First large-scale study of antimicrobial susceptibility data, and genetic resistance determinants, in Fusobacterium necrophorum highlighting the importance of continuing focused susceptibility trend surveillance.

OBJECTIVES: The objective of the study was to explore antimicrobial resistance gene determinant, and phenotypic antibiotic susceptibility, data for Fusobacterium necrophorum from a collection of UK strains. Antimicrobial resistance genes detected in publicly available assembled whole genome sequences were investigated for comparison. METHODS: Three hundred and eighty five F. necrophorum strains (1982-2019) were revived from cryovials (Prolab). Subsequent to sequencing (Illumina) and quality checking, 374 whole genomes were available for analysis. Genomes were interrogated, using BioNumerics (bioMérieux; v 8.1), for the presence of known antimicrobial resistance genes (ARGs). Agar dilution susceptibility results for 313 F. necrophorum isolates (2016-2021) were also examined. RESULTS: The phenotypic data for the 313 contemporary strains demonstrated potential resistance to penicillin in three isolates, using EUCAST v 11.0 breakpoints, and 73 (23%) strains using v 13.0 analysis. All strains were susceptible to multiple agents using v 11.0 guidance other than clindamycin (n = 2). Employing v 13.0 breakpoints, metronidazole (n = 3) and meropenem (n = 13) resistance were also detected. The tet(O), tet(M), tet(40), aph(3')-III, ant(6)-la and blaOXA-85 ARGs were present in publicly available genomes. tet(M), tet(32), erm(A) and erm(B) were found within the UK strains, with correspondingly raised clindamycin and tetracycline minimum inhibitory concentrations. CONCLUSIONS: Susceptibility to antibiotics recommended for the treatment of F. necrophorum infections should not be assumed. With evidence of potential ARG transmission from oral bacteria, and the detection of a transposon-mediated beta-lactamase resistance determinant in F. necrophorum, surveillance of both phenotypic and genotypic antimicrobial susceptibility trends must continue, and increase.

UK Bacteroides species surveillance survey: Change in antimicrobial resistance over 16 years (2000-2016).

OBJECTIVES: To assess the differences in antimicrobial susceptibility of UK Bacteroides species across two distinct cohorts from 2000 to 2016. METHODS: Strain identification was performed using matrix-assisted laser-desorption ionisation time of flight mass spectrometry (MALDI-TOF MS) or by partial 16S rRNA sequencing. Minimum inhibitory concentrations (MICs) were determined using agar dilution, following CLSI guidelines (CLSI, 2012; 2017). RESULTS: 224 isolates were included from 2000 to 168 from 2016. Bacteroides fragilis was the most common species, comprising 68% of the 2000 cohort, and 77% in 2016. For all antimicrobials tested, there was an overall increase in the rates of non-susceptible isolates between the cohorts. CONCLUSIONS: The antibiogram of Bacteroides species in the UK is no longer predictable. Multi-drug resistant isolates although rare, are on the rise, and require testing to guide therapy. The monitoring and surveillance of resistance trends is imperative, as is the development of standardised, robust and accessible antimicrobial susceptibility testing methodology for clinical laboratories.

Development of a EUCAST disk diffusion method for the susceptibility testing of rapidly growing anaerobic bacteria using Fastidious Anaerobe Agar (FAA): a development study using Bacteroides species.

OBJECTIVES: Antimicrobial resistance among anaerobic bacteria is increasing, leading to a growing demand for inexpensive and reliable susceptibility testing methods. The aim of this study was to determine the suitability of Fastidious Anaerobe Agar (FAA) as a medium for disk diffusion for rapidly growing anaerobic bacteria. METHODS: Reproducibility of zone diameters and quality of growth were tested using six quality control (QC) strains. We compared four anaerobic incubation systems, two incubation temperatures (35°C and 37°C), and FAA from four manufacturers. The effect of incubation for 16-20 hours instead of 24 hours was tested on ten randomly selected isolates of the Bacteroides fragilis group. The final method was tested on 170 clinical B. fragilis-group isolates and compared to agar dilution MICs. RESULTS: After 24 hours' incubation, all QC strains demonstrated confluent growth. The different anaerobic incubation systems were equal regarding quality of growth and zone diameters. Incubation at 35°C resulted in slightly larger zones (1-2 mm) than at 37°C. Except for Acumedia FAA, the different manufacturers showed good agreement in zone diameters. All B. fragilis-group isolates displayed confluent growth after 16-20 hours. Metronidazole inhibition zones correlated well with the reference MICs. There was an area of poorer separation for meropenem and piperacillin-tazobactam between 19-27 and 14-23 mm respectively. Prolonged incubation (40-44 h) of clindamycin resulted in better separation and the area of overlap was reduced from 13 to 8 mm compared with 16-20 hours' incubation. CONCLUSION: FAA is a suitable medium for disk diffusion of these rapidly growing anaerobic bacteria.

Axon and muscle spindle hyperplasia in the myostatin null mouse.

Germline deletion of the myostatin gene results in hyperplasia and hypertrophy of the tension-generating (extrafusal) fibres in skeletal muscle. As this gene is expressed predominantly in myogenic tissues it offers an excellent model with which to investigate the quantitative relationship between muscle and axonal development. Here we show that skeletal muscle hyperplasia in myostatin null mouse is accompanied by an increase in nerve fibres in major nerves of both the fore- and hindlimbs. We show that axons within these nerves undergo hypertrophy. Furthermore, we provide evidence that the age-related neural atrophic process is delayed in the absence of myostatin. Finally, we show that skeletal muscle hyperplasia in the myostatin null mouse is accompanied by an increase in the number of muscle spindles (also called stretch receptors or proprioceptors). However, our work demonstrates that the mechanisms regulating intrafusal fibre hyperplasia and hypertrophy differ from those that control the aetiology of extrafusal fibres.