Mycoplasmoides genitalium nucleic acid semi-quantitation and molecular macrolide resistance detection via automated assays: gender and specimen source considerations.

A laboratory-developed test (LDT) using analyte-specific reagents has been optimized on a commercial platform to detect macrolide resistance-associated mutations (MRM) in 23S rRNA from Mycoplasmoides genitalium from primary clinical specimens. In this study, MRM-LDT was applied to a multi-specimen source study set. One thousand four hundred ninety-five primary specimens testing positive for M. genitalium by commercial transcription-mediated amplification (TMA) were initially titered by the TMA assay using serial 10-fold dilutions to semi-quantitate target nucleic acid burden. Primary specimens were then processed for MRM detection using the MRM-LDT. Findings were stratified by gender and specimen source. The mean log10 target nucleic acid titer of a TMA-positive specimen was 3.51 (median 3; range 0-10). Male specimens (n = 1145) demonstrated a mean log10 M. genitalium TMA titer of 3.67; that value observed in 350 female specimens was 2.98 (P < 0.0001). The MRM-LDT detection rate (88.7%) from specimens with log10 M. genitalium TMA titers ≥ 4 was increased over specimens with log10 titers ≤ 1 (4.5%; P < 0.0002). In females, MRM-LDT was positive from 51.3% of vaginal swab and 34.7% of urine specimens (P = 0.01). In males, MRM-LDT was positive from 65.0% of rectal swab and 55.7% of urine specimens (P = 0.002). Differences were also observed in log10 M. genitalium TMA titers as a function of specimen source. M. genitalium macrolide resistance rates among multiple specimen sources, as determined by MRM-LDT, are high in the United States and can be consistent with target nucleic acid burden within the primary specimen. Caveats experienced within subgroupings support MRM reflex testing on primary M. genitalium-positive specimens. IMPORTANCE: First-line macrolide treatment failure is of increasing concern with Mycoplasmoides genitalium in multiple settings. Recent sexually-transmitted infection treatment guidelines from the United States Centers for Disease Control and Prevention have predicated therapeutic approaches on the availability of a macrolide resistance/susceptibility result from a primary clinical specimen. In this report, we investigate potential correlation between macrolide resistance mutation detection rates (identified by a molecular amplified laboratory-developed test) and transcription-mediated amplification-based rRNA target semi-quantitation. Data reveal that rRNA semi-quantitation and laboratory-developed test detection rate differences exist as a function of gender and specimen source. These data can guide providers in proper specimen selection not only for the laboratory diagnosis of M. genitalium but also macrolide resistance mutation determination from primary clinical specimens.

Mycoplasmoides genitalium Macrolide Resistance Detection is Needed in University Settings.

Background: Mycoplasmoides genitalium remains a difficult sexually-transmitted infection (STI) to manage due to its potential for antimicrobial resistance and post-infection sequelae. University students are especially vulnerable, as this demographic has the highest rate of STI in the United States. As a result, investigating prevalence rates and therapeutic outcomes in this population is essential to minimize future impact of M. genitalium The purpose of this study was to investigate a university student population for M. genitalium distribution and treatment outcome.Design: Retrospective chart-review of university health clinic attendees, augmented by laboratory detection of M. genitalium following therapeutic intervention.Methods: A total of 1617 student encounters at a midwestern United States university health clinic over a 28-month interval from November 2017 through February 2020 were analyzed for M. genitalium and Chlamydia trachomatis positivity rates and prevalence. Detection of these sexually-transmitted pathogens occurred by commercial RNA amplification testing. Chart review was focused on participant outcomes following initial M. genitalium detection and therapeutic intervention.Results: C. trachomatis positivity and prevalence rates were 7.05% and 9.00%, respectively, while analogous rates for M. genitalium were 7.05% and 6.51%, respectively. An average of 1.83 positive results was generated from participants infected with M. genitalium at any time, with an average of 1.17 positive results for C. trachomatis (P < 0.0002). For students treated with azithromycin, 30.3% generated a negative M. genitalium result upon follow-up, with 1g daily and 2-day 500mg dosing regimens demonstrating less efficacy than a 4-day 250mg regimen or moxifloxacin.Conclusion: Data indicate a need for molecular M. genitalium macrolide resistance determination from primary specimens in the university setting.

An update on novel taxa and revised taxonomic status of bacteria isolated from non-domestic animals described in 2022.

Numbers of new and revised microbial taxa are continuously expanding, and the rapid accumulation of novel bacterial species is challenging to keep up with in the best of circumstances. With that in mind, following the template of reports on prokaryotic species isolated from humans, this is now the second publication summarizing new and revised taxa in non-domestic animal species in the Journal of Clinical Microbiology. The majority of new taxa were obtained as part of programs to identify bacteria from mucosal surfaces and the gastrointestinal tract from healthy wildlife. A few notable bacteria included new Erysipelothrix spp. from mammalian and aquatic sources and a novel Bartonella spp. isolated from a rodent, both of which could be considered members of emerging and re-emerging genera with pathogenic potential in humans and animals.

Valid and accepted novel bacterial taxa derived from human clinical specimens and taxonomic revisions published in 2022.

Although some nomenclature changes have caused consternation among clinical microbiologists, the discovery of novel taxa and improving classification of existing groups of organisms is exciting and adds to our understanding of microbial pathogenesis. In this mini-review, we present an in-depth summary of novel taxonomic designations and revisions to prokaryotic taxonomy that were published in 2022. Henceforth, these bacteriology taxonomic summaries will appear annually. Several of the novel Gram-positive organisms have been associated with disease, namely, the Corynebacterium kroppenstedtii-like organisms Corynebacterium parakroppenstedtii sp. nov. and Corynebacterium pseudokroppenstedtii sp. nov. A newly described Streptococcus species, Streptococcus toyakuensis sp. nov., is noteworthy for exhibiting multi-drug resistance. Among the novel Gram-negative pathogens, Vibrio paracholerae sp. nov. stands out as an organism associated with diarrhea and sepsis and has probably been co-circulating with pandemic Vibrio cholerae for decades. Many new anaerobic organisms have been described in this past year largely from genetic assessments of gastrointestinal microbiome collections. With respect to revised taxa, as discussed in previous reviews, the genus Bacillus continues to undergo further division into additional genera and reassignment of existing species into them. Reassignment of two subspecies of Fusobacterium nucleatum to species designations (Fusobacterium animalis sp. nov. and Fusobacterium vincentii sp. nov.) is also noteworthy. As was typical of previous reviews, literature updates for selected clinically relevant organisms discovered between 2017 and 2021 have been included.

Update on novel validly published taxa of bacteria isolated from domestic animals described in 2022.

Expansion of our knowledge of the microbial world continues to progress at a rapid rate and carries with it an associated need for recognizing and understanding the implications of those changes. Here, we describe additions of novel taxa from domestic animals published in 2022 that are validly published per the International Code of Nomenclature of Prokaryotes. These included new members of Staphylococcaceae, Moraxella nasovis sp. nov. in sheep with respiratory disease, three additions to Campylobacteraceae (including one from chickens with spotty liver disease), and multiple additions of organisms from the microbiota of dogs, pigs, and especially honeybees and other important pollinators. Noteworthy additions were associated with diseases of cattle, including mastitis, endocarditis, orchitis, and endometritis. Also described in 2022 was Pseudochrobactrum algeriense sp. nov., a member of the Brucellaceae family, isolated from the mammary lymph nodes of cows.

Optimization and Clinical Evaluation of an Automated Commercial Analyte-Specific Reagent Assay for Mycoplasmoides genitalium Macrolide Resistance Detection in Primary Clinical Specimens.

With improvement in laboratory diagnosis of Mycoplasmoides genitalium infection through molecular diagnostics, macrolide resistance determination within M. genitalium-positive patients is necessary. In this study, we report baseline parameters for an analyte-specific reagent (ASR) macrolide resistance real-time reverse transcriptase PCR on an open access analyzer and evaluated detection of macrolide resistance-mediated mutation (MRM) within 23S rRNA in a clinical specimen set. Initial use of 1.2 µM M. genitalium primer and 0.8 µM M. genitalium detection probe concentrations yielded an 80% false-positive detection rate when challenged with 10,000 copies of wild-type RNA. Optimization experiments showed that lowering primer/detection probe and MgCl2 concentrations minimized these false-detections of wild-type 23S rRNA, while higher levels of KCl increased rates of MRM detection with concomitant lower cycle threshold values and higher fluorescence emission. Lower limit of A2058G mutation detection was 5000 copies/mL (180 copies/reaction; 20/20 detections). Utilization of a baseline correction slope limit of 250 units further mitigated false-detection from wild-type 23S rRNA at challenges up to 3.3 billion copies/mL. MRM was detected in 583/866 (67.3%) clinical specimens initially positive for M. genitalium by commercial transcription-mediated amplification. These data included 392/564 detections (69.5%) from M. genitalium-positive swab specimens and 191/302 (63.2%) from M. genitalium-positive-positive first-void urine specimens (P = 0.06). Overall resistance detection rates did not vary by gender (P = 0.76). Specificity of the M. genitalium macrolide resistance ASR was 100% (141 urogenital determinations). MRM detection by the ASR was confirmed at a concordance rate of 90.9% by Sanger sequencing of a clinical specimen subset.

Point-Counterpoint: What's in a Name? Clinical Microbiology Laboratories Should Use Nomenclature Based on Current Taxonomy.

INTRODUCTION The mnemonic SPICE (Serratia, Pseudomonas, indole-positive Proteus, Citrobacter, and Enterobacter) has served as a reminder to consider when a Gram-negative organism may carry a chromosomal copy of blaampC, with the associated risk of developing resistance to first-, second-, and third-generation cephalosporins. However, in 2017, there was a well-founded proposal to rename Enterobacter aerogenes to Klebsiella aerogenes, based on whole-genome sequencing (WGS), and the SPICE mnemonic lost its relevance. With the increased use of WGS for taxonomy, it seems like bacteria and fungi are undergoing constant name changes. These changes create unique challenges for clinical microbiology laboratories, who would like to issue reports that are readily understood and that help clinicians determine empirical antibiotic therapy, interpret antimicrobial resistance, and understand clinical significance. In this Point-Counterpoint, Drs. Karen Carroll and Erik Munson discuss the pros of updating bacterial taxonomy and why clinical labs must continue to update reporting, while Drs. Susan Butler-Wu and Sheila Patrick argue for caution in adopting new names for microorganisms.

An Update on Novel Taxa and Revised Taxonomic Status of Bacteria (Including Members of the Phylum Planctomycetota) Isolated from Aquatic Host Species Described in 2018 to 2021.

Increased interest in farmed aquatic species, aquatic conservation measures, and microbial metabolic end-product utilization have translated into a need for awareness and recognition of novel microbial species and revisions to bacterial taxonomy. Because this need has largely been unmet, through a 4-year literature review, we present lists of novel and revised bacterial species (including members of the phylum Planctomycetota) derived from aquatic hosts that can serve as a baseline for future biennial summaries of taxonomic revisions in this field. Most new and revised taxa were noted within oxidase-positive and/or nonglucose fermentative Gram-negative bacilli, including members of the Tenacibaculum, Flavobacterium, and Vibrio genera. Valid and effectively published novel members of the Streptococcus, Erysipelothrix, and Photobacterium genera are additionally described from disease pathogenesis perspectives.

Antimicrobial Activity of Tebipenem and Comparators against Enterobacterales from diverse Outpatient Centers and Nursing Homes in the United States.

INTRODUCTION: Tebipenem is a potential option for the treatment of a range of infections because of its oral dosing coupled with the safety profile of the ß-lactam antimicrobial class. OBJECTIVES: To evaluate tebipenem in vitro activity against a challenge set of clinical Enterobacterales collected from outpatient and community settings. METHODS: 618 Enterobacterales isolates were submitted by 11 geographically dispersed U.S medical centers that processed cultures from affiliated outpatient centers in 2022. Susceptibility tests for tebipenem and comparator agents were performed by broth microdilution. Extended-spectrum-ß-lactamase (ESBL)-like isolates were identified phenotypically. Multidrug-resistant isolates were non-susceptible to ≥1 agent in ≥3 antimicrobial classes. Genotypic testing (CarbaR) was conducted on select isolates. RESULTS: Isolates (59% Escherichia coli) were recovered from patients seen predominantly in urology/nephrology (24%), nursing home/long-term care (21%), and ambulatory/primary care (21%) clinics. Comparator agent susceptibility rates against all isolates were as follows: levofloxacin (67.5%), amoxicillin/clavulanate (73.6%), cefixime (70.4%), cefpodoxime (70%), cephalexin (61.7%), ceftriaxone (74.4%), cefazolin (63.8%), ertapenem (97.6%), meropenem (99.7%), nitrofurantoin (64.9%), and sulfamethoxazole/trimethoprim (70.9%). Overall, 90.3% (558/619) of isolates were inhibited at a tebipenem MIC of ≤0.125 mg/L (MIC50/90, 0.016/0.125 mg/L), including 85.7% inhibition of ESBL-phenotype isolates (n=161; MIC50/90, 0.03/0.25 mg/L), 86.3% of levofloxacin and sulfamethoxazole/trimethoprim co-resistant isolates (n=95; MIC50/90, 0.016/0.25 mg/L) and 84.3% of multidrug-resistant isolates (n = 172; MIC50/90, 0.03/0.25 mg/L). Carbapenemase genes were observed in 2 ESBL-phenotype isolates with a tebipenem MIC of ≥0.5 mg/L. CONCLUSION: Relative to common oral comparators, these data demonstrate excellent tebipenem in vitro activity against Enterobacterales isolated from patients receiving care in outpatient settings, including urology clinics and nursing homes.

Incidence and laboratory diagnosis of sexually-transmitted infections among university students in a high-prevalence community.

Objective: The purpose of this study was to determine the incidence of four sexually-transmitted infection (STI) agents among university students. Participants: A total of 1744 Milwaukee, Wisconsin metropolitan area student encounters at two university health clinics. Methods: Following consent, participants completed an eight-item demographic and behavioral questionnaire and provided a specimen for Chlamydia trachomatis, Neisseria gonorrhoeae, Trichomonas vaginalis, and Mycoplasma genitalium RNA testing. Results: C. trachomatis (7.2%) and M. genitalium (6.7%) detection rates were highest, with C. trachomatis more likely to result in symptomatic infection in males and females. Female M. genitalium detection rates did not differ as a function of symptomatic status. Students identifying as African American were more likely to test positive for M. genitalium than other races/ethnicities. M. genitalium detection in both genders was a function of reported condom usage. Conclusions: Students would benefit from familiarity with emerging STI agents, as well as behaviors associated with increased risk of STI.

An Update on Novel Taxa and Revised Taxonomic Status of Bacteria Isolated from Domestic Animals Described in 2018 to 2021.

Novel bacterial taxonomy and nomenclature revisions can have significant impacts on clinical practice, disease epidemiology, and veterinary microbiology laboratory operations. Expansion of research on the microbiota of humans, animals, and insects has significant potential impacts on the taxonomy of organisms of clinical interest. Implications of taxonomic changes may be especially important when considering zoonotic diseases. Here, we address novel taxonomy and nomenclature revisions of veterinary significance. Noteworthy discussion centers around descriptions of novel mastitis pathogens in Streptococcaceae, Staphylococcaceae, and Actinomycetaceae; bovine reproductive tract pathogens in Corynebacteriaceae; novel members of Mannheimia spp., Leptospira spp., and Mycobacterium spp.; the transfer of Ochrobactrum spp. to Brucella spp.; and revisions to the genus Mycoplasma.

Update on Accepted Novel Bacterial Isolates Derived from Human Clinical Specimens and Taxonomic Revisions Published in 2020 and 2021.

A number of factors, including microbiome analyses and the increased utilization of whole-genome sequencing in the clinical microbiology laboratory, has contributed to the explosion of novel prokaryotic species discovery, as well as bacterial taxonomy revision. This review attempts to summarize such changes relative to human clinical specimens that occurred in 2020 and 2021, per primary publication in the International Journal of Systematic and Evolutionary Microbiology or acceptance on Validation Lists published by the International Journal of Systematic and Evolutionary Microbiology. Of particular significance among valid and effectively published taxa within the past 2 years were novel Corynebacterium spp., coagulase-positive staphylococci, Pandoraea spp., and members of family Yersiniaceae. Noteworthy taxonomic revisions include those within the Bacillus and Lactobacillus genera, family Staphylococcaceae (including unifications of subspecies designations to species level taxa), Elizabethkingia spp., and former members of Clostridium spp. and Bacteroides spp. Revisions within the Brucella genus have the potential to cause deleterious effects unless the relevance of such changes is properly communicated by microbiologists to stakeholders in clinical practice, infection prevention, and public health.

Update on Novel Taxa and Revised Taxonomic Status of Bacteria Isolated from Nondomestic Animals Described in 2018 to 2021.

Revisions and new additions to bacterial taxonomy can have a significant widespread impact on clinical practice, infectious disease epidemiology, veterinary microbiology laboratory operations, and wildlife conservation efforts. The expansion of genome sequencing technologies has revolutionized our knowledge of the microbiota of humans, animals, and insects. Here, we address novel taxonomy and nomenclature revisions of veterinary significance that impact bacteria isolated from nondomestic wildlife, with emphasis being placed on bacteria that are associated with disease in their hosts or were isolated from host animal species that are culturally significant, are a target of conservation efforts, or serve as reservoirs for human pathogens.

Changes in Streptococcus pneumoniae Susceptibility in Wisconsin: Implications for Clinical Treatment Decisions for Respiratory Infections.

Objective: In 2019, the American Thoracic Society and Infectious Diseases Society of America updated clinical practice guidelines for community-acquired pneumonia (CAP). In contrast to guidelines published in 2007, macrolide monotherapy for outpatients was made a conditional recommendation based on resistance levels. Local knowledge of current antimicrobial susceptibility is needed to guide management of CAP and other bacterial respiratory pathogens. The purpose of this study was to investigate antimicrobial susceptibility profiles and trending for Wisconsin Streptococcus pneumoniae isolates.Design: Multi-center laboratory surveillance, with testing at a central location utilizing standardized susceptibility testing protocols.Methods: Data published by the Wisconsin Department of Health Services (DHS) were augmented with data from the Surveillance of Wisconsin Organisms for Trends in Antimicrobial Resistance and Epidemiology (SWOTARE) program. Data were stratified by invasive or non-invasive sources, as well as DHS region and compared to data compiled from 2006-2010.Results: Susceptibility rates for ≥ 916 invasive S. pneumoniae assessed from 2016-2020 were greater than 91% for ceftriaxone, tetracycline, and fluoroquinolone agents and were generally higher than those from 354 non-invasive isolates. Low susceptibility rates were observed for invasive isolates of penicillin (78.7%) and erythromycin (64.8%) and were even lower for non-invasive isolates (73.8% and 59.9%, respectively). This erythromycin susceptibility rate was a significant reduction from that observed in 2006-2010 (80.4; P < 0.0002). 24.8% of isolates generated an erythromycin MIC ≥ 8 µg/mL. Statewide geographic variability was noted.Conclusions: Rates of S. pneumoniae susceptibility to parenteral penicillins and cephems, and oral tetracycline and fluoroquinolone agents, remain high throughout Wisconsin. However, low oral penicillin susceptibility rates, taken together with declining macrolide susceptibility rates, should cause clinicians to consider alternative treatment options for respiratory tract infections, especially with macrolides.

Surveillance of Fluoroquinolone Resistance in Wisconsin: Geographic Variation and Impact of Revised CLSI Breakpoints.

Objective: Many clinical microbiology laboratories procure antimicrobial susceptibility testing data using guidelines established by Clinical and Laboratory Standards Institute (CLSI). When necessary, CLSI revises interpretive breakpoints in efforts to improve clinical correlation, with two revisions relative to fluoroquinolone agents occurring in 2019. The purpose of this investigation was to determine the impact of fluoroquinolone breakpoint revisions on Wisconsin clinical isolates of Escherichia coli, Proteus mirabilis, and Pseudomonas aeruginosa.Design: Multi-center laboratory surveillance, with testing at a single location utilizing standardized media and susceptibility testing protocols.Methods: From the Surveillance of Wisconsin Organisms for Trends in Antimicrobial Resistance and Epidemiology (SWOTARE) program, levofloxacin and ciprofloxacin minimum inhibitory concentration (MIC) values for 1911, 1521, and 1463 Wisconsin isolates of E. coli, P. mirabilis, and P. aeruginosa, respectively, were determined by broth microdilution testing. In separate data analyses, all MIC data were interpreted using CLSI breakpoints published prior to 2019, then secondarily by using CLSI breakpoints published since 2019 (which reflect lower breakpoints for both levofloxacin and ciprofloxacin resistance). Findings were further stratified by Wisconsin Department of Health Services region.Results: Up to 3.2% decreased statewide fluoroquinolone susceptibility was observed for E. coli isolates, while 5.1% and 6.3% decreases in levofloxacin susceptibility were noted for P. aeruginosa and P. mirabilis isolates, respectively, when revised breakpoints were applied. E. coli isolates from the Western region and P. mirabilis isolates from the Southeastern region demonstrated significant shifts toward decreased fluoroquinolone susceptibility upon application of revised breakpoints. Northern region P. mirabilis isolates exhibited consistently decreased fluoroquinolone susceptibility.Conclusions: Fluoroquinolone resistance has been underreported in Wisconsin as a whole, yet geographic variability continues to exist. Targeted annual surveillance is important to identify and monitor resistance trending. Compilations of SWOTARE surveillance data can be utilized to predict the impact of future CLSI interpretive breakpoint revisions in Wisconsin.