Antimicrobial Resistance Laboratory Network's multisite evaluation of the ThermoFisher Sensititre GN7F broth microdilution panel for antimicrobial susceptibility testing.

In 2017, the Centers for Disease Control and Prevention (CDC) established the Antimicrobial Resistance Laboratory Network to improve domestic detection of multidrug-resistant organisms. CDC and four laboratories evaluated a commercial broth microdilution panel. Antimicrobial susceptibility testing using the Sensititre GN7F (ThermoFisher Scientific, Lenexa, KS) was evaluated by testing 100 CDC and Food and Drug Administration AR Isolate Bank isolates [40 Enterobacterales (ENT), 30 Pseudomonas aeruginosa (PSA), and 30 Acinetobacter baumannii (ACB)]. We assessed multiple amounts of transfer volume (TV) between the inoculum and tubed 11-mL cation-adjusted Mueller-Hinton broth: 1 µL [tribe Proteeae (P-tribe) only] and 10, 30, and 50 µL, resulting in respective CFU per milliter of 1 × 104, 1 × 105, 3 × 105, and 5 × 105. Four TV combinations were analyzed: standard (STD) [1 µL (P-tribe) and 10 µL], enhanced standard (E-STD) [1 µL (P-tribe) and 30 µL], 30 µL, and 50 µL. Essential agreement (EA), categorical agreement, major error (ME), and very major error (VME) were analyzed by organism then TVs. For ENT, the average EA across laboratories was <90% for 7 of 15 ß-lactams using STD and E-STD TVs. As TVs increased, EA increased (>90%), and VMEs decreased. For PSA, EA improved as TVs increased; however, MEs also increased. For ACB, increased TVs provided slight EA improvements; all TVs yielded multiple VMEs and MEs. For ENT and ACB, Minimum inhibitory concentrations (MICs) trended downward using a 1 or 10 µL TV; there were no obvious MIC trends by TV for PSA. The public health and clinical consequences of missing resistance warrant increased TV of 30 µL for the GN7F, particularly for P-tribe, despite being considered "off-label" use.

Longitudinal serologic and viral testing post-SARS-CoV-2 infection and post-receipt of mRNA COVID-19 vaccine in a nursing home cohort-Georgia, October 2020‒April 2021.

There are limited data describing SARS-CoV-2-specific immune responses and their durability following infection and vaccination in nursing home residents. We conducted a prospective longitudinal evaluation of 11 consenting SARS-CoV-2-positive nursing home residents to evaluate the quantitative titers and durability of binding antibodies detected after SARS-CoV-2 infection and subsequent COVID-19 vaccination. The evaluation included nine visits over 150 days from October 25, 2020, through April 1, 2021. Visits included questionnaire administration, blood collection for serology, and paired anterior nasal specimen collection for testing by BinaxNOW™ COVID-19 Ag Card (BinaxNOW), reverse transcription polymerase chain reaction (RT-PCR), and viral culture. We evaluated quantitative titers of binding SARS-CoV-2 antibodies post-infection and post-vaccination (beginning after the first dose of the primary series). The median age among participants was 74 years; one participant was immunocompromised. Of 10 participants with post-infection serology results, 9 (90%) had detectable Pan-Ig, IgG, and IgA antibodies, and 8 (80%) had detectable IgM antibodies. At first antibody detection post-infection, two-thirds (6/9, 67%) of participants were RT-PCR-positive, but none were culture- positive. Ten participants received vaccination; all had detectable Pan-Ig, IgG, and IgA antibodies through their final observation ≤90 days post-first dose. Post-vaccination geometric means of IgG titers were 10-200-fold higher than post-infection. Nursing home residents in this cohort mounted robust immune responses to SARS-CoV-2 post-infection and post-vaccination. The augmented antibody responses post-vaccination are potential indicators of enhanced protection that vaccination may confer on previously infected nursing home residents.

Repeated antigen testing among severe acute respiratory coronavirus virus 2 (SARS-CoV-2)-positive nursing home residents.

Repeated antigen testing of 12 severe acute respiratory coronavirus virus 2 (SARS-CoV-2)-positive nursing home residents using Abbott BinaxNOW identified 9 of 9 (100%) culture-positive specimens up to 6 days after initial positive test. Antigen positivity lasted 2-24 days. Antigen positivity might last beyond the infectious period, but it was reliable in residents with evidence of early infection.

Prevalence of SARS-CoV-2 Antibodies in New York City Adults, June-October 2020: A Population-Based Survey.

BACKGROUND: Serosurveys help to ascertain burden of infection. Prior severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) serosurveys in New York City (NYC) used nonrandom samples. During June-October 2020, the NYC Health Department conducted a population-based survey estimating SARS-CoV-2 antibody prevalence in NYC adults. METHODS: Participants were recruited from the NYC 2020 Community Health Survey. We estimated citywide and stratified antibody prevalence using a hybrid design: serum tested with the DiaSorin LIAISON SARS-CoV-2 S1/S2 IgG assay and self-reported antibody test results were used together. We estimated univariate frequencies and 95% confidence intervals (CI), accounting for complex survey design. Two-sided P values ≤ .05 were statistically significant. RESULTS: There were 1074 respondents; 497 provided blood and 577 provided only a self-reported antibody test result. Weighted prevalence was 24.3% overall (95% CI, 20.7%-28.3%). Latino (30.7%; 95% CI, 24.1%-38.2%; P < .01) and black (30.7%; 95% CI, 21.9%-41.2%; P = .02) respondents had a higher weighted prevalence compared with white respondents (17.4%; 95% CI, 12.5%-23.7%). CONCLUSIONS: By October 2020, nearly 1 in 3 black and 1 in 3 Latino NYC adults had SARS-CoV-2 antibodies, highlighting unequal impacts of the coronavirus disease 2019 (COVID-19) pandemic on black and Latino NYC adults.

Performance Evaluation of Serial SARS-CoV-2 Rapid Antigen Testing During a Nursing Home Outbreak.

BACKGROUND: To address high COVID-19 burden in U.S. nursing homes, rapid SARS-CoV-2 antigen tests have been widely distributed in those facilities. However, performance data are lacking, especially in asymptomatic people. OBJECTIVE: To evaluate the performance of SARS-CoV-2 antigen testing when used for facility-wide testing during a nursing home outbreak. DESIGN: A prospective evaluation involving 3 facility-wide rounds of testing where paired respiratory specimens were collected to evaluate the performance of the BinaxNOW antigen test compared with virus culture and real-time reverse transcription polymerase chain reaction (RT-PCR). Early and late infection were defined using changes in RT-PCR cycle threshold values and prior test results. SETTING: A nursing home with an ongoing SARS-CoV-2 outbreak. PARTICIPANTS: 532 paired specimens collected from 234 available residents and staff. MEASUREMENTS: Percentage of positive agreement (PPA) and percentage of negative agreement (PNA) for BinaxNOW compared with RT-PCR and virus culture. RESULTS: BinaxNOW PPA with virus culture, used for detection of replication-competent virus, was 95%. However, the overall PPA of antigen testing with RT-PCR was 69%, and PNA was 98%. When only the first positive test result was analyzed for each participant, PPA of antigen testing with RT-PCR was 82% among 45 symptomatic people and 52% among 343 asymptomatic people. Compared with RT-PCR and virus culture, the BinaxNOW test performed well in early infection (86% and 95%, respectively) and poorly in late infection (51% and no recovered virus, respectively). LIMITATION: Accurate symptom ascertainment was challenging in nursing home residents; test performance may not be representative of testing done by nonlaboratory staff. CONCLUSION: Despite lower positive agreement compared with RT-PCR, antigen test positivity had higher agreement with shedding of replication-competent virus. These results suggest that antigen testing could be a useful tool to rapidly identify contagious people at risk for transmitting SARS-CoV-2 during nascent outbreaks and help reduce COVID-19 burden in nursing homes. PRIMARY FUNDING SOURCE: None.

Thinking outside the (pill) box: Does toxic metal exposure thwart antibiotic stewardship best practices?

Multi-antibiotic resistant (MAR) bacteria cost billions in medical care and tens of thousands of lives annually but perennial calls to limit agricultural and other misuse of antibiotics and to fund antibiotic discovery have not slowed this MAR deluge. Since mobile genetic elements (MGEs) stitch single antibiotic resistance genes into clinically significant MAR arrays, it is high time to focus on how MGEs generate MAR and how disabling them could ameliorate the MAR problem. However, to consider only antibiotics as the drivers of MAR is to miss the significant impact of exposure to non-antibiotic toxic chemicals, specifically metals, on the persistence and spread of MAR. Toxic metals were among the earliest discovered targets of plasmid-encoded resistance genes. Recent genomic epidemiology clearly demonstrated the co-prevalence of metal resistances and antibiotic multi-resistance, uniquely in humans and domestic animals. Metal resistances exploit the same, ancient "transportation infrastructure" of plasmids, transposons, and integrons that spread the antibiotic resistance genes and will continue to do so even if all antibiotic misuse were stopped today and new antibiotics were flowing from the pipeline monthly. In a key experiment with primates, continuous oral exposure to mercury (Hg) released from widely used dental amalgam fillings co-selected for MAR bacteria in the oral and fecal commensal microbiomes and, most importantly, when amalgams were replaced with non-metal fillings, MAR bacteria declined dramatically. Could that also be happening on the larger public health scale as use of amalgam restorations is curtailed or banned in many countries? This commentary covers salient past and recent findings of key metal-antibiotic resistance associations and proposes that the shift from phenotyping to genotyping in surveillance of resistance loci will allow a test of whether declining exposure to this leading source of Hg is accompanied by a decline in MAR compared to countries where amalgam is still used. If this hypothesis is correct, the limited success of antibiotic stewardship practices may be because MAR is also being driven by continuous, daily exposure to Hg, a non-antibiotic toxicant widely used in humans.

Correction to: Transcriptional responses of Escherichia coli during recovery from inorganic or organic mercury exposure.

After publication of the original article [1] the authors noted that the key displayed in Figure 1a was incorrect, as the PMA and HgCl2 conditions had been switched.

Transcriptional responses of Escherichia coli during recovery from inorganic or organic mercury exposure.

BACKGROUND: The protean chemical properties of mercury have long made it attractive for diverse applications, but its toxicity requires great care in its use, disposal, and recycling. Mercury occurs in multiple chemical forms, and the molecular basis for the distinct toxicity of its various forms is only partly understood. Global transcriptomics applied over time can reveal how a cell recognizes a toxicant and what cellular subsystems it marshals to repair and recover from the damage. The longitudinal effects on the transcriptome of exponential phase E. coli were compared during sub-acute exposure to mercuric chloride (HgCl2) or to phenylmercuric acetate (PMA) using RNA-Seq. RESULTS: Differential gene expression revealed common and distinct responses to the mercurials throughout recovery. Cultures exhibited growth stasis immediately after each mercurial exposure but returned to normal growth more quickly after PMA exposure than after HgCl2 exposure. Correspondingly, PMA rapidly elicited up-regulation of a large number of genes which continued for 30 min, whereas fewer genes were up-regulated early after HgCl2 exposure only some of which overlapped with PMA up-regulated genes. By 60 min gene expression in PMA-exposed cells was almost indistinguishable from unexposed cells, but HgCl2 exposed cells still had many differentially expressed genes. Relative expression of energy production and most metabolite uptake pathways declined with both compounds, but nearly all stress response systems were up-regulated by one or the other mercurial during recovery. CONCLUSIONS: Sub-acute exposure influenced expression of ~45% of all genes with many distinct responses for each compound, reflecting differential biochemical damage by each mercurial and the corresponding resources available for repair. This study is the first global, high-resolution view of the transcriptional responses to any common toxicant in a prokaryotic model system from exposure to recovery of active growth. The responses provoked by these two mercurials in this model bacterium also provide insights about how higher organisms may respond to these ubiquitous metal toxicants.

Organic and inorganic mercurials have distinct effects on cellular thiols, metal homeostasis, and Fe-binding proteins in Escherichia coli.

The protean chemical properties of the toxic metal mercury (Hg) have made it attractive in diverse applications since antiquity. However, growing public concern has led to an international agreement to decrease its impact on health and the environment. During a recent proteomics study of acute Hg exposure in E. coli, we also examined the effects of inorganic and organic Hg compounds on thiol and metal homeostases. On brief exposure, lower concentrations of divalent inorganic mercury Hg(II) blocked bulk cellular thiols and protein-associated thiols more completely than higher concentrations of monovalent organomercurials, phenylmercuric acetate (PMA) and merthiolate (MT). Cells bound Hg(II) and PMA in excess of their available thiol ligands; X-ray absorption spectroscopy indicated nitrogens as likely additional ligands. The mercurials released protein-bound iron (Fe) more effectively than common organic oxidants and all disturbed the Na(+)/K(+) electrolyte balance, but none provoked efflux of six essential transition metals including Fe. PMA and MT made stable cysteine monothiol adducts in many Fe-binding proteins, but stable Hg(II) adducts were only seen in CysXxx(n)Cys peptides. We conclude that on acute exposure: (a) the distinct effects of mercurials on thiol and Fe homeostases reflected their different uptake and valences; (b) their similar effects on essential metal and electrolyte homeostases reflected the energy dependence of these processes; and (c) peptide phenylmercury-adducts were more stable or detectable in mass spectrometry than Hg(II)-adducts. These first in vivo observations in a well-defined model organism reveal differences upon acute exposure to inorganic and organic mercurials that may underlie their distinct toxicology.

Discovering mercury protein modifications in whole proteomes using natural isotope distributions observed in liquid chromatography-tandem mass spectrometry.

The identification of peptides that result from post-translational modifications is critical for understanding normal pathways of cellular regulation as well as identifying damage from, or exposures to xenobiotics, i.e. the exposome. However, because of their low abundance in proteomes, effective detection of modified peptides by mass spectrometry (MS) typically requires enrichment to eliminate false identifications. We present a new method for confidently identifying peptides with mercury (Hg)-containing adducts that is based on the influence of mercury's seven stable isotopes on peptide isotope distributions detected by high-resolution MS. Using a pure protein and E. coli cultures exposed to phenyl mercuric acetate, we show the pattern of peak heights in isotope distributions from primary MS single scans efficiently identified Hg adducts in data from chromatographic separation coupled with tandem mass spectrometry with sensitivity and specificity greater than 90%. Isotope distributions are independent of peptide identifications based on peptide fragmentation (e.g. by SEQUEST), so both methods can be combined to eliminate false positives. Summing peptide isotope distributions across multiple scans improved specificity to 99.4% and sensitivity above 95%, affording identification of an unexpected Hg modification. We also illustrate the theoretical applicability of the method for detection of several less common elements including the essential element, selenium, as selenocysteine in peptides.