Fecal microbiota transplantation promotes reduction of antimicrobial resistance by strain replacement.

Multidrug-resistant organism (MDRO) colonization is a fundamental challenge in antimicrobial resistance. Limited studies have shown that fecal microbiota transplantation (FMT) can reduce MDRO colonization, but its mechanisms are poorly understood. We conducted a randomized, controlled trial of FMT for MDRO decolonization in renal transplant recipients called PREMIX (NCT02922816). Eleven participants were enrolled and randomized 1:1 to FMT or an observation period followed by delayed FMT if stool cultures were MDRO positive at day 36. Participants who were MDRO positive after one FMT were treated with a second FMT. At last visit, eight of nine patients who completed all treatments were MDRO culture negative. FMT-treated participants had longer time to recurrent MDRO infection versus PREMIX-eligible controls who were not treated with FMT. Key taxa (Akkermansia muciniphila, Alistipes putredinis, Phocaeicola dorei, Phascolarctobacterium faecium, Alistipes species, Mesosutterella massiliensis, Barnesiella intestinihominis, and Faecalibacterium prausnitzii) from the single feces donor used in the study that engrafted in recipients and metabolites such as short-chain fatty acids and bile acids in FMT-responding participants uncovered leads for rational microbiome therapeutic and diagnostic development. Metagenomic analyses revealed a previously unobserved mechanism of MDRO eradication by conspecific strain competition in an FMT-treated subset. Susceptible Enterobacterales strains that replaced baseline extended-spectrum ß-lactamase-producing strains were not detectable in donor microbiota manufactured as FMT doses but in one case were detectable in the recipient before FMT. These data suggest that FMT may provide a path to exploit strain competition to reduce MDRO colonization.

Concordance of SARS-CoV-2 Results in Self-collected Nasal Swabs vs Swabs Collected by Health Care Workers in Children and Adolescents.

Importance: Despite the expansion of SARS-CoV-2 testing, available tests have not received Emergency Use Authorization for performance with self-collected anterior nares (nasal) swabs from children younger than 14 years because the effect of pediatric self-swabbing on SARS-CoV-2 test sensitivity is unknown. Objective: To characterize the ability of school-aged children to self-collect nasal swabs for SARS-CoV-2 testing compared with collection by health care workers. Design, Setting, and Participants: Cross-sectional study of 197 symptomatic children and adolescents aged 4 to 14 years old. Individuals were recruited based on results of testing in the Children's Healthcare of Atlanta system from July to August 2021. Exposures: Children and adolescents were given instructional material consisting of a short instructional video and a handout with written and visual steps for self-swab collection. Participants first provided a self-collected nasal swab. Health care workers then collected a second specimen. Main Outcomes and Measures: The primary outcome was SARS-CoV-2 detection and relative quantitation by cycle threshold (Ct) in self- vs health care worker-collected nasal swabs when tested with a real-time reverse transcriptase-polymerase chain reaction test with Emergency Use Authorization. Results: Among the study participants, 108 of 194 (55.7%) were male and the median age was 9 years (IQR, 6-11). Of the 196 participants, 87 (44.4%) tested positive for SARS-CoV-2 and 105 (53.6%) tested negative by both self- and health care worker-collected swabs. Two children tested positive by self- or health care worker-collected swab alone; 1 child had an invalid health care worker swab. Compared with health care worker-collected swabs, self-collected swabs had 97.8% (95% CI, 94.7%-100.0%) and 98.1% (95% CI, 95.6%-100.0%) positive and negative percent agreement, respectively, and SARS-CoV-2 Ct values did not differ significantly between groups (mean [SD] Ct, self-swab: 26.7 [5.4] vs health care worker swab: 26.3 [6.0]; P = .65). Conclusions and Relevance: After hearing and seeing simple instructional materials, children and adolescents aged 4 to 14 years self-collected nasal swabs that closely agreed on SARS-CoV-2 detection with swabs collected by health care workers.

Association of Secretor Status and Recent Norovirus Infection With Gut Microbiome Diversity Metrics in a Veterans Affairs Population.

Norovirus infection causing acute gastroenteritis could lead to adverse effects on the gut microbiome. We assessed the association of microbiome diversity with norovirus infection and secretor status in patients from Veterans Affairs medical centers. Alpha diversity metrics were lower among patients with acute gastroenteritis but were similar for other comparisons.

Sampling for SARS-CoV-2 Aerosols in Hospital Patient Rooms.

Evidence varies as to how far aerosols spread from individuals infected with SARS-CoV-2 in hospital rooms. We investigated the presence of aerosols containing SARS-CoV-2 inside of dedicated COVID-19 patient rooms. Three National Institute for Occupational Safety and Health BC 251 two-stage cyclone samplers were set up in each patient room for a six-hour sampling period. Samplers were place on tripods, which each held two samplers at various heights above the floor. Extracted samples underwent reverse transcription polymerase chain reaction for selected gene regions of the SARS-CoV-2 virus nucleocapsid. Patient medical data were compared between participants in rooms where virus-containing aerosols were detected and those where they were not. Of 576 aerosols samples collected from 19 different rooms across 32 participants, 3% (19) were positive for SARS-CoV-2, the majority from near the head and foot of the bed. Seven of the positive samples were collected inside a single patient room. No significant differences in participant clinical characteristics were found between patients in rooms with positive and negative aerosol samples. SARS-CoV-2 viral aerosols were detected from the patient rooms of nine participants (28%). These findings provide reassurance that personal protective equipment that was recommended for this virus is appropriate given its spread in hospital rooms.

Single-Amplicon Multiplex Real-Time Reverse Transcription-PCR with Tiled Probes To Detect SARS-CoV-2 spike Mutations Associated with Variants of Concern.

To provide an accessible and inexpensive method to surveil for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) mutations, we developed a multiplex real-time reverse transcription-PCR (rRT-PCR) assay, the Spike single-nucleotide polymorphism (SNP) assay, to detect specific mutations in the spike receptor binding domain. A single primer pair was designed to amplify a 348-bp region of spike, and probes were initially designed to detect K417, E484K, and N501Y. The assay was evaluated using characterized variant sample pools and residual nasopharyngeal samples. Variant calls were confirmed by SARS-CoV-2 genome sequencing in a subset of samples. Subsequently, a fourth probe was designed to detect L452R. The lower limit of 95% detection was 2.46 to 2.48 log10 genome equivalents (GE)/ml for the three initial targets (∼1 to 2 GE/reaction). Among 253 residual nasopharyngeal swabs with detectable SARS-CoV-2 RNA, the Spike SNP assay was positive in 238 (94.1%) samples. All 220 samples with threshold cycle (CT) values of <30 for the SARS-CoV-2 N2 target were detected, whereas 18/33 samples with N2 CT values of ≥30 were detected. Spike SNP results were confirmed by sequencing in 50/50 samples (100%). Addition of the 452R probe did not affect performance for the original targets. The Spike SNP assay accurately identifies SARS-CoV-2 mutations in the receptor binding domain, and it can be quickly modified to detect new mutations that emerge.

Validation of High-Sensitivity Severe Acute Respiratory Syndrome Coronavirus 2 Testing for Stool-Toward the New Normal for Fecal Microbiota Transplantation.

INTRODUCTION: Mounting evidence demonstrates potential for fecal-oral transmission of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The US Food and Drug Administration now requires SARS-CoV-2 testing of potential feces donors before the use of stool manufactured for fecal microbiota transplantation. We sought to develop and validate a high-sensitivity SARS-CoV-2 reverse transcriptase polymerase chain reaction (RT-PCR) procedure for testing stool specimens. METHODS: A modified extraction method was used with an RT-PCR assay adapted from the Centers for Disease Control and Prevention PCR protocol for respiratory specimens. Contrived specimens were created using pre-COVID-19 banked stool specimens and spiking in known concentrations of SARS-CoV-2-specific nucleic acid. The highest transcript concentration at which 2/2 or 1/2 SARS-CoV-2 targets were detected in 9/10 replicates was defined as the dual-target limit and single-target limit of detection, respectively. The clinical performance of the assay was evaluated with stool samples collected from 17 nasopharyngeal swab RT-PCR-positive patients and 14 nasopharyngeal RT-PCR-negative patients. RESULTS: The dual-target and single-target limit of detection were 56 copies/µL and 3 copies/µL, respectively. SARS-CoV-2 was detected at concentrations as low as 0.6 copies/µL. Clinical stool samples from known COVID-19-positive patients demonstrated the detection of SARS-CoV-2 in stool up to 29 days from symptom onset with a high agreement with nasopharyngeal swab tests (kappa statistic of 0.95, P value < 0.001). DISCUSSION: The described RT-PCR test is a sensitive and flexible approach for the detection of SARS-CoV-2 in stool specimens. We propose an integrated screening approach that incorporates this stool test to support continuation of fecal microbiota transplantation programs.

Bioaerosol Sampling for Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) in a Referral Center with Critically Ill Coronavirus Disease 2019 (COVID-19) Patients March-May 2020.

BACKGROUND: Previous research has shown that rooms of patients with coronavirus disease 2019 (COVID-19) present the potential for healthcare-associated transmission through aerosols containing severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). However, data on the presence of these aerosols outside of patient rooms are limited. We investigated whether virus-containing aerosols were present in nursing stations and patient room hallways in a referral center with critically ill COVID-19 patients. METHODS: Eight National Institute for Occupational Safety and Health BC 251 2-stage cyclone samplers were set up throughout 6 units, including nursing stations and visitor corridors in intensive care units and general medical units, for 6 h each sampling period. Samplers were placed on tripods which held 2 samplers positioned 102 cm and 152 cm above the floor. Units were sampled for 3 days. Extracted samples underwent reverse transcription polymerase chain reaction for selected gene regions of the SARS-CoV-2 virus nucleocapsid and the housekeeping gene human RNase P as an internal control. RESULTS: The units sampled varied in the number of laboratory-confirmed COVID-19 patients present on the days of sampling. Some of the units included patient rooms under negative pressure, while most were maintained at a neutral pressure. Of 528 aerosol samples collected, none were positive for SARS-CoV-2 RNA by the estimated limit of detection of 8 viral copies/m3 of air. CONCLUSIONS: Aerosolized SARS-CoV-2 outside of patient rooms was undetectable. While healthcare personnel should avoid unmasked close contact with each other, these findings may provide reassurance for the use of alternatives to tight-fitting respirators in areas outside of patient rooms during the current pandemic.

Metagenomic Sequencing To Detect Respiratory Viruses in Persons under Investigation for COVID-19.

Broad testing for respiratory viruses among persons under investigation (PUIs) for SARS-CoV-2 has been performed inconsistently, limiting our understanding of alternative viral infections and coinfections in these patients. RNA metagenomic next-generation sequencing (mNGS) offers an agnostic tool for the detection of both SARS-CoV-2 and other RNA respiratory viruses in PUIs. Here, we used RNA mNGS to assess the frequencies of alternative viral infections in SARS-CoV-2 RT-PCR-negative PUIs (n = 30) and viral coinfections in SARS-CoV-2 RT-PCR-positive PUIs (n = 45). mNGS identified all viruses detected by routine clinical testing (influenza A [n = 3], human metapneumovirus [n = 2], and human coronavirus OC43 [n = 2], and human coronavirus HKU1 [n = 1]). mNGS also identified both coinfections (1, 2.2%) and alternative viral infections (4, 13.3%) that were not detected by routine clinical workup (respiratory syncytial virus [n = 3], human metapneumovirus [n = 1], and human coronavirus NL63 [n = 1]). Among SARS-CoV-2 RT-PCR-positive PUIs, lower cycle threshold (CT ) values correlated with greater SARS-CoV-2 read recovery by mNGS (R2, 0.65; P < 0.001). Our results suggest that current broad-spectrum molecular testing algorithms identify most respiratory viral infections among SARS-CoV-2 PUIs, when available and implemented consistently.

Performance evaluation of the Aptima HIV-1 RNA Quant assay on the Panther system using the standard and dilution protocols.

BACKGROUND: Currently, FDA-approved HIV-1 viral load (VL) assays use venipuncture-derived plasma. The Hologic Panther system uses 0.7 mL total volume for the Aptima HIV-1 Quant Assay standard (APT-Quant-std) and dilution (APT-Quant-dil) protocols. However, smaller plasma volumes from fingerstick whole blood (FSB) collected in EDTA-microtainer tubes (MCT) could provide an easier sample collection method for HIV-1 VL testing. OBJECTIVES: To evaluate the performance of the APT-Quant-std compared to the Roche CAP/CTM and Abbott m2000RT VL assays and an alternative APTQuant 1:7 dilution protocol, the latter using 100 µL of MCT-derived plasma from FSB. STUDY DESIGN: Linearity was determined using commercial HIV-1 RNA plasma controls. Dilutions ranging 1.56-2.95 log10 copies/mL were prepared to determine the APT-Quant-dil Limit of Quantitation (LOQ) using Probit analysis. Specificity of APT-Quant-std was calculated using 326 HIVnegative samples. To evaluate agreement, 329 plasma specimens were tested with APT-Quant-std, CAP/CTM, and m2000RT. Forty-seven matched venipuncture and MCT-derived plasma specimens were tested with APT-Quant-std and APT-Quant-dil. RESULTS: Among the RNA controls, specificity was 99.69 % for APT-Quant-std. The R2 values were 0.988 (APT-Quant-std/CAP/CTM), 0.980 (APT-Quant-std/ m2000RT), and 0.997 (APT-Quant-std/APT-Quant-dil). The APT-Quant-dil LOQ was estimated at 2.7 log10 copies/mL (500 copies/mL) (95 %CI 2.62-2.87). At 2.3 log10 copies/mL (200 copies/mL), the overall agreement was 91.0 % for APT-Quant-std/CAP/CTM, 85.7 % for APT-Quant-std/m2000RT, and 82.9 % for APT-Quant-std/APT-Quant-dil. Quantified APT-Quant-std results were on average 0.2 log10 copies/mL higher than CAP/CTM and m2000RT and 0.14 log10 copies/mL higher than APT-Quant-dil. CONCLUSION: APT-Quant showed similar performance compared to the CAP/CTM and m2000RT assays and remains sensitive and accurate using the dilution protocol.

Characterization of 2009 pandemic influenza A (H1N1) virus specimens with a positive hemagglutinin 1 signal in the Luminex xTAG respiratory viral panel assay.

This retrospective review analyzed Luminex xTAG respiratory viral panel (RVP) results for 2009 pandemic influenza A (H1N1) virus specimens. Comparing median fluorescence intensity (MFI) signals for the influenza A virus and hemagglutinin 1 (H1) reactions for specimens with very low positive (MFI < 1,000) or "no-call" H1 results reliably distinguished 2009 H1N1 from seasonal virus.