Genome sequencing and analysis reveals possible determinants of Staphylococcus aureus nasal carriage

<p>Abstract</p> <p>Background</p> <p>Nasal carriage of <it>Staphylococcus aureus </it>is a major risk factor in clinical and community settings due to the range of etiologies caused by the organism. We have identified unique immunological and ultrastructural properties associated with nasal carriage isolates denoting a role for bacterial factors in nasal carriage. However, despite extensive molecular level characterizations by several groups suggesting factors necessary for colonization on nasal epithelium, genetic determinants of nasal carriage are unknown. Herein, we have set a genomic foundation for unraveling the bacterial determinants of nasal carriage in <it>S. aureus.</it></p> <p>Results</p> <p>MLST analysis revealed no lineage specific differences between carrier and non-carrier strains suggesting a role for mobile genetic elements. We completely sequenced a model carrier isolate (D30) and a model non-carrier strain (930918-3) to identify differential gene content. Comparison revealed the presence of 84 genes unique to the carrier strain and strongly suggests a role for Type VII secretion systems in nasal carriage. These genes, along with a putative pathogenicity island (SaPIBov) present uniquely in the carrier strains are likely important in affecting carriage. Further, PCR-based genotyping of other clinical isolates for a specific subset of these 84 genes raise the possibility of nasal carriage being caused by multiple gene sets.</p> <p>Conclusion</p> <p>Our data suggest that carriage is likely a heterogeneic phenotypic trait and implies a role for nucleotide level polymorphism in carriage. Complete genome level analyses of multiple carriage strains of <it>S. aureus </it>will be important in clarifying molecular determinants of <it>S. aureus </it>nasal carriage.</p

Does the presence of multiple β-lactamases in Gram-negative bacilli impact the results of antimicrobial susceptibility tests and extended-spectrum β-lactamase and carbapenemase confirmation methods?

Objectives: Many multidrug-resistant Gram-negative bacilli (MDR-GNB) harbour multiple β-lactamases. The aim of this study was to assess the impact of multiple β-lactamase carriage on the accuracy of susceptibility tests and extended-spectrum β-lactamase (ESBL) and carbapenemase confirmation methods. Methods: A total of 50 MDR-GNB, of which 29 carried multiple β-lactamases, underwent broth microdilution (BMD) and disk diffusion (DD) testing as well as confirmation tests for ESBLs and carbapenemases. Whole-genome sequencing (WGS) was used for β-lactamase gene identification. Results: Categorical agreement of BMD and DD testing results ranged from 86.5 to 97.7% for 10 β-lactam agents. BMD and DD algorithms for ESBL detection were highly variable; 6 of 8 positive strains carried an ESBL plus a carbapenemase or an AmpC enzyme, which may confound antimicrobial selection. The sensitivity and specificity of the modified carbapenem inactivation method (mCIM) were both 100%, whilst mCIM and EDTA-modified carbapenem inactivation method (eCIM) when used together to differentiate serine from metallo-β-lactamase carriage were both 96%. Xpert® Carba-R results (in vitro diagnostic test) were consistent with WGS results. Predicting phenotypic carbapenem resistance from WGS data overall showed 100% specificity but only 66.7% sensitivity for Enterobacterales isolates that were non-susceptible to imipenem and meropenem. Conclusions: Multiple β-lactamases in MDR-GNB does not impact DD results, the utility of mCIM/eCIM tests, or Xpert Carba-R results. However, ESBL algorithms produced inconsistent results and predicting carbapenem resistance from WGS data was problematic in such strains

Does the Presence of Multiple β-Lactamases in Gram-Negative Bacilli Impact the Results of Antimicrobial Susceptibility Tests and Extended-Spectrum β-Lactamase and Carbapenemase Confirmation Methods?

Objectives: Many multidrug-resistant Gram-negative bacilli (MDR-GNB) harbour multiple β-lactamases. The aim of this study was to assess the impact of multiple β-lactamase carriage on the accuracy of susceptibility tests and extended-spectrum β-lactamase (ESBL) and carbapenemase confirmation methods. Methods: A total of 50 MDR-GNB, of which 29 carried multiple β-lactamases, underwent broth microdilution (BMD) and disk diffusion (DD) testing as well as confirmation tests for ESBLs and carbapenemases. Whole-genome sequencing (WGS) was used for β-lactamase gene identification. Results: Categorical agreement of BMD and DD testing results ranged from 86.5 to 97.7% for 10 β-lactam agents. BMD and DD algorithms for ESBL detection were highly variable; 6 of 8 positive strains carried an ESBL plus a carbapenemase or an AmpC enzyme, which may confound antimicrobial selection. The sensitivity and specificity of the modified carbapenem inactivation method (mCIM) were both 100%, whilst mCIM and EDTA-modified carbapenem inactivation method (eCIM) when used together to differentiate serine from metallo-β-lactamase carriage were both 96%. Xpert® Carba-R results (in vitro diagnostic test) were consistent with WGS results. Predicting phenotypic carbapenem resistance from WGS data overall showed 100% specificity but only 66.7% sensitivity for Enterobacterales isolates that were non-susceptible to imipenem and meropenem. Conclusions: Multiple β-lactamases in MDR-GNB does not impact DD results, the utility of mCIM/eCIM tests, or Xpert Carba-R results. However, ESBL algorithms produced inconsistent results and predicting carbapenem resistance from WGS data was problematic in such strains

Updating Molecular Diagnostics for Detecting Methicillin- Susceptible and Methicillin-Resistant \u3cem\u3eStaphylococcus aureus \u3c/em\u3eIsolates in Blood Culture Bottles

Molecular diagnostic tests can be used to provide rapid identification of staphylococcal species in blood culture bottles to help improve antimicrobial stewardship. However, alterations in the target nucleic acid sequences of the microorganisms or their antimicrobial resistance genes can lead to false-negative results. We determined the whole-genome sequences of 4 blood culture isolates of Staphylococcus aureus and 2 control organisms to understand the genetic basis of genotypephenotype discrepancies when using the Xpert MRSA/SA BC test (in vitro diagnostic medical device [IVD]). Three methicillin-resistant S. aureus (MRSA) isolates each had a different insertion of a genetic element in the staphylococcal cassette chromosome (SCCmec)-orfX junction region that led to a misclassification as methicillin-susceptible S. aureus (MSSA). One strain contained a deletion in spa, which produced a false S. aureus-negative result. A control strain of S. aureus that harbored an SCCmec element but no mecA (an empty cassette) was correctly called MSSA by the Xpert test. The second control contained an SCCM1 insertion. The updated Xpert MRSA/SA BC test successfully detected both spa and SCCmec variants of MRSA and correctly identified empty-cassette strains of S. aureus as MSSA. Among a sample of 252 MSSA isolates from the United States and Europe, 3.9% contained empty SCCmec cassettes, 1.6% carried SCCM1, \u3c1% had spa deletions, and \u3c1% contained SCCmec variants other than those with SCCM1. These data suggest that genetic variations that may interfere with Xpert MRSA/SA BC test results remain rare. Results for all the isolates were correct when tested with the updated assay

Whole-exome sequencing-based discovery of STIM1 deficiency in a child with fatal classic Kaposi sarcoma

Whole-exome sequencing reveals a homozygous splice-site mutation in the gene encoding STIM1 in a child with classic Kaposi sarcoma

\u3cem\u3eStreptococcus agalactiae \u3c/em\u3eStrains with Chromosomal Deletions Evade Detection with Molecular Methods

Surveillance of circulating microbial populations is critical for monitoring the performance of a molecular diagnostic test. In this study, we characterized 31 isolates of Streptococcus agalactiae (group B Streptococcus [GBS]) from several geographic locations in the United States and Ireland that contain deletions in or adjacent to the region of the chromosome that encodes the hemolysin gene cfb, the region targeted by the Xpert GBS and GBS LB assays. PCR-negative, culture-positive isolates were recognized during verification studies of the Xpert GBS assay in 12 laboratories between 2012 and 2018. Whole-genome sequencing of 15 GBS isolates from 11 laboratories revealed four unique deletions of chromosomal DNA ranging from 181 bp to 49 kb. Prospective surveillance studies demonstrated that the prevalence of GBS isolates containing deletions in the convenience sample wa

Incidence and Tracking of Escherichia coli O157:H7 in a Major Produce Production Region in California

Fresh vegetables have become associated with outbreaks caused by Escherichia coli O157:H7 (EcO157). Between 1995–2006, 22 produce outbreaks were documented in the United States, with nearly half traced to lettuce or spinach grown in California. Outbreaks between 2002 and 2006 induced investigations of possible sources of pre-harvest contamination on implicated farms in the Salinas and San Juan valleys of California, and a survey of the Salinas watershed. EcO157 was isolated at least once from 15 of 22 different watershed sites over a 19 month period. The incidence of EcO157 increased significantly when heavy rain caused an increased flow rate in the rivers. Approximately 1000 EcO157 isolates obtained from cultures of>100 individual samples were typed using Multi-Locus Variable-number-tandem-repeat Analysis (MLVA) to assist in identifying potential fate and transport of EcO157 in this region. A subset of these environmental isolates were typed by Pulse Field Gel Electrophoresis (PFGE) in order to make comparisons with human clinical isolates associated with outbreak and sporadic illness. Recurrence of identical and closely related EcO157 strains from specific locations in the Salinas and San Juan valleys suggests that transport of the pathogen is usually restricted. In a preliminary study, EcO157 was detected in water at multiple locations in a low-flow creek only within 135 meters of a point source. However, possible transport up to 32 km was detected during periods of higher water flow associated with flooding. During the 2006 baby spinach outbreak investigation, transport was also detected where water was unlikely to be involved. These results indicate that contamination of the environment is a dynamic process involving multiple sources and methods of transport. Intensive studies of the sources, incidence, fate and transport of EcO157 near produce production are required to determine the mechanisms of pre-harvest contamination and potential risks for human illness

Origins Space Telescope: Baseline mission concept

The Origins Space Telescope will trace the history of our origins from the time dust and heavy elements permanently altered the cosmic landscape to present-day life. How did galaxies evolve from the earliest galactic systems to those found in the Universe today? How do habitable planets form? How common are life-bearing worlds? To answer these alluring questions, Origins will operate at mid-and far-infrared (IR) wavelengths and offer powerful spectroscopic instruments and sensitivity three orders of magnitude better than that of the Herschel Space Observatory, the largest telescope flown in space to date. We describe the baseline concept for Origins recommended to the 2020 US Decadal Survey in Astronomy and Astrophysics. The baseline design includes a 5.9-m diameter telescope cryocooled to 4.5 K and equipped with three scientific instruments. A mid-infrared instrument (Mid-Infrared Spectrometer and Camera Transit spectrometer) will measure the spectra of transiting exoplanets in the 2.8 to 20 μm wavelength range and offer unprecedented spectrophotometric precision, enabling definitive exoplanet biosignature detections. The far-IR imager polarimeter will be able to survey thousands of square degrees with broadband imaging at 50 and 250 μm. The Origins Survey Spectrometer will cover wavelengths from 25 to 588 μm, making wide-area and deep spectroscopic surveys with spectral resolving power R ∼ 300, and pointed observations at R ∼ 40,000 and 300,000 with selectable instrument modes. Origins was designed to minimize complexity. The architecture is similar to that of the Spitzer Space Telescope and requires very few deployments after launch, while the cryothermal system design leverages James Webb Space Telescope technology and experience. A combination of current-state-of-the-art cryocoolers and next-generation detector technology will enable Origins\u27 natural background-limited sensitivity