Journal of Clinical Microbiology

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ISSN / EISSN : 0095-1137 / 1098-660X
Published by: American Society for Microbiology (10.1128)
Total articles ≅ 46,311
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James A. Karlowsky, Melanie R. Baxter, Alyssa R. Golden, Heather J. Adam, , Philippe R. S. Lagacé-Wiens, George G. Zhanel
Journal of Clinical Microbiology; https://doi.org/10.1128/jcm.01635-21

Abstract:
Clinical isolates of Enterobacterales other than Escherichia coli (EOTEC), non-fermenting Gram-negative bacilli, and Gram-positive cocci were tested for susceptibility to fosfomycin using Etest ® and reference agar dilution. Applying EUCAST (v. 11.0, 2021) intravenous fosfomycin breakpoints, Etest ® MICs for EOTEC showed essential agreement (EA), categorical agreement (CA), major error (ME), and very major error (VME) rates of 70.4%, 88.4%, 4.1%, and 32.1%, respectively. No species of EOTEC tested with acceptable rates for all of EA (≥90%), CA (≥90%), ME (≤3%), and VME (≤3%). Etest ® MICs for Enterococcus faecalis , interpreted using CLSI oral/urine criteria (M100, 2021), showed EA, CA, minor error, ME, and VME rates of 98.5%, 81.2%, 18.8%, 0%, and 0%. Against Staphylococcus aureus , EA, CA, and ME rates were 84.1%, 98.7%, and 1.3% (EUCAST intravenous criteria). S. aureus isolates with fosfomycin MICs >32 μg/ml (resistant) were not identified by agar dilution. We conclude performing fosfomycin Etest ® on isolates of S. aureus will reliably identify fosfomycin-susceptible isolates with low, acceptable rates of MEs and VMEs. Testing of urinary isolates of E. faecalis by Etest ® is associated with an unacceptably high rate of minor errors (18.8%) but low, acceptable rates of MEs and VMEs when results are interpreted using CLSI criteria. Isolates of EOTEC tested by Etest ® with resulting MICs interpreted by EUCAST criteria were associated with an unacceptably high VME rate (32.1%). In vitro testing of clinical isolates beyond E. coli , E. faecalis , and S. aureus to determine susceptibility to fosfomycin is problematic with current methods and breakpoints.
, , , , Andreas van Arkel, Robbert G. Bentvelsen, Tom Rijpstra, Simone Van Der Sar-Van Der Brugge, Katrien Lagrou, Nico A.F. Janssen, et al.
Journal of Clinical Microbiology; https://doi.org/10.1128/jcm.01229-21

Abstract:
Background The literature regarding COVID-19 associated pulmonary aspergillosis (CAPA) has shown conflicting observations, including survival of CAPA patients not receiving antifungal therapy and discrepancy between CAPA diagnosis and autopsy findings. To gain insight into the pathophysiology of CAPA we performed a case-control study, in which we compared Aspergillus test profiles in CAPA patients and controls in relation to ICU-mortality. Methods A multinational case-control study, in which Aspergillus test results, use of antifungal therapy and mortality were collected from critically-ill COVID-19 patients. Patients were classified using the 2020 ECMM/ISHAM consensus case definitions. Results 219 critically-ill COVID-19 cases were analyzed, including one proven, 38 probable, 19 possible CAPA cases, 21 Aspergillus colonized patients, seven patients only positive for serum (1, 3)-ß-D-glucan (BDG), and 133 cases with no evidence of CAPA. Mortality was 53.8% in CAPA patients compared to 24.1% in patients without CAPA (p=0.001). Positive serum galactomannan (GM) and BDG were associated with increased mortality compared to serum biomarker negative CAPA patients (87.5% versus 41.7%, p=0.046; 90.0% versus 42.1%, p=0.029, respectively). For each point increase in GM or ten-point BDG serum concentration, the odds of death increased (GM, OR 10.208, 95%CI 1.621-64.291, p=0.013; BDG, OR 1.247, 95%CI 1.029-1.511, p=0.024). Conclusions CAPA is a complex disease, probably involving a continuum of respiratory colonization, tissue-invasion and angioinvasion. Serum biomarkers are useful for staging CAPA disease progression and, if positive, indicate angioinvasion and a high probability of mortality. There is need for a biomarker that distinguishes between respiratory tract colonization and tissue invasive CAPA disease.
, Ana Miqueleiz, María Eugenia Portillo, Miguel Fernández-Huerta, Ana Navascués, Pilar Sola Sara, Paula López Moreno, Gonzalo R Ordoñez, Jesús Castilla, Carmen Ezpeleta
Journal of Clinical Microbiology; https://doi.org/10.1128/jcm.01736-21

Abstract:
With the emergence of new SARS-CoV-2 variants and the acquisition of novel mutations in exiting lineages, the need to implement methods capable of monitoring viral dynamics arises. We report the emergence and spread of a new SARS-CoV-2 variant within B.1.575 lineage containing the E484K mutation in the spike protein (named B.1.575.2) in a region of Northern Spain between May and June 2021. SARS-CoV-2 positive samples with cycle threshold value less than or equal to 30 were selected to screen of presumptive variants using the TaqPath TM COVID-19 RT-PCR kit and TaqMan TM SARS-CoV-2 Mutation Panel. Confirmation of variants was performed by whole genome sequencing. Of the 200 samples belonging to the B.1.575 lineage, 194 (97%) corresponded to the B.1.575.2 sub-lineage, which was related to the presence of the E484K mutation. Of 197 cases registered in GISAID EpiCoV database as lineage B.1.575.2, 194 (99.5%) were identified in Pamplona (Spain) This report emphasizes the importance of complementing surveillance of SARS-CoV-2 with sequencing for the rapid control of emerging viral variants.
Stephanie Minnies, Byron W.P. Reeve, Loren Rockman, Georgina Nyawo, Charissa C. Naidoo, Natasha Kitchin, Cornelia Rautenbach, Colleen A. Wright, Andrew Whitelaw, Pawel Schubert, et al.
Journal of Clinical Microbiology; https://doi.org/10.1128/jcm.01316-21

Abstract:
Background: Tuberculosis lymphadenitis (TBL) is the most common extrapulmonary TB (EPTB) manifestation. Xpert MTB/RIF Ultra (Ultra) is a World Health Organization-endorsed diagnostic test, but performance data for TBL, including on non-invasive specimens, are limited. Methods: Fine needle aspiration biopsies (FNABs) from outpatients (≥18 years) with presumptive TBL (n=135) underwent: 1) routine Xpert (later Ultra once programmatically available), 2) a MGIT 960 culture (if Xpert- or Ultra-negative, or rifampicin-resistant), and 3) study Ultra. Concentrated paired urine underwent Ultra. Primary analyses used a microbiological reference standard (MRS). Results: In a head-to-head comparison (n=92) of FNAB study Ultra and Xpert, Ultra had increased sensitivity [91% (95% confidence interval 79, 98) vs. 72% (57, 84); p=0.016] and decreased specificity [76% (61, 87) vs. 93% (82, 99); p=0.020], and detected patients not on treatment. HIV nor alternative reference standards affected sensitivity and specificity. In patients with both routine and study Ultras, the latter detected more cases [+20% (0, 42); p=0.034] and, further indicative of potential laboratory-based room-for-improvement (e.g., specimen processing optimisation), false-negative study Ultras were more inhibited than true-positives. Study Ultra false-positives had less mycobacterial DNA than true-positives [trace-positive proportions 59% (13/22) vs. 12% (5/51); p<0.001]. “Trace” exclusion or recategorization removed potential benefits offered over Xpert. Urine Ultra had low sensitivity [18% (7, 35)]. Conclusions: Ultra on FNABs is highly sensitive and detects more TBL than Xpert. Patients with FNAB Ultra-positive “trace” results, most of whom will be culture-negative, may require additional clinical investigation. Urine Ultra could reduce the number of patients needing invasive sampling.
Mary Elizabeth Sexton,
Journal of Clinical Microbiology; https://doi.org/10.1128/jcm.01564-21

Abstract:
The utility of rapid antigen testing for SARS-CoV-2 is measured within the context for which it is applied; diagnostic accuracy must be considered in determining if rapid antigen testing is appropriate for the clinical situation. In this issue of the Journal of Clinical Microbiology , J.N. Kanji et al (J Clin Microbiol 59:e01411-21, 2021, https://doi.org/10.1128/JCM.01411-21 ) evaluate two rapid antigen tests that demonstrate high false-positive rates in asymptomatic healthcare workers. The assays may not be useful in situations where there is a shortage of staff, such as healthcare, since isolation would occur unnecessarily for these employees.
Carine Truyens, , Jackeline Alger, Maria Luisa Cafferata, Alvaro Ciganda, Luz Gibbons, , Sergio Sosa-Estani, Pierre Buekens
Journal of Clinical Microbiology; https://doi.org/10.1128/jcm.01062-21

Abstract:
Chagas disease is a neglected disease caused by Trypanosoma cruzi parasites. Most of diagnosis is based on serological tests but the lack of a gold standard test complicates the measurement of test performance. To overcome this limitation, we used samples from a cohort of well-characterized T. cruzi infected women to evaluate the reactivity of two rapid diagnostic tests and one ELISA assay. Our cohort derived from a previous study on congenital transmission of T. cruzi , and consisted in 481 blood/plasma samples from Argentina (n=149), Honduras (n=228) and Mexico (n=104) with at least one positive T. cruzi PCR. Reactivity of the three tests ranged from 70.5% for the Wiener ELISA to 81.0% for the T-Detect and 90.4% for the Stat-Pak rapid tests. Test reactivity varied significantly among countries, and was highest in Argentina, and lowest in Mexico. When considering at least two reactive serological tests to confirm seropositivity, over 12% of T. cruzi infection cases from Argentina were missed by serological tests, over 21% in Honduras, and an alarming 72% in Mexico. Differences in test performance among countries were not due to differences in parasitemia, but differences in antibody levels against ELISA test antigens were observed. Geographic differences in T. cruzi parasite strains as well as genetic differences among human populations may both contribute to the discrepancies in serological testing. Improvements in serological diagnostics for T. cruzi infections are critically needed to ensure an optimum identification of cases.
, Anna Aspán, Robert Söderlund, Annette Backhans, Marie Sjölund, Bengt Guss, Magdalena Jacobson
Journal of Clinical Microbiology; https://doi.org/10.1128/jcm.01297-21

Abstract:
Streptococcus suis is an important bacterial pathogen in pigs that may also cause zoonotic disease in humans. The aim of the study was to evaluate MALDI-TOF MS identification of S. suis case isolates from diseased pigs and tonsil isolates from healthy pigs and wild boar, using sequence analysis methods. Isolates (n=348) which had been classified as S. suis by MALDI-TOF MS were whole-genome sequenced and investigated using analysis of i) the 16S rRNA gene, ii) the recN gene, and iii) whole-genome average nucleotide identity (ANI). Analysis of the 16S rRNA gene indicated that 82.8% (288 out of 348) of the isolates were S. suis , while recN -gene analysis indicated that 75.6% (263 out of 348) were S. suis . ANI analysis classified 44.3% (154 out of 348) as S. suis . In total, 44% (153 out of 348) of the investigated isolates were classified as S. suis by all of the species identification methods employed. The mean MALDI-TOF MS score was significantly higher for the S. suis case isolates compared to the tonsil isolates, however, the difference is of limited practical use. The results show that species confirmation beyond MALDI-TOF MS is needed for S. suis isolates. Since the resolution of 16S rRNA gene analysis is too low for Streptococcus spp., ANI analysis with a slightly lowered cutoff of 94% may be used instead of, or in addition to, recN -gene analysis. Supplementation of the MALDI-TOF MS reference library with mass spectra from S. orisratti , S. parasuis , S. ruminantium , and additional S. suis serotypes, should be considered in order to produce more accurate classifications.
, Yuting Huang, Alyssa Lee, Xianming Zhu, Ruchee Shrestha, , Kirsten Littlefield, , Evan M. Bloch, Aaron A.R. Tobian, et al.
Journal of Clinical Microbiology; https://doi.org/10.1128/jcm.01186-21

Abstract:
Serologic, point-of-care tests to detect antibodies against SARS-CoV-2 are an important tool in the COVID-19 pandemic. The majority of current point-of-care antibody tests developed for SARS-CoV-2 rely on lateral flow assays, but these do not offer quantitative information. To address this, we developed a novel antibody test leveraging hemagglutination, employing a dry card format currently used for typing ABO blood groups. 200 COVID-19 patient and 200 control plasma samples were reconstituted with O-negative RBCs to form whole blood and added to dried viral-antibody fusion protein, followed by a stirring step and a tilting step, 3-minute incubation, and a second tilting step. The sensitivity for the hemagglutination test, Euroimmun IgG ELISA test and RBD-based CoronaChek lateral flow assay was 87.0%, 86.5%, and 84.5%, respectively, using samples obtained from recovered COVID-19 individuals. Testing pre-pandemic samples, the hemagglutination test had a specificity of 95.5%, compared to 97.3% and 98.9% for the ELISA and CoronaChek, respectively. A distribution of agglutination strengths was observed in COVID-19 convalescent plasma samples, with the highest agglutination score (4) exhibiting significantly higher neutralizing antibody titers than weak positives (2) (p<0.0001). Strong agglutinations were observed within 1 minute of testing, and this shorter assay time also increased specificity to 98.5%. In conclusion, we developed a novel rapid, point-of-care RBC agglutination test for the detection of SARS-CoV-2 antibodies that can yield semi-quantitative information on neutralizing antibody titer in patients. The five-minute test may find use in determination of serostatus prior to vaccination, post-vaccination surveillance and travel screening.
, Katherine Immergluck, Samuel D. Stampfer, Anuradha Rao, Leda Bassit, Max Su, Vi Nguyen, Victoria Stittleburg, Jessica M. Ingersoll, Heath L. Bradley, et al.
Journal of Clinical Microbiology; https://doi.org/10.1128/jcm.01446-21

Abstract:
To provide an accessible and inexpensive method to surveil for SARS-CoV-2 mutations, we developed a multiplex real-time RT-PCR (the Spike 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 log 10 GE/mL for the three initial targets (∼1-2 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 Ct values < 30 for the SARS-CoV-2 N2 target were detected, whereas 18/33 samples with N2 Ct values ≥ 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 receptor binding domain, and it can be quickly modified to detect new mutations that emerge.
, Mathew D. Esona, Rashi Gautam, Michael D. Bowen
Journal of Clinical Microbiology; https://doi.org/10.1128/jcm.02602-20

Abstract:
Since 2013, group A rotavirus strains characterized as novel DS-1-like inter-genogroup reassortant ‘equine-like G3’ strains have emerged and spread across five continents among human populations in at least 14 countries. Here we report a novel one-step TaqMan quantitative real-time reverse transcription-PCR assay developed to genotype and quantify the viral load for samples containing rotavirus equine-like G3 strains. Using a universal G forward primer and a newly designed reverse primer and TaqMan probe, we developed and validated an assay with a linear dynamic range of 2.3 × 10 9 – 227 copies per reaction and a limit of detection of 227 copies. The percent positive agreement, percent negative agreement, and precision of our assay were 100.00%, 99.63%, and 100.00%, respectively. This assay can simultaneously detect and quantify the viral load for samples containing DS-1-like inter-genogroup reassortant equine-like G3 strains with high sensitivity and specificity, faster turnaround time, and decreased cost and will be valuable for high-throughput screening of stool samples collected to monitor equine-like G3 strain prevalence and circulation among human populations throughout the world.
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