Metagenomic sequencing for infectious disease diagnostics is an important tool that holds promise for use in the clinical laboratory. Challenges for implementation so far include high cost, the length of time to results, and the need for technical and bioinformatics expertise. However, the recent technological innovation of nanopore sequencing from Oxford Nanopore Technologies (ONT) has the potential to address these challenges. ONT sequencing is an attractive platform for clinical laboratories to adopt due to its low cost, rapid turnaround time, and user-friendly bioinformatics pipelines. However, this method still faces the problem of base-calling accuracy compared to other platforms. This review highlights the general challenges of pathogen detection in clinical specimens by metagenomic sequencing, the advantages and disadvantages of the ONT platform, and how research to date supports the potential future use of nanopore sequencing in infectious disease diagnostics.

Fungal infections are a rising threat to our immunocompromised patient population as well as other non-immunocompromised patients with various medical conditions. However, little progress has been made in the past decade to improve fungal diagnostics. To jointly address this diagnostic challenge, the Fungal Diagnostics Laboratory Consortium (FDLC) was recently created. The FDLC consists of 26 laboratories from the United States and Canada that routinely provide fungal diagnostic services for patient care. A survey of fungal diagnostic capacity among the 26 members of the FDLC was recently completed, identifying the following diagnostic gaps: lack of molecular detection of mucormycosis; lack of an optimal diagnostic algorithm incorporating fungal biomarkers and molecular tools for early and accurate diagnosis of Pneumocystis pneumonia, aspergillosis, candidemia, and endemic mycoses; lack of a standardized molecular approach to identify fungal pathogens directly in formalin-fixed paraffin-embedded tissues; lack of robust databases to enhance mold identification with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry; suboptimal diagnostic approaches for mold blood cultures, tissue culture processing for Mucorales, and fungal respiratory cultures for cystic fibrosis patients; inadequate capacity for fungal point-of-care testing, to detect and identify new, emerging or under-recognized, rare or uncommon fungal pathogens, and performance of antifungal susceptibility testing. In this commentary, the FDLC delineates the most pressing unmet diagnostic needs and provides expert opinion on how to fulfill them. Most importantly, the FDLC provides a robust laboratory network to tackle these diagnostic gaps and ultimately to improve and enhance the clinical laboratory’s capability to rapidly and accurately diagnose fungal infections.

bRectal swabs from high-risk patients were screened for carbapenem-resistant organisms (CROs) using several methods. The direct MacConkey plate method was the most sensitive for CROs (95%), while chromID CARBA and the Check-Direct CPE screen assay were the most sensitive for the detection of carbapenemase-producing organisms (CPOs) (100%; all bla KPC ). All methods had a specificity of >90% for CROs, and for CPOs, the specificity ranged from 85 to 98%. Broth enrichment methods performed poorly compared to direct inoculation methods, negating the need for the broth enrichment step. In 2013, the U.S. Centers for Disease Control and Prevention (CDC) assigned the highest threat level to carbapenem-resistant Enterobacteriaceae (CRE). Additionally, the CDC designated multidrug-resistant (MDR) Pseudomonas aeruginosa and Acinetobacter baumannii as serious threats because they are resistant to nearly all available antibiotics, including carbapenems-declaring that they require urgent public health attention (1). Optimal screening methods for rapid detection of carbapenem-resistant organisms (CROs) have yet to be established (2). Currently described methods include broth enrichment, direct selective culture, chromogenic media, and detection of carbapenemase genes directly from rectal swabs (3-9). The objectives of this study were to (i) evaluate multiple methods for screening CROs from rectal swabs and (ii) to determine the prevalence of gastrointestinal colonization with CROs among high-risk inpatient populations.Two-hundred thirteen remnant vancomycin-resistant enterococcus (VRE) surveillance rectal ESwabs (Copan, Murrieta, CA) were collected in a non-outbreak setting from four distinct inpatient populations at The Johns Hopkins Hospital, Baltimore, MD. ESwabs were collected upon hospital unit admission and weekly thereafter until unit discharge. Consecutive rectal ESwabs were collected from medical and surgical intensive care units (MICUs and SICUs, respectively), an oncology ward, and an organ transplant ward over a 2-week period. The ESwabs were vortexed, and the remnant liquid Amies broth was aliquoted to cryovials and frozen at Ϫ70°C until further testing was performed. From each ESwab broth, five different methods for the detection of CRO were set up in parallel. The five methods included (i) the CDC broth enrichment method with ertapenem for selection (3), (ii) a modified CDC broth enrichment method using ertapenem and vancomycin for selection (3), (iii) a direct MacConkey plate with ertapenem disks (4, 6), (iv) a chromogenic chromID CARBA agar plate method (with the new reformulated medium; package insert version 20157 A-en-2013/02, reference no. 414012 [bioMérieux, Marcy l'Étoile, France]), and (v) the Check-Direct CPE screen assay for the BD MAX instrument (Check-Points, Wageningen, The Netherlands; Becton Dickinson, Sparks, MD). The ESwab broth was first vortexed for 5 s, 100 l of ESwab broth was inoculated into each of the medium types, and 25 l was inoculated into a DNA sample buffer tube (SBT) for the Check...

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