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Strains of two novel Corynebacterium species were cultured from samples of human nostrils and skin collected in the United States and Botswana. These strains demonstrated growth on Columbia Colistin-Nalidixic Acid agar with 5% sheep blood and in liquid media (brain heart infusion and tryptic soy broth) supplemented with Tween 80, a source of the fatty acid oleic acid. Cells were Gram-positive, non-spore-forming, non-motile bacilli that showed catalase but not oxidase activity. Major fatty acids in both of these species were 18:1 ω9c (oleic acid), 16:0 (palmitic acid), and 18:0 (stearic acid). Analysis of the 16S ribosomal RNA gene sequences identified these strains as belonging to the genus Corynebacterium (family Corynebacteriaceae). Whole-genome sequencing revealed that these strains formed distinct branches on a phylogenomic tree, with C. tuberculostearicum being the closest relative but with average nucleotide identities of < 95% relative to all previously described species. These results indicate that these strains represent novel species of Corynebacterium , for which we propose the names Corynebacterium hallux sp. nov., with the type strain CTNIH22 T (=ATCC TSD-435 T =DSM 117774 T ), and Corynebacterium nasorum sp. nov., with the type strain KPL3804 T (=ATCC TSD-439 T =DSM 117767 T ). We also describe the characteristics of two strains isolated from human nasal passages that are members of the recently named species Corynebacterium yonathiae ., Competing Interests: 1.7Conflicts of interest The authors declare that there are no conflicts of interest.

Four strains (MSK211, MSK294 T , MSK312, MSK433) of a novel Dolosigranulum species were cultured from nasopharyngeal swabs collected from mother-infant dyads in southern Botswana. These strains grew optimally on tryptic soy agar with 5% sheep blood solid medium and in fastidious bacteria broth. Colonies on tryptic soy agar with 5% sheep blood agar appeared grey or white with a flat, smooth surface and variable alpha haemolysis. Cells were Gram-positive, non-spore-forming, non-motile cocci that lacked catalase or oxidase activity. Major fatty acids were C 16 : 0 (palmitic acid), C 18 : 1  ω9 c (oleic acid), and C 18 : 0 (stearic acid). Analyses of 16S rRNA gene sequences identified these strains as belonging to the genus Dolosigranulum (family Carnobacteriaceae ), which currently contains only a single validly published species ( Dolosigranulum pigrum ). Whole-genome sequencing revealed that the genomes of these strains are 1.98-2.07 Mbp in size and have a G+C content of 39.6-39.9 mol%. Comparisons of these genomes to publicly available genomes of D. pigrum yielded average nucleotide identities and in silico DNA-DNA hybridization values of 92.3-92.9% and 49.1-51.4%, respectively. These results indicate that these strains represent a novel species of Dolosigranulum , for which we propose the name Dolosigranulum savutiense sp. nov., with the type strain MSK294 T (=DSM 117171 T =JCM 36673 T ).

This study compares three of the most inclusive and widely used panels for respiratory syndromic testing in the United States, namely, Luminex NxTAG Respiratory Pathogen Panel (RPP), BioFire FilmArray Respiratory Panel (RP), and GenMark eSensor Respiratory Viral Panel (RVP). We compared the three assays using nasopharyngeal swab samples ( n  = 350) collected from symptomatic patients ( n  = 329) in the pre-coronavirus disease 2019 (COVID-19) era. There was no significant difference in the overall accuracies of BioFire and Luminex assays ( P  = 0.6171); however, significant differences were found between BioFire and GenMark ( P  = 0.0003) and between GenMark and Luminex ( P  = 0.0009). The positive percent agreement of the BioFire RP assay was 94.1%, compared to 97.3% for GenMark RVP and 96.5% for Luminex RPP. Overall negative percent agreement values were high for all three assays, i.e., 99.9% for BioFire and Luminex and 99.5% for GenMark. The three assays were equivalent for adenovirus, human metapneumovirus, influenza A, and respiratory syncytial virus. Increased false-positive results were seen with BioFire for the endemic coronaviruses and with GenMark for influenza B and the parainfluenza viruses. IMPORTANCE Clinical laboratories have multiple choices when it is comes to syndromic respiratory testing. Here, the Luminex NxTAG RPP is compared to the BioFire FilmArray RP and GenMark eSensor RVP for overall and per-target accuracy. As new tests come to market, it is important to ascertain their performance characteristics, compared to other widely used in vitro diagnostic products.

This study was conducted to determine the prevalence of respiratory viral infections (RVI) in persons living with HIV (PLH) admitted with a respiratory complaint using real-time reverse transcription polymerase chain reaction and primer-independent next-generation sequencing (NGS). Of 82 subjects, respiratory viruses were the most common pathogen identified in 27 (33%), followed by fungus and bacteria in 8 (10%) and 4 (5%) subjects, respectively. Among subjects with RVI, 11 (41%) required ICU admission and 16 (59%) required mechanical ventilation. The proportion of respiratory viruses identified, and the associated complicated hospital course highlights the significant role that RVIs play in the lung health of PLH., (© 2020 The Authors. Influenza and Other Respiratory Viruses Published by John Wiley & Sons Ltd.)

Quantitative bacterial culture of bronchoalveolar lavage fluids (BALF) is labor-intensive, and the delay involved in performing culture, definitive identification, and susceptibility testing often results in prolonged use of broad-spectrum antibiotics. The Unyvero lower respiratory tract (LRT) panel (Curetis, Holzgerlingen, Germany) allows the multiplexed rapid detection and identification of 20 potential etiologic agents of pneumonia within 5 h of collection. In addition, the assay includes detection of gene sequences that confer antimicrobial resistance. We retrospectively compared the performance of the molecular panel to routine quantitative bacterial culture methods on remnant BALF. Upon testing 175 BALF, we were able to analyze positive agreement of 181 targets from 129 samples, and 46 samples were negative. The positive percent agreement (PPA) among the microbial targets was 96.5%, and the negative percent agreement (NPA) was 99.6%. The targets with a PPA of <100% were Staphylococcus aureus (34/37 [91.9%]), Streptococcus pneumoniae (10/11 [90.9%]), and Enterobacter cloacae complex (2/4 [50%]). For the analyzable resistance targets, concordance with phenotypic susceptibility testing was 79% (14/18). This study found the Unyvero LRT panel largely concordant with culture results; however, no outcome or clinical impact studies were performed., (Copyright © 2020 American Society for Microbiology.)

Molecular diagnostics for influenza and respiratory syncytial virus (RSV) have become commonplace, and various tests and systems have been cleared by the FDA for use in the United States. We performed a retrospective study to compare the Cepheid Xpress Flu/RSV assay with the Xpert Flu/RSV XC assay, using laboratory-developed tests (LDTs) as the reference method. The Xpress assay was 100% accurate compared to LDTs, whereas the Xpert Flu/RSV XC assay was 96.0% accurate. The Xpress test was determined to be faster and more sensitive than the XC assay., (Copyright © 2018 American Society for Microbiology.)

Clostridium difficile colonizes the gastrointestinal (GI) tract, resulting in either asymptomatic carriage or a spectrum of diarrheal illness. If clinical suspicion for C. difficile is low, stool samples are often submitted for analysis by multiplex molecular assays capable of detecting multiple GI pathogens, and some institutions do not report this organism due to concerns for high false-positive rates. Since clinical disease correlates with organism burden and molecular assays yield quantitative data, we hypothesized that numerical cutoffs could be utilized to improve the specificity of the Luminex xTAG GI pathogen panel (GPP) for C. difficile infection. Analysis of cotested liquid stool samples ( n = 1,105) identified a GPP median fluorescence intensity (MFI) value cutoff of ≥1,200 to be predictive of two-step algorithm (2-SA; 96.4% concordance) and toxin enzyme immunoassay (EIA) positivity. Application of this cutoff to a second cotested data set ( n = 1,428) yielded 96.5% concordance. To determine test performance characteristics, concordant results were deemed positive or negative, and discordant results were adjudicated via chart review. Test performance characteristics for the MFI cutoff of ≥150 (standard), MFI cutoff of ≥1,200, and 2-SA were as follows (respectively): concordance, 95, 96, and 97%; sensitivity, 93, 78, and 90%; specificity, 95, 98, and 98%; positive predictive value, 67, 82, and 81%;, and negative predictive value, 99, 98, and 99%. To capture the high sensitivity for organism detection (MFI of ≥150) and high specificity for active infection (MFI of ≥1,200), we developed and applied a reporting algorithm to interpret GPP data from patients ( n = 563) with clinician orders only for syndromic panel testing, thus enabling accurate reporting of C. difficile for 95% of samples (514 negative and 5 true positives) irrespective of initial clinical suspicion and without the need for additional testing., (Copyright © 2018 American Society for Microbiology.)

Setting: Xpert® MTB/RIF is the most widely used molecular assay for rapid diagnosis of tuberculosis (TB). The number of polymerase chain reaction cycles after which detectable product is generated (cycle threshold value, CT) correlates with the bacillary burden.OBJECTIVE To investigate the association between Xpert CT values and smear status through a systematic review and individual-level data meta-analysis., Design: Studies on the association between CT values and smear status were included in a descriptive systematic review. Authors of studies including smear, culture and Xpert results were asked for individual-level data, and receiver operating characteristic curves were calculated., Results: Of 918 citations, 10 were included in the descriptive systematic review. Fifteen data sets from studies potentially relevant for individual-level data meta-analysis provided individual-level data (7511 samples from 4447 patients); 1212 patients had positive Xpert results for at least one respiratory sample (1859 samples overall). ROC analysis revealed an area under the curve (AUC) of 0.85 (95%CI 0.82-0.87). Cut-off CT values of 27.7 and 31.8 yielded sensitivities of 85% (95%CI 83-87) and 95% (95%CI 94-96) and specificities of 67% (95%CI 66-77) and 35% (95%CI 30-41) for smear-positive samples., Conclusion: Xpert CT values and smear status were strongly associated. However, diagnostic accuracy at set cut-off CT values of 27.7 or 31.8 would not replace smear microscopy. How CT values compare with smear microscopy in predicting infectiousness remains to be seen.

Norovirus is the most common cause of sporadic gastroenteritis and outbreaks worldwide. The rapid identification of norovirus has important implications for infection prevention measures and may reduce the need for additional diagnostic testing. The Xpert Norovirus assay recently received FDA clearance for the detection and differentiation of norovirus genogroups I and II (GI and GII), which account for the vast majority of infections. In this study, we evaluated the performance of the Xpert Norovirus assay with both fresh, prospectively collected (n = 914) and frozen, archived (n = 489) fecal specimens. A Centers for Disease Control and Prevention (CDC) composite reference method was used as the gold standard for comparison. For both prospective and frozen specimens, the Xpert Norovirus assay showed positive percent agreement (PPA) and negative percent agreement (NPA) values of 98.3% and 98.1% for GI and of 99.4% and 98.2% for GII, respectively. Norovirus prevalence in the prospective specimens (collected from March to May of 2014) was 9.9% (n = 90), with the majority of positives caused by genogroup II (82%, n = 74). The positive predictive value (PPV) of the Xpert Norovirus assay was 75% for GI-positive specimens, whereas it was 86.5% for GII-positive specimens. The negative predictive values (NPV) for GI and GII were 100% and 99.9%, respectively., (Copyright © 2015, American Society for Microbiology. All Rights Reserved.)

Background: Methicillin-resistant Staphylococcus aureus (MRSA) infects ∼25% of patients with cystic fibrosis (CF) in the United States. We hypothesized that health-related outcomes differed between healthcare-associated (staphylococcal cassette chromosome mec [SCCmec] II) vs community-associated (SCCmec IV) MRSA strains in patients chronically infected with CF., Methods: At 7 CF centers, MRSA isolates were prospectively obtained from patients ≤18 years old with 2 or more positive MRSA cultures within 1 year. Isolates were classified by SCCmec type and Panton-Valentine-leukocidin (PVL) status at a core laboratory, and sites remained blinded to SCCmec type and PVL results. Prospective clinical data including antibiotic use, respiratory symptoms, and pulmonary exacerbations were obtained., Results: Among the 295 cohort participants with typeable MRSA isolates, 69.5% had SCCmec II PVL(-), 13.2% had SCCmec IV PVL(-), and 17.3% had SCCmec IV PVL(+) strains. During follow-up of 287 patients with prospective data after enrollment, the risk for pulmonary exacerbations was significantly higher among participants with SCCmec II than SCCmec IV strains (risk ratio [RR] = 1.13; P = .03) and higher in those with SCCmec IV PVL(-) than SCCmec IV PVL(+) strains (RR = 1.62; P < .0001). Neither decline in lung function nor changes in nutritional outcomes differed by SCCmec type or PVL status during the study period., Conclusions: Participants harboring chronic SCCmec II MRSA received more antibiotics and may have more lung disease than those with SCCmec IV; PVL(+) isolates were not associated with more advanced disease.

The Xpert Flu/RSV XC assay was compared to laboratory-developed tests (LDTs) (n = 207) and the Xpert Flu assay (n = 147) using archived nasopharyngeal swabs. The percentages of positive agreements with LDTs were 97.8% for influenza A, 97.2% for influenza B, and 89.3% for RSV. The sensitivity of influenza detection was improved with the Xpert Flu/RSV XC assay compared to the Xpert Flu assay., (Copyright © 2015, American Society for Microbiology. All Rights Reserved.)

Rationale: Methicillin-resistant Staphylococcus aureus (MRSA) prevalence continues to increase in patients with cystic fibrosis (CF) in the United States, reaching 26.5% in 2012. Approximately 30% of strains are SCCmec (staphylococcal cassette chromosome mec) IV type, frequently USA300, which in the general population have different genotypic and phenotypic features than SCCmec II type., Objectives: We hypothesized that risk factors for acquisition and outcomes in patients with CF differed for "health care-associated" (SCCmec II) versus "community-associated" (SCCmec IV) MRSA strains., Methods: To determine the role of SCCmec type and Panton-Valentine leukocidin (PVL), MRSA isolates from patients not more than 18 years old at seven CF centers were typed and the association of potential risk factors and subsequent clinical course was assessed, using data provided by the CF Patient Registry., Measurements and Main Results: Participants with chronic MRSA (295) had typeable isolates and clinical data; 205 (69.5%) had SCCmec II PVL(-), 39 (13.2%) had SCCmec IV PVL(-), and 51 (17.3%) had SCCmec IV PVL(+) strains. SCCmec IV, compared with SCCmec II, increased during the study period, 1996-2010 (P = 0.03). SCCmec II was associated with Pseudomonas aeruginosa-positive cultures and three or more clinic visits in the 6 months preceding the first positive MRSA culture (adjusted odds ratio, 2.05; 95% confidence interval, 1.13-3.74; P = 0.019). Lung function and anthropometrics remained unchanged in the 6 months after initial MRSA detection compared with the 6 months prior. Although CF care increased for participants in both groups in the 6 months after MRSA detection, inhaled antibiotics were prescribed more frequently in those with SCCmec II strains and increased hospitalizations occurred in those with SCCmec IV PVL(-) strains compared with those with PVL(+) strains (adjusted difference, 34.10%; 95% confidence interval, 7.58-60.61; P = 0.012). Participants in both groups had an increase in CF care in the 2 years after MRSA detection compared with the 2 years prior., Conclusions: Increased exposure to CF clinics and P. aeruginosa may constitute risk factors for acquisition of SCCmec II MRSA strains. Clinical interventions increased 6 months and 2 years after initial MRSA detection regardless of SCCmec type.

Background: In the United States, individuals with presumptive pulmonary tuberculosis are placed in airborne infection isolation (AII) and assessed by smear microscopy on 3 respiratory specimens collected 8-24 hours apart. Xpert MTB/RIF assay (Xpert) on 1, 2, or 3 specimens may be more efficient for determining AII discontinuation., Methods: This single-center, observational cohort study of inpatients with presumptive pulmonary tuberculosis enrolled adults with 1 or more sputum specimens submitted for smear microscopy. Smear microscopy and Xpert were performed on each sputum specimen. Clinicians were blinded to Xpert results. The primary endpoint was AII duration. Secondary endpoints were laboratory processing time, strategy-based tuberculosis detection, and sensitivity and specificity., Results: Among 207 subjects, the median AII duration was 68.0 hours (interquartile range [IQR], 47.1-97.5) for smear microscopy compared with 20.8 hours (IQR, 16.8-32.0) for the 1-specimen Xpert, 41.2 hours (IQR, 26.6-54.8) for the 2-specimen Xpert, and 54.0 hours (IQR, 43.3-80.0) for the 3-specimen Xpert strategies (P ≤ .004). Median laboratory processing time for smear microscopy was 2.5 times as long as Xpert (P < .001). The 2- and 3-specimen Xpert and smear microscopy strategies captured all 6 tuberculosis cases. The 1-specimen Xpert strategy missed 1 case. No difference was observed between smear microscopy and Xpert in sensitivity or specificity for detection of Mycobacterium tuberculosis., Conclusions: Xpert-based strategies significantly reduced AII duration compared with the smear-based strategy. The 2-specimen Xpert strategy was most efficient in minimizing AII time while identifying all tuberculosis cases among individuals with presumptive tuberculosis in this low-burden setting., (© The Author 2014. Published by Oxford University Press on behalf of the Infectious Diseases Society of America. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.)

Objectives: The prevalence of methicillin-resistant Staphylococcus aureus (MRSA) in cystic fibrosis (CF) patients in the United States is approximately 25%. Little is known about the relative proportion of hospital- versus community-associated strains or the antimicrobial susceptibility of MRSA in different CF centers. We hypothesized that the majority of MRSA isolates obtained from children with CF are those endemic in the hospital and that those associated with community acquisition (SCCmec IV) would be more resistant than typically seen in non-CF MRSA isolates., Methods: We studied MRSA strains from seven pediatric CF centers to determine the clonal distribution based on DNA sequencing of the staphylococcal protein A gene (spa typing), the type of staphylococcal chromosomal cassette mec (SCCmec), and the proportion of strains with Panton-Valentine leukocidin (PVL). Antimicrobial susceptibility to systemic and topical antibiotics was compared between different MRSA types., Results: We analyzed 277 MRSA isolates from unique patients (mean age 11.15 ± 4.77 years, 55% male). Seventy % of isolates were SCCmec II PVL negative and the remainder SCCmec IV. Overall 17% MRSA strains were PVL positive (all SCCmec IV). Spa typing of 118 isolates showed most of the SCCmec II strains being t002, while SCCmec IV PVL positive isolates were t008, and SCCmec IV PVL negative isolates represented a variety of spa-types. The proportions of SCCmec II strains and spa-types were similar among centers. Overall rates of resistance to trimethoprim-sulfamethoxazole (4%), tetracycline (7%), tigecycline (0.4%), linezolid (0.4%) as well as fosfomycin (0.4%), fusidic acid (3%), and mupirocin (1%) were low. No strains were resistant to vancomycin. SCCmec II strains had higher rates of resistance to ciprofloxacin and clindamycin (P < 0.001) than SCCmec IV strains., Conclusions: In this U.S. study, most MRSA isolates in the pediatric CF population were SCCmec II PVL negative. Rates of resistance were low, including to older and orally available antibiotics such as trimethoprim-sulfamethoxazole., (© 2013 Wiley Periodicals, Inc.)

There are several U.S. FDA-cleared molecular respiratory virus panels available today, each with advantages and disadvantages. This study compares four multiplex panels, the BioFire Diagnostics FilmArray RP (respiratory panel), the GenMark Dx eSensor RVP (respiratory viral panel), the Luminex xTAG RVPv1, and the Luminex xTAG RVP fast. Three hundred specimens (200 retrospective and 100 consecutive) were tested using all four platforms to determine performance characteristics. The overall sensitivity and specificity, respectively, and 95% confidence interval (CI; in parentheses) for each panel were as follows: FilmArray RP, 84.5% (79.2, 88.6) and 100% (96.2, 100); eSensor RVP, 98.3% (95.5, 99.5) and 99.2% (95.4, 100); xTAG RVPv1, 92.7% (88.5, 95.4) and 99.8% (96.0, 100); and xTAG RVP fast, 84.4% (78.5, 88.9) and 99.9% (96.1, 100). The sensitivity of each assay fluctuated by viral target, with the greatest discrepancies noted for adenovirus and influenza virus B detection. Hands-on time and time to result were recorded and ease of use was assessed to generate a complete profile of each assay.

Using retrospective (n = 200) and prospective (n = 150) nasopharyngeal specimens, we evaluated the Nanosphere Verigene RV+ and the Focus Diagnostics Simplexa Flu A/B & RSV tests. Overall, RV+ demonstrated sensitivities and specificities of 96.6% and 100% for influenza A virus, 100% and 99.7% for influenza B virus, and 100% and 100% for respiratory syncytial virus (RSV), while the Simplexa test sensitivities and specificities were 82.8 and 99.7%, 76.2 and 100%, and 94.6 and 100%, respectively.

The Cepheid Xpert MTB/RIF research-use-only (RUO) assay and a laboratory-developed test (LDT) targeting IS6110 were evaluated and compared to mycobacterial culture as the gold standard. The performance characteristics of both molecular assays were determined by using 112 specimens from 90 patients, including 89 pulmonary specimens and 23 extrapulmonary specimens. Of the specimens tested, 37 (33%) were culture positive for the Mycobacterium tuberculosis complex; 29 were pulmonary, and 8 were extrapulmonary. Of these culture-positive specimens, 83% of the pulmonary specimens and 50% of the extrapulmonary specimens were smear positive. There was complete concordance between the smear-positive culture-positive specimens, independent of the anatomical site (100% sensitivity). The sensitivity of the MTB/RIF RUO assay for smear-negative specimens was 60% for pulmonary and 75% for extrapulmonary specimens, while the IS6110 LDT sensitivities were 40% and 0%, respectively. There was also complete concordance among the culture-negative specimens tested. Both assays showed 95% specificity, with four culture-negative specimens testing as positive. A review of patient records indicated that there was a high likelihood of the presence of M. tuberculosis complex DNA in the false-positive specimens. Biosafety analysis was performed and showed an acceptable reduction in organism viability using the processing methods described above. Both molecular assays are suitable for the detection of M. tuberculosis isolates in smear-positive pulmonary and extrapulmonary specimens, while the sensitivity of the detection of M. tuberculosis isolates in smear-negative specimens was variable.

We performed a retrospective (n = 121) and prospective (n = 305) verification of the Cepheid Xpert Flu assay to determine its performance characteristics. The overall sensitivity and specificity were 93% and 100%, respectively. Nasopharyngeal specimen sensitivities were 100% for seasonal influenza A/H1 virus and influenza A/H3 virus, 90% for influenza A/2009/H1N1 virus, and 95% for influenza B virus.

Children attending child care centers (CCCs) are at increased risk for infections, including those caused by methicillin-resistant Staphylococcus aureus (MRSA). Nasal colonization often precedes infection, and MRSA colonization has been associated with increased infection risk. Community-associated MRSA (CA-MRSA) has caused increased MRSA infections in the general population, including children. Little is known about the frequency of MRSA nasal colonization in young children, particularly in those attending CCCs where disease transmission is common. We sampled the nares of 1,163 children in 200 classrooms from 24 CCCs in North Carolina and Virginia to assess S. aureus colonization. MRSA strains were molecularly analyzed for staphylococcal cassette chromosome mec (SCCmec) type, Panton-Valentine leukocidin status, and multilocus sequence type. A case-control study was performed to identify risk factors for MRSA colonization. We found that 18.1% children were colonized with S. aureus and 1.3% with MRSA. Molecular analysis of the MRSA strains identified 47% as CA-MRSA and 53% as health care-associated MRSA (HA-MRSA). Although two centers had multiple children colonized with MRSA, genotyping indicated that no transmission had occurred within classrooms. The case-control study did not detect statistically significant risk factors for MRSA colonization. However, MRSA-colonized children were more likely to be nonwhite and to have increased exposure to antibiotics and skin infections in the home. Both CA-MRSA and HA-MRSA strains were found colonizing the nares of children attending CCCs. The low frequency of colonization observed highlights the need for a large multicenter study to determine risk factors for MRSA colonization and subsequent infection in this highly susceptible population.