared with L. pneumophila str. Paris, and two large, horizontally-transferred genomic loci, among other polymorphisms. We also found a completely unassociated ST36 strain that displayed remarkable genetic similarity to the historical Philadelphia isolates. This similar strain implies the presence of a potential clonal population, and suggests important implications may exist for considering epidemiological context when interpreting phylogenetic relationships among outbreak-associated isolates. Additional extensive archival research identified the Philadelphia isolate associated with a non-Legionnaire case of "Broad Street pneumonia", and provided new historical and genetic insights into the 1976 epidemic. This retrospective analysis has underscored the utility of fully-assembled WGS data for Legionella outbreak investigations, highlighting the increased resolution that comes from long-read sequencing and a sequence type-matched genomic data set. |
Complete Genome Sequences of the Historical Legionella pneumophila Strains OLDA and Pontiac.
Mercante JW , Morrison SS , Raphael BH , Winchell JM . Genome Announc 2016 4 (4) Here, we report the complete genome sequences of Legionella pneumophila serogroup 1 strains OLDA and Pontiac, which predate the 1976 Philadelphia Legionnaires' disease outbreak. Strain OLDA was isolated in 1947 from an apparent sporadic case, and strain Pontiac caused an explosive outbreak at a Michigan health department in 1968. |
Complete Genome Sequences of Three Outbreak-Associated Legionella pneumophila Isolates.
Morrison SS , Desai HP , Mercante JW , Lapierre P , Raphael BH , Musser K , Winchell JM . Genome Announc 2016 4 (4) We report here the complete genome sequences of three Legionella pneumophila isolates that are associated with a Legionnaires' disease outbreak in New York in 2012. Two clinical isolates (D7630 and D7632) and one environmental isolate (D7631) were recovered from this outbreak. A single isolate-specific virulence gene was found in D7632. These isolates were included in a large study evaluating the genomic resolution of various bioinformatics approaches for L. pneumophila serogroup 1 isolates. |
Genomic resolution of outbreak-associated Legionella pneumophila serogroup 1 isolates from New York State.
Raphael BH , Baker DJ , Nazarian E , Lapierre P , Bopp D , Kozak-Muiznieks NA , Morrison SS , Lucas CE , Mercante JW , Musser KA , Winchell JM . Appl Environ Microbiol 2016 82 (12) 3582-90 A total of 30 Legionella pneumophila serogroup 1 isolates representing 10 separate legionellosis laboratory investigations ("outbreaks") that occurred in New York State between 2004 and 2012 were selected for evaluation of whole-genome sequencing (WGS) approaches for molecular subtyping of this organism. Clinical and environmental isolates were available for each outbreak and were initially examined by pulsed-field gel electrophoresis (PFGE). Sequence-based typing alleles were extracted from WGS data yielding complete sequence types (ST) for isolates representing 8 out of the 10 outbreaks evaluated in this study. Isolates from separate outbreaks sharing the same ST also contained the fewest differences in core genome single nucleotide polymorphisms (SNPs) and the greatest proportion of identical allele sequences in a whole-genome multilocus sequence typing (wgMLST) scheme. Both core SNP and wgMLST analyses distinguished isolates from separate outbreaks, including those from two outbreaks sharing indistinguishable PFGE profiles. Isolates from a hospital-associated outbreak spanning multiple years shared indistinguishable PFGE profiles but displayed differences in their genome sequences, suggesting the presence of multiple environmental sources. Finally, the rtx gene demonstrated differences in the repeat region sequence among ST1 isolates from different outbreaks, suggesting that variation in this gene may be useful for targeted molecular subtyping approaches for L. pneumophila This study demonstrates the utility of various genome sequence analysis approaches for L. pneumophila for environmental source attribution studies while furthering the understanding of Legionella ecology. IMPORTANCE: We demonstrate that whole-genome sequencing helps to improve resolution of Legionella pneumophila isolated during laboratory investigations of legionellosis compared to traditional subtyping methods. These data can be important in confirming the environmental sources of legionellosis outbreaks. Moreover, we evaluated various methods to analyze genome sequence data to help resolve outbreak-related isolates. |
Simultaneous detection of Legionella species and L. anisa, L. bozemanii, L. longbeachae and L. micdadei using conserved primers and multiple probes in a multiplex real-time PCR assay
Cross KE , Mercante JW , Benitez AJ , Brown EW , Diaz MH , Winchell JM . Diagn Microbiol Infect Dis 2016 85 (3) 295-301 Legionnaires' disease is a severe respiratory disease that is estimated to cause between 8,000 and 18,000 hospitalizations each year, though the exact burden is unknown due to under-utilization of diagnostic testing. Although Legionella pneumophila is the most common species detected in clinical cases (80-90%), other species have also been reported to cause disease. However, little is known about Legionnaires' disease caused by these non-pneumophila species. We designed a multiplex real-time PCR assay for detection of all Legionella spp. and simultaneous specific identification of four clinically-relevant Legionella species, L. anisa, L. bozemanii, L. longbeachae, and L. micdadei, using 5'-hydrolysis probe real-time PCR. The analytical sensitivity for detection of nucleic acid from each target species was ≤50fg per reaction. We demonstrated the utility of this assay in spiked human sputum specimens. This assay could serve as a tool for understanding the scope and impact of non-pneumophila Legionella species in human disease. |
Mycobacterium tuberculosis pncA polymorphisms that do not confer pyrazinamide resistance at a breakpoint concentration of 100 µg/ml in MGIT.
Whitfield MG , Warren RM , Streicher EM , Sampson SL , Sirgel FA , van Helden PD , Mercante A , Willby M , Hughes K , Birkness K , Morlock G , van Rie A , Posey JE . J Clin Microbiol 2015 Sequencing of the Mycobacterium tuberculosis pncA gene allows for pyrazinamide susceptibility testing. We summarize data on pncA polymorphisms which do not confer resistance at a susceptibility breakpoint of 100 mug/ml pyrazinamide in MGIT within a cohort of isolates from South Africa and the US Centre for Disease Control. |
Current and emerging Legionella diagnostics for laboratory and outbreak investigations
Mercante JW , Winchell JM . Clin Microbiol Rev 2015 28 (1) 95-133 Legionnaires' disease (LD) is an often severe and potentially fatal form of bacterial pneumonia caused by an extensive list of Legionella species. These ubiquitous freshwater and soil inhabitants cause human respiratory disease when amplified in man-made water or cooling systems and their aerosols expose a susceptible population. Treatment of sporadic cases and rapid control of LD outbreaks benefit from swift diagnosis in concert with discriminatory bacterial typing for immediate epidemiological responses. Traditional culture and serology were instrumental in describing disease incidence early in its history; currently, diagnosis of LD relies almost solely on the urinary antigen test, which captures only the dominant species and serogroup, Legionella pneumophila serogroup 1 (Lp1). This has created a diagnostic "blind spot" for LD caused by non-Lp1 strains. This review focuses on historic, current, and emerging technologies that hold promise for increasing LD diagnostic efficiency and detection rates as part of a coherent testing regimen. The importance of cooperation between epidemiologists and laboratorians for a rapid outbreak response is also illustrated in field investigations conducted by the CDC with state and local authorities. Finally, challenges facing health care professionals, building managers, and the public health community in combating LD are highlighted, and potential solutions are discussed. |
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