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Microbiological analysis and whole-genome sequencing of Neisseria gonorrhoeae from the microbiological failures in the international, zoliflodacin, phase 3, clinical trial for treatment of uncomplicated urogenital gonorrhoea: a retrospective, genomic, observational study.

BACKGROUND: Zoliflodacin, a first-in-class oral bacterial, DNA gyrase (GyrB) inhibitor, showed non-inferiority to ceftriaxone combined with azithromycin in a recent large international, phase 3, randomised controlled trial for treatment of uncomplicated urogenital gonorrhoea. The aim of this study was to describe the microbiological and whole-genome sequencing (WGS) analyses of paired baseline (pre-treatment) and test-of-cure (TOC) gonococcal isolates from the zoliflodacin phase 3, randomised controlled trial to further characterise and evaluate the protocol-specified microbiological failures with zoliflodacin (n=22) or ceftriaxone and azithromycin (n=1). METHODS: In this retrospective, genomic, observational study, results from antimicrobial susceptibility testing (agar dilution method) of isolates (n=960; 936 baseline isolates from 763 participants and 24 TOC isolates [23 with a paired baseline isolate in the same anatomical site] from 20 participants) collected during the zoliflodacin phase 3, randomised controlled trial done in 16 outpatient clinics in Belgium, the Netherlands, South Africa, Thailand, and the USA (Nov 6, 2019-March 16, 2023) are described. WGS analysis was performed on paired baseline and TOC isolates from participants with microbiological failures (zoliflodacin 44 isolates [19 participants]; ceftriaxone and azithromycin two isolates [one participant]), and the three baseline isolates with highest zoliflodacin minimum inhibitory concentration (MIC 0·5 mg/L). FINDINGS: All isolates were inhibited by the same zoliflodacin concentrations (MICs ≤0·008 to 0·5 mg/L) as wild-type strains cultured internationally in 2013-23. In participants with a microbiological failure after zoliflodacin treatment (n=22, 19 participants), zoliflodacin MIC values for baseline and TOC isolates were similar, and resistance selection was lacking. WGS showed that five (23%) of 22 infections (95% CI 10-43 [in four participants]) of zoliflodacin microbiological failures had different strains at TOC versus baseline. In 17 zoliflodacin microbiological failures (15 participants), isolates at baseline and TOC were indistinguishable. 13 of these 17 microbiological failures, corresponding to 59% (95% CI 39-77; 13 of 22) of all zoliflodacin microbiological failures, were in urogenital or rectal sites in 11 participants and the isolates had zoliflodacin MICs less than or equal to 0·008 to 0·25 mg/L. The single microbiological failure after ceftriaxone and azithromycin treatment had different strains at TOC versus at baseline. No sequenced isolates had mutations associated with elevated zoliflodacin MICs. INTERPRETATION: In the zoliflodacin phase 3, randomised controlled trial, 23% of the zoliflodacin microbiological failures and the single ceftriaxone and azithromycin microbiological failure had different gonococcal strains at TOC versus baseline, which suggests reinfections and not treatment failures. In addition, 59% of the zoliflodacin microbiological failures, all in anogenital sites, had no obvious microbiological explanation based on the low zoliflodacin MICs, previous pharmacodynamic studies, and no evidence of resistance selection after zoliflodacin therapy. A reinfection as the cause for these microbiological failures could not be excluded. We recommend that WGS is implemented in future randomised controlled trials for gonorrhoea treatment to further evaluate possible microbiological failures, exclude reinfections (to avoid underestimating the cure rates), and characterise antimicrobial resistance determinants. FUNDING: GARDP through grants from Germany BMFTR (03KA1831), UK DHSC as part of GAMRIF, Japan MHLW, the Netherlands' Ministry of Health, Welfare and Sport and Directorate-General for International Cooperation, the Federal Office of Public Health of Switzerland, the Canton of Geneva, Switzerland, and Örebro University Hospital, Sweden.

Humans

[Application of a new method for the calculation and description of the resistance of microbiological indicators. I. Testing of several common microbiological sterilization indicators (author's transl)].

The method described by SPICHER and PETERS (1975) for the calculation and description of the resistance of microbiological indicators was tested. As test objects served spore-containing earth according to DIN 58946, Attest indicators (3 M Company, Minnesota) and Oxoid Spore Strips (Oxoid Ltd., London). The tests were performed not only for different batches of indicators but also for preparations of different age. After application of steam (120 degrees C), the indicators were examined for the presence of surviving germs capable of multiplication. When plotting the frequency of indicators with surviving germs (q) against the duration of steam action, S-shaped curves were obtained as expected. By altering the scale of the ordinate (y = lg (-ln(1 - q))), the S-shaped curves could be transformed into straight lines. Thus, the experimentally established paired values could be used for a calculation of regression. This method of calculation proved to be suitable in all cases studied. By indicating the position and the slope of these straight regression lines, the resistance of microbiological indicators can be exactly described (cf. Table 2). This method is applicable not only to indicators containing culture spores but also for native spore-containing earth. The indicators examined differed in their resistance and stability. Seven out of eight batches of Attest indicators (cf Figs. 1 and 2 and Table 1) fulfilled the requirements of DIN 58946, Part 4, for the resistance of bio-indicators for steam sterilization. One of the batches had a slightly higher resistance. The Attest indicators tested were of good stability (see Fig. 1 and Table 1). Where surviving germs were present on the indicators after treatment by steam, their growth was recognizable, in 99% of cases, already after incubation of the cultures for 24 hours. Only two batches of Oxoid Spore Strips were available for testing. One batch was of a higher resistance than required by DIN 58946. The second batch was slightly above the lower limit of the permissible range (see Fig. 3). During storage for 12 months, the resistance of both batches was reduced by 3--4 min. Where the indicators exhibited surviving germs after treatment by steam, growth was recognizable in 87% of the cases after incubation for 24 hours, while for the other indicators, incubation for 48 hours was necessary. The experiments confirmed the good stability of native spore-containing earth (see Fig. 5). Within 4--5 years, the steam resistance of the preparations decreased only by 3--4 min.

Bacteriological Techniques

[Microbiological Characterization of Exacerbations in Severe Asthma and Their Impact on Therapeutic Decision-Making].

INTRODUCTION: Severe asthma (SA) exacerbations impose a substantial healthcare burden. Microbiological characterization using molecular techniques may improve pathogen identification and contribute to a more individualized therapeutic approach. OBJECTIVE: To characterize the microbiological profile of exacerbations in patients with severe asthma and to analyze the prescription patterns for antibiotics (ATB) and systemic corticosteroids (SC). METHODS: This retrospective observational study was conducted in a Severe Asthma Unit. A total of 103 exacerbations were investigated using conventional microbiological methods and multiplex polymerase chain reaction (FilmArray™) performed on respiratory samples. Bacterial findings were classified according to operational criteria compatible with infection or colonization based on genomic load and culture results. Associations between clinical, microbiological, and therapeutic variables were explored using univariate analyses. RESULTS: Microbiological detection was achieved in 78.6% of exacerbations. Viruses were identified in 59.2% of episodes, with rhinovirus representing the predominant pathogen (62.3% of viral detections). Bacteria were identified in 53.4% of exacerbations (H. influenzae 36,6%), frequently in association with viral coinfection. Bronchiectasis was associated with a higher probability of bacterial detection (OR 2.50; p = 0.031). ATB and SC were prescribed in 61.2% and 44.6% of exacerbations, respectively, with frequent use of combination therapy. No significant differences in overall microbiological detection rates were observed according to biologic therapy status. Considerable microbiological variability was observed across recurrent exacerbations in the same patient. CONCLUSIONS: Microbiological findings were common during severe asthma exacerbations, with respiratory viruses, particularly rhinovirus, being the most frequently identified pathogens. Bronchiectasis was associated with higher rates of bacterial detection and ATB use. The marked variability observed between episodes supports the potential value of individualized microbiological assessment during exacerbations and warrants prospective studies aimed at optimizing therapeutic decision-making.

Biologic therapies.

Computerization in microbiology.

Computerization of the clinical microbiology laboratory is finally coming of age. Operations and functions easily adapted from other clinical laboratories have already been implemented in the microbiology laboratory, e.g., fiscal, clerical, and other administrative housekeeping chores. Similarly, storage, retrieval, and analysis of banks of data easily collected and filed have also been successfully accomplished in microbiology for several years. The more challenging problems of computerization still deserve and require the attention of microbiologists. These problems lie in the smooth sequential formulation and transmission of clinical microbiological test results from the laboratory to clinical records; the manipulation of relevant data by the computer prospectively for the detection and prediction of nosocomial infections or miniepidemics; and the development of programed lessons and examinations for computerized instruction, retraining, and examination of technologists and other individuals interested in microbiology.

Clinical Laboratory Techniques

[Problems of maintaining of hygienic and microbiological quality standards of foodstuffs, particularly of vegetable origin (author's transl)].

As more and more components of our food are nowadays subjected to industrial processing, microbiological controls of raw materials and plant hygiene are important factors in protecting the consumer. In many cases preventive measures and specifications of microbiological characteristics of certain foodstuffs governed by legal regulations do not suffice to guarantee protection of the consumer as desired. For this reason conscientious food manufacturers maintain controls in line with self-imposed and very strict hygienic and microbiological rules. Insufficiencies of the relevant dietetic regulations are exposed and measures to ensure quality recommended. It is shown that, to determine the hygienic and microbiological condition of dehydrated vegetable raw materials, the conditions applying to fresh vegetables must be taken into consideration. Peculiarities pertaining to the microbiological examination of seasonings and methodical problems of determining and judging the spore contents in foods are described.

Bacteriological Techniques

Microbiology Galaxy Lab: The first community-driven gateway for reproducible and FAIR analysis of microbial data.

The explosion of microbial omics data has outpaced the ability of many researchers to analyze it, with complex tools and limited computational resources creating barriers to discovery. To address this gap, we present the Microbiology Galaxy Lab: a free, globally accessible, community-supported platform that combines state-of-the-art analytical power with user-friendly accessibility. Supported by the Galaxy and global microbiology communities, this platform integrates over 315 tool suites and 115 curated workflows, enabling comprehensive metabarcoding, (meta)genomic, (meta)transcriptomic, and (meta)proteomic data analysis within a FAIR-aligned environment. It also supports research in the health and infectious disease sectors, as well as in environmental microbiology. The platform's utility is exemplified through various use cases, including antimicrobial resistance tracking, biomarker prediction, microbiome classification, and functional annotation of key microbes. Built on reproducibility and community engagement, it supports creation, sharing, and updating of best-practice workflows. Over 35 tutorials and learning paths empower scientists, fostering an ecosystem that keeps resources at the forefront of microbial science. The Microbiology Galaxy Lab enables collective analysis, democratising research, thereby accelerating discovery across the global microbiology community (microbiology.usegalaxy.org, .eu, .org.au, .fr).

Journal Article