Monkeypox virus genomic accordion strategies.

The 2023 monkeypox (mpox) epidemic was caused by a subclade IIb descendant of a monkeypox virus (MPXV) lineage traced back to Nigeria in 1971. Person-to-person transmission appears higher than for clade I or subclade IIa MPXV, possibly caused by genomic changes in subclade IIb MPXV. Key genomic changes could occur in the genome's low-complexity regions (LCRs), which are challenging to sequence and are often dismissed as uninformative. Here, using a combination of highly sensitive techniques, we determine a high-quality MPXV genome sequence of a representative of the current epidemic with LCRs resolved at unprecedented accuracy. This reveals significant variation in short tandem repeats within LCRs. We demonstrate that LCR entropy in the MPXV genome is significantly higher than that of single-nucleotide polymorphisms (SNPs) and that LCRs are not randomly distributed. In silico analyses indicate that expression, translation, stability, or function of MPXV orthologous poxvirus genes (OPGs), including OPG153, OPG204, and OPG208, could be affected in a manner consistent with the established "genomic accordion" evolutionary strategies of orthopoxviruses. We posit that genomic studies focusing on phenotypic MPXV differences should consider LCR variability.

Prevalence of fosA3 Gene in Fosfomycin-Resistant Proteus mirabilis in the Autonomous Community of Madrid (Spain) and Characterization of Two Positive Strains.

Plasmid-mediated resistance to fosfomycin has been seldom described in Proteus mirabilis. We report two strains harboring fosA3 gene. Whole-genome sequencing revealed a plasmid that encoded fosA3 gene flanked by two insertion sequence (IS)26 mobile elements. Both strains also produced the blaCTX-M-65 gene that was located in the same plasmid. The sequence detected was IS1182-blaCTX-M-65-orf1-orf2-IS26-IS26-fosA3-orf1-orf2-orf3-IS26. The importance of this transposon lies in its ability to spread in Enterobacterales, therefore, epidemiological surveillance should be carried out.

In vitro activity of delafloxacin against highly levofloxacin-resistant invasive isolates of Streptococcus pneumoniae.

INTRODUCTION: We report the activity of delafloxacin, a new fluoroquinolone with high affinity for both topoisomerase IV and DNA gyrase, against highly-levofloxacin-resistant invasive strains of Streptococcus pneumoniae. METHODS: A total of 173 highly-levofloxacin-resistant (MIC>32mg/L) S. pneumoniae invasive isolates were studied. The strains were isolated from blood (n=162) and other sterile fluids (n=11). Serotyping was performed by the Pneumotest-Latex and Quellung reaction. Delafloxacin, levofloxacin, penicillin, cefotaxime, erythromycin and vancomycin MICs were determined by the gradient diffusion method following EUCAST guidelines and breakpoints. RESULTS: Among the isolates, 32.9% were penicillin non-susceptible, 19.7% cefotaxime non-susceptible, and 76.9% erythromycin resistant. All were susceptible to vancomycin. Delafloxacin MIC50 and MIC90 (mg/L) values were 0.064 and 0.12, respectively; 60% (15/25) of serotype 9V isolates showed delafloxacin MICs≥0.12mg/L. CONCLUSIONS: Delafloxacin was very active against highly-levofloxacin-resistant invasive isolates of S. pneumoniae. Isolates belonging to serotype 9V showed higher delafloxacin MIC values.

In vitro activity of delafloxacin against highly levofloxacin-resistant invasive isolates of Streptococcus pneumoniae / Actividad in vitro de delafloxacino frente a aislados invasivos de Streptococcus pneumoniae altamente resistentes a levofloxacino

IntroductionWe report the activity of delafloxacin, a new fluoroquinolone with high affinity for both topoisomerase IV and DNA gyrase, against highly-levofloxacin-resistant invasive strains of Streptococcus pneumoniae.MethodsA total of 173 highly-levofloxacin-resistant (MIC>32mg/L) S. pneumoniae invasive isolates were studied. The strains were isolated from blood (n=162) and other sterile fluids (n=11). Serotyping was performed by the Pneumotest-Latex and Quellung reaction.Delafloxacin, levofloxacin, penicillin, cefotaxime, erythromycin and vancomycin MICs were determined by the gradient diffusion method following EUCAST guidelines and breakpoints.ResultsAmong the isolates, 32.9% were penicillin non-susceptible, 19.7% cefotaxime non-susceptible, and 76.9% erythromycin resistant. All were susceptible to vancomycin. Delafloxacin MIC50 and MIC90 (mg/L) values were 0.064 and 0.12, respectively; 60% (15/25) of serotype 9V isolates showed delafloxacin MICs≥0.12mg/L.ConclusionsDelafloxacin was very active against highly-levofloxacin-resistant invasive isolates of S. pneumoniae. Isolates belonging to serotype 9V showed higher delafloxacin MIC values.

IntroducciónAnalizamos la actividad de delafloxacino, una fluoroquinolona con elevada afinidad por la topoisomerasa i.v. y la ADN girasa, frente a cepas invasivas de Streptococcus pneumoniae altamente resistentes a levofloxacino.MétodosSe estudiaron 173 cepas con CMI de levofloxacino >32mg/l procedentes de sangre (n=162) y otros fluidos estériles (n=11). El serotipado se realizó mediante Pneumotest-Latex y test de Quellung.Las CMI de delafloxacino, levofloxacino, penicilina, cefotaxima, eritromicina y vancomicina se determinaron mediante gradiente de difusión siguiendo las recomendaciones del EUCAST.ResultadosEl 32,9% de las cepas fueron no-sensibles a penicilina, el 19,7% no-sensibles a cefotaxima y el 76,9% resistentes a eritromicina. Todas fueron sensibles a vancomicina. Las CMI50 y CMI90 de delafloxacino (mg/l) fueron 0,064 y 0,120, respectivamente; 60% (15/25) de cepas del serotipo 9V presentaron CMI de delafloxacino ≥0,12mg/l.ConclusionesDelafloxacino fue muy activo frente a S. pneumoniae altamente resistente a levofloxacino. Los aislados del serotipo 9V presentaron mayores valores de CMI de delafloxacino.

Prevalence, detection and characterisation of fosfomycin-resistant Escherichia coli strains carrying fosA genes in Community of Madrid, Spain.

OBJECTIVES: The aim of this this study was to describe the presence of different variants of the fosA gene in fosfomycin-resistant Escherichia coli strains in Madrid, Spain. METHODS: fos genes were searched for in 55 E. coli strains collected from seven representative hospitals located in Madrid. A phenotypic screening test was performed following the disk diffusion method with sodium phosphonoformate added as described by Nakamura et al. Additionally, a molecular study based on PCR was used to confirm the screening results. Positive strains for fos genes were further subjected to whole-genome sequencing (WGS). RESULTS: Phenotypic screening was positive in 9/55 strains (16.4%), although genotypic detection was positive in only 3 (fosA3, fosA4 and fosA6). Thus, the prevalence of fos genes in Madrid was 5.5% (3/55). WGS data were not available for the fosA6-positive strain. One isolate with fosA3 (ST69) carried a blaCTX-M-55 gene and seven virulence genes (air, eilA, iha, iss, lpfA, sat and senB). The fosA4-positive isolate (ST4038) carried the virulence genes iss, lpfA, iroN and mchF. Both fos genes were located between two IS26 mobile elements of a plasmid. CONCLUSION: We detected the presence of different variants of plasmid-mediated fosA genes in fosfomycin-resistant E. coli strains in Madrid, Spain. Despite the few reports in Europe, it would be of interest to monitor the spread of these acquired resistance genes.

Rapid identification of bacteria directly from positive blood cultures by a modified method using a serum separator tube and matrix-assisted laser desorption ionization - time of flight MS.

Introduction. Several studies have used matrix-assisted laser desorption ionization-time of flight MS (MALDI-TOF) with a serum separator tube (SST) to perform rapid identification of microorganisms directly from positive blood cultures (BCs), with different performances and methodologies.Hypothesis / Gap Statement. The use of TSS could significantly reduce the time of identification of microorganisms that produce bacteremia.Aim. Our goals were to evaluate bacterial identification by MALDI-TOF using a method based on an SST and compare it with MALDI-TOF after subculture for 18-24 h.Methodology. BCs no more than 1 h after a positive growth signal were included in the study. Analysis of results was expressed as a score. Information about time to a positive signal and number of microorganisms was collected.Results. In total, 253 BCs were analysed; 45.5 % gave a reliable result, 23.3 % an unreliable result and 31.2 % an error in identification. In gram-negative and gram-positive bacteria, the percentages of reliable results were 83.5 and 21.8 %, respectively. According to time to positive signal, the percentages of correct identification and mean score were 81.1 % (99/122) and 1.89±0.30 in Group 1 (<15 h); and 57.2 % (75/131) and 1.70±0.32 in Group 2 (>15 h), respectively (P <0.001). According to the number of microorganisms, the corresponding percentages of correct identification and mean scores were: Group 1 [≤50 microorganisms observed per field (MOF)], 50/94 (53.19 %) and 1.72±0.32; Group 2 (51-100 MOF): 44/66 (66.67 %) and 1.85±0.34; Group 3 (>100 MOF): 79/93 (84.94 %) and 1.84±0.31.Conclusion. This method allowed us to obtain a high percentage of the aetiological agent of bacteraemia in less than 30 min after a positive BC.

In vitro activity of delafloxacin against highly levofloxacin-resistant invasive isolates of Streptococcus pneumoniae.

INTRODUCTION: We report the activity of delafloxacin, a new fluoroquinolone with high affinity for both topoisomerase IV and DNA gyrase, against highly-levofloxacin-resistant invasive strains of Streptococcus pneumoniae. METHODS: A total of 173 highly-levofloxacin-resistant (MIC>32mg/L) S. pneumoniae invasive isolates were studied. The strains were isolated from blood (n=162) and other sterile fluids (n=11). Serotyping was performed by the Pneumotest-Latex and Quellung reaction. Delafloxacin, levofloxacin, penicillin, cefotaxime, erythromycin and vancomycin MICs were determined by the gradient diffusion method following EUCAST guidelines and breakpoints. RESULTS: Among the isolates, 32.9% were penicillin non-susceptible, 19.7% cefotaxime non-susceptible, and 76.9% erythromycin resistant. All were susceptible to vancomycin. Delafloxacin MIC50 and MIC90 (mg/L) values were 0.064 and 0.12, respectively; 60% (15/25) of serotype 9V isolates showed delafloxacin MICs≥0.12mg/L. CONCLUSIONS: Delafloxacin was very active against highly-levofloxacin-resistant invasive isolates of S. pneumoniae. Isolates belonging to serotype 9V showed higher delafloxacin MIC values.

Escherichia coli resistant to fosfomycin from urinary tract infections: Detection of the fosA3 gene in Spain.

OBJECTIVES: The aim of this study was to report the epidemiological and genetic background of fosfomycin-resistant Escherichia coli isolates causing urinary tract infections (UTIs) in Spain. METHODS: A retrospective observational study of 39 randomly selected fosfomycin-resistant E. coli from urine samples collected during 2017 in Getafe (Spain) was performed. Medical records of 39 patients were reviewed. Phylogenetic groups were identified and the pandemic E. coli ST131 and clades thereof were sought by PCR and multilocus sequence typing (MLST). Screening and identification of fos genes and determination of their genetic environment (linkage to IS26) was performed by PCR. RESULTS: Of the 39 E. coli strains, 49% were ESBL-producers. Most of the strains belonged to phylogenetic group B2 (23/39; 59%), and all of these belonged to ST131 but to different clades (20 to clade C and 3 to clade B). Two isolates from phylogenetic group A (both ST10) carried a plasmid-borne fosA3 gene flanked by IS26. Of the 39 patients, 31 were female (mean age 78 years) and 8 were male (mean age 71 years). Moreover, 27 patients (69%) were diagnosed with complicated UTIs and the remaining 12 (31%) had uncomplicated UTIs, and 33 patients (85%) had been previously treated with fosfomycin. CONCLUSION: This study shows that fosfomycin-resistant E. coli strains are mainly isolated from elderly people with complicated UTIs and belong to the pandemic ST131 clone. To our knowledge, here we describe the fosA3 gene for the first time in Spain, which alerts for potential future dissemination that should be monitored.