Patel, Robin

Abstract Background Next-generation sequencing (NGS) is increasingly used for periprosthetic joint infection (PJI) diagnosis, but its clinical utility is poorly defined. Shotgun metagenomic sequencing (sNGS) has been reported to identify PJI pathogens undetected by culture in sonicate fluid. However, sNGS is complex and costly. Here, 16S ribosomal RNA (rRNA) gene-based targeted metagenomic sequencing (tNGS) was compared to sNGS of sonicate fluid for microbial detection and identification in patients with total hip arthroplasty (THA) and total knee arthroplasty (TKA) failure. Methods A convenience sample of sonicate fluids derived from patients who had undergone THA or TKA removal, enriched with culture negative PJI cases, was tested. Samples had been previously tested by sNGS. For tNGS, samples were extracted, amplified by polymerase chain reaction targeting the V1 to V3 regions of the 16S rRNA gene, and sequenced on an Illumina MiSeq. Results A total of 395 sonicate fluids, including 208 from subjects with PJI, were studied. Compared with sonicate fluid culture, tNGS had higher positive percent agreement (72.1 vs 52.9%, P < .001), detecting potential pathogens in 48.0% of culture-negative PJIs. There was no difference between the positive percent agreement of tNGS (72.1%) and sNGS (73.1%, P = .83). Conclusions 16S rRNA gene-based tNGS is a potential diagnostic tool for PJI pathogen identification in sonicate fluid from failed THAs and TKAs in culture-negative cases, with similar performance characteristics to sNGS.

Whole-genome sequencing (WGS) is rapidly replacing traditional typing methods for the investigation of infectious disease outbreaks. Additionally, WGS data are being used to predict phenotypic antimicrobial susceptibility. Acinetobacter baumannii , which is often multidrug-resistant, is a significant culprit in outbreaks in health care settings.

Abstract Background Increasing antimicrobial resistance combined with a lagging pipeline of novel antimicrobial compounds have resulted in a resurgence of interest in phage therapy. To select optimal phage or phage combinations for patients for whom phage therapy is considered, assessment of activity of a panel of phages against the patients’ bacterial isolate(s) should ideally be performed. Classical phage susceptibility testing methods (i.e., agar overlay) may be laborious, with expertise outside of normal training and competency of medical laboratory science staff needed. Content Adaptive Phage Therapeutics™ leveraged a commercially available phenotyping system (Biolog OmniLog®) to generate the PhageBank Susceptibility Test™, which uses a custom data analysis pipeline (PhageSelect™) to measure the delay in reaching log-phase metabolic activity (“hold time”) when a given isolate is challenged with a specific phage. The goal of this study was to preliminarily assess reproducibility of this approach by testing 2 bacterial species at 2 sites, APT and an academic site. Nineteen Escherichia coli phages were tested against 18 bacterial isolates, and 21 Staphylococcus aureus phages, against 11 bacterial isolates. Result comparisons were statistically excellent for E. coli (κ = 0.7990) and good/fair for S. aureus (κ = 0.6360). Summary The described method provides good/fair to excellent statistical reproducibility for assessment of phage susceptibility of 2 commonly encountered bacterial species.

Elizabethkingia species are Gram-negative bacilli that were most recently linked to a cluster of infections in the Midwestern United States from 2016 to 2017. Inappropriate empirical and directed antibiotic selection for this organism is common among providers and is an independent risk factor for mortality.

The prevalence of carbapenem-resistant Klebsiella pneumoniae (CR Kp ) infections in nosocomial settings remains a public health challenge. High-risk clones such as clonal group 258 (CG258) are particularly concerning due to their association with bla KPC carriage, which can severely complicate antimicrobial treatments.

Abstract Background The yield of next-generation sequencing (NGS) added to a Sanger sequencing–based 16S ribosomal RNA (rRNA) gene polymerase chain reaction (PCR) assay was evaluated in clinical practice for diagnosis of bacterial infection. Methods PCR targeting the V1 to V3 regions of the 16S rRNA gene was performed, with amplified DNA submitted to Sanger sequencing and/or NGS (Illumina MiSeq) or reported as negative, depending on the cycle threshold value. A total of 2146 normally sterile tissues or body fluids were tested between August 2020 and March 2021. Clinical sensitivity was assessed in 579 patients from whom clinical data were available. Results Compared with Sanger sequencing alone (400 positive tests), positivity increased by 87% by adding NGS (347 added positive tests). Clinical sensitivity of the assay that incorporated NGS was 53%, which was higher than culture (42%, P < .001), with an impact on clinical decision-making in 14% of infected cases. Clinical sensitivity in the subgroup that received antibiotics at sampling was 41% for culture and 63% for the sequencing assay (P < .001). Conclusions Adding NGS to Sanger sequencing of the PCR-amplified 16S rRNA gene substantially improved test positivity. In the patient population studied, the assay was more sensitive than culture, especially in patients who had received antibiotic therapy.

The epidemiology of macrolide resistance in Mycoplasma ( Mycoplasmoides ) pneumoniae in the United States is incompletely described. Using a PCR assay targeting common mutations associated with macrolide resistance in M. pneumoniae (23S rRNA gene, A2063G/A2064G), the frequency of macrolide resistance was estimated to be 10% based on analysis of 114 samples tested from January 2014 to September 2021 at Mayo Clinic Laboratories.

Surgical site infections (SSI) are a significant burden to patients and health care systems. They increase mortality rates, length of hospital stays, and associated health care costs.

Ureaplasma -induced hyperammonemia is a deadly complication affecting around 4% of lung transplant recipients and, to a lesser extent, other solid organ transplant patients. Understanding the underlying mechanisms will inform patient management, potentially decreasing mortality and morbidity. Here, it is shown that uremia is a plausible contributing factor to the pathophysiology of the condition.