Non-tuberculous mycobacteria (NTM) represent an expanding group of environmental microorganisms with growing clinical significance, particularly among immunocompromised patients. In this study, we describe the isolation and comprehensive characterization of a novel Mycobacterium species, strain HUMS_1102779-3, recovered from a sputum sample of a patient undergoing diagnostic bronchoscopy for hilar lymphadenopathy. Initial identification attempts using MALDI-TOF MS and commercial DNA∙STRIP assays were inconclusive, prompting whole-genome sequencing for comprehensive genomic and phenotypic characterization. De novo genome assembly resulted in a 6.2 Mb draft genome with a GC content of 69.03 mol

Cefiderocol is a newly developed siderophore-conjugated cephalosporin specifically designed to overcome multidrug-resistant (MDR) Gram-negative bacilli. Nevertheless, reduced susceptibility and resistance to cefiderocol has increasingly been reported, especially derived from alterations in iron transport systems and plasmid-mediated determinants and the expression of structural variants of β-lactamases. Moreover, cefiderocol resistance derived from target alteration remains insufficiently defined. This work aimed to characterize the genomic and structural basis of cefiderocol resistance emerging in vivo during therapy in Klebsiella pneumoniae. Comparative genomics revealed conserved plasmid architecture and no acquisition of novel β-lactamases or iron-uptake determinants. Eighty-two chromosomal variants were identified, among which a missense substitution in ftsI (FtsI G306V), located adjacent to the catalytic serine (S307) of PBP3, emerged as a plausible resistance determinant. Structural modelling suggests that the G306V substitution introduces a significant side-chain volumetric increase, resulting in locally increased steric hindrance within the active-site, possibly impairing cefiderocol interaction with its target. The absence of canonical iron-transport alterations further distinguishes this case from the predominant mechanism described in global molecular epidemiology studies. Large-scale genomic screening revealed a very low prevalence of substitutions affecting ftsI, including in resistance isolates, although possible clonal expansion cannot be ruled out. Our findings align with the broader epidemiological landscape in which cefiderocol resistance is heterogeneous and flexible. Therefore, incorporating analysis of ftsI variants within or proximal to the active-site into genomic surveillance frameworks could improve resistance detection to this last-resort antibiotic. • Resistance occurred without alterations in iron-uptake systems or siderophore pathways. • Mutations in FtsI (G306V) and AcrR (E91K) were identified as key candidates. • FtsI G306V is rare across cefiderocol resistant and global K. pneumoniae genomes.

The emergence of antimicrobial resistance (AMR) is a global threat, with livestock antibiotic use proposed as a significant contributor. We investigated Streptococcus suis, a multidrug-resistant porcine pathogen, as a potential source of AMR gene dissemination to human-associated streptococci in Aragón, Spain. Among 91 porcine isolates recovered across Spain, erythromycin and tetracycline resistance was linked to erm(B) and tet(O) genes, frequently co-located on Tn5252-family integrative and conjugative elements (ICEs) and integrative and mobilizable elements (IMEs). Tn5252-family ICEs shared 32-100% of their sequence and carried different AMR gene combinations. Screening of 2388 human clinical streptococcal isolates of different species obtained in Aragón revealed ∼20% erythromycin resistance, particularly S. pneumoniae and S. agalactiae. Analysis of erm(B)-tet(O) linkage in a set of erythromycin-resistant isolates and subsequent genome sequencing, revealed twelve isolates carrying Tn5252-family ICEs with both AMR genes. Eight S. agalactiae isolates harbored Tn5252-family ICEs with >95% nucleotide identity across >91% of their sequence compared with two different ICEs identified in S. suis isolates recovered in the same region and period. Conjugation assays confirmed ICE transfer from S. suis to S. agalactiae, S. pneumoniae, and S. pyogenes, while in vitro experiments demonstrated that recombination between ICEs promotes diversification of AMR gene cargo within ICEs. Our results identify S. suis as a reservoir and generator of ICEs carrying distinct AMR gene combinations that can be horizontally transferred to other human pathogenic streptococci, leading to multidrug resistance in a single step. These findings provide strong evidence supporting interspecies AMR dissemination in regions with intensive pig production and underscore the need for integrated One Health strategies combining veterinary and clinical genomic surveillance, and targeted interventions to limit the spread of mobile genetic elements across species.

High-risk human papillomavirus (HR-HPV) and other sexually transmitted infections (STIs-O) are promoters to the development of cervical cancer (CC), especially when they co-exist. This study aims to determine the prevalence of the major STIs-O and the rate of co-infection in women previously diagnosed with HR-HPV infection. For this observational study, 254 women aged 25–65 years who were being followed up for HR-HPV infection (without a CC history) were recruited at a hospital’s Gynaecology Department from February 2024 to November 2024. Their endocervical specimens were collected and processed for HR-HPV, Chlamydia trachomatis, Neisseria gonorrhoeae, Mycoplasma genitalium, and Trichomonas vaginalis detection by RT-PCR using commercially available reagents and equipment. The overall rate of infection was 38.6% for HPV and 4.3% for ITSs-O (3.8% in HPV-negative women and 5.1% in HPV-positive women). The presence of ITSs-O in women aged 25–34 was higher in those with a persistent positive result for HR-HPV (20.0% vs. 4.2%). Diverse multiple co-infections were found in HPV-positive women, whilst some single STIs-O were found in HPV-negative women. These results support the benefits of STI-O screening beyond an HR-HPV positive result, especially in those women under 35 years old.

Candida auris, a yeast that is highly resistant to antifungal treatments, represents a significant healthcare concern due to its rapid spread through surface contamination and patient colonization. A recent outbreak at the Miguel Servet University Hospital in Zaragoza, Spain, highlighted the urgent need for rapid diagnostic responses. The current 4-day duration for microbiological culture results hinders timely patient isolation and transmission prevention. This study aims to evaluate the VIASURE Candida auris Real-Time PCR Detection Kit (Certest Biotec) for diagnostic performance and analytical specificity compared to the standard methodology based on culture and MALDI-TOF MS characterization. From August 2023 to December 2023, the Microbiology Service of the Miguel Servet University Hospital processed 816 diagnostic samples. Accuracy testing resulted in adequate clinical validation values, as presented here: sensitivity, 0.98 (95% confidence interval [CI], 0.90 to 1); specificity, 1 (95% CI, 0.99 to 1); positive predictive value, 0.98 (95% CI, 0.90 to 1); and negative predictive value, 1 (95% CI, 0.99 to 1). The implementation of this molecular biology test in the laboratory promises to improve diagnostic efficiency by allowing rapid patient isolation, initiation of appropriate treatment, and effective isolation measures. IMPORTANCE:Candida auris, a resilient and antifungal-resistant yeast, poses a significant healthcare threat due to its rapid spread. A recent outbreak at a hospital in Zaragoza, Spain, emphasized the urgent need for faster diagnostics. The current four-day wait for culture results hampers timely patient isolation. This retrospective and comparative study demonstrates the favorable clinical parameters of a commercially available molecular diagnostic kit. The kit allows enhanced diagnostic efficiency, swifter patient isolation, and more effective control measures in the laboratory. In conclusion, the kit addresses the pressing challenges presented by C. auris in healthcare.

Giardiasis and cryptosporidiosis, caused by Giardia lamblia and Cryptosporidium spp., are parasitic infections transmitted through faecal–oral routes or contaminated water. Although less common in Spain compared to developing countries, they pose a public health concern, particularly for vulnerable groups like children and immunocompromised individuals. This study aims to analyse the cases reported to the Microbiological Information System (MIS) in children between 2012 and 2021, as well as their distribution across sociodemographic variables. Proportions and infectivity rates were determined for epidemiological and sociodemographic data, and the incidence rate for giardiasis and cryptosporidiosis was calculated annually and by health sector. The variables analysed included sex, age, health sector and weather. For both diseases, there was a significant decrease in the number of cases in 2020, suggesting the importance of person-to-person transmission. Children were infected by Giardia in significantly higher proportion (p < 0.001), being the majority in age groups 5–14 years, while the proportion of boys and girls infected by Cryptosporidium was almost identical (1.4% vs. 1.3%), in children aged 2–4 years. Periodically there was a significant increase in cases of cryptosporidiosis, apparently related to the presence of torrential rains. Transmission is related to increased temperature and rainfall. Person-to-person transmission in the paediatric population needs further investigation. This study provides the foundation for future research on the evolution of cases of giardiasis and cryptosporidiosis in Spanish children. The data emphasise the need for informational campaigns on hygienic measures and efforts by public health authorities to maintain water resources in optimal condition to prevent parasite spread.

This study explores the relationship between specific SARS-CoV-2 mutations and obesity, focusing on how these mutations may influence COVID-19 severity and outcomes in high-BMI individuals. We analyzed 205 viral mutations from a cohort of 675 patients, examining the association of mutations with BMI, hospitalization, and mortality rates. Logistic regression models and statistical analyses were applied to assess the impact of significant mutations on clinical outcomes, including inflammatory markers and antibody levels. Our findings revealed three key mutations—C14599T, A20268G, and C313T—that were associated with elevated BMI. Notably, C14599T appeared to be protective against hospitalization, suggesting context-dependent effects, while A20268G was linked to a 50% increase in hospitalization risk and elevated antibody levels, potentially indicating an adaptive immune response. C313T showed a 428% increase in mortality risk, marking it as a possible poor-prognosis marker. Interestingly, all three mutations were synonymous, suggesting adaptive roles in obesity-driven environments despite not altering viral protein structures. These results emphasize the importance of studying mutations within the broader context of comorbidities, other mutations, and regional factors to enhance our understanding of SARS-CoV-2 adaptation in high-risk groups. Further validation in larger cohorts is necessary to confirm these associations and to assess their clinical significance.

Background/Objectives: Infections caused by multidrug-resistant (MDR)bacteria pose a significant public health threat by worsening patient outcomes, contributing to hospital outbreaks, and increasing health and economic burdens. Advanced genomic tools enhance the detection of resistance genes, virulence factors, and high-risk clones, thus improving the management of MDR infections. In the Autonomous Community of Aragon, the diversity and incidence of carbapenemase-producing Enterobacteriaceae (CPE) have increased during the last years. This study analyses CPE trends at a tertiary hospital in Spain from 2021 to 2023, aiming to optimize personalized medicine. Methods: CPE isolates were the first isolate per patient, year, species, and carbapenemase from January 2021 to December 2023. Additional metadata were collected from the laboratory's information system. Antibiotic susceptibility testing was performed by broth microdilution. Whole-genome sequencing (WGS) was performed using Illumina short reads. De novo assembly was used to generate draft genomes in order to determine their complete taxonomic classification, resistome, plasmidome, sequence type (ST), core-genome multilocus sequence typing (cgMLST), and phylogenetic relationships using a suite of bioinformatics tools and in-house scripts. Results: Between 2021 and 2023, 0.4% out of 38,145 Enterobacteriaceae isolates were CPE. The CPE rate tripled in 2022 and doubled again in 2023. The most common species was Klebsiella pneumoniae (51.8%) and the most common carbapenemase was blaOXA-48. WGS revealed concordant species identification and the carbapenemase distribution in detail. Resistance rates to critical antibiotics, such as carbapenems, were variable, but in most cases were above 70%. Genetic diversity was observed in WGS and phylogenetic analyses, with plasmids often mediating carbapenemase dissemination. Conclusions: The increasing rate of CPE in healthcare settings highlights a critical public health challenge, with limited treatment options. Genomic characterization is essential to understanding resistance mechanisms, aiding therapy, limiting outbreaks, and improving precision medicine.

Background/Objectives: Cefiderocol is a novel siderophore cephalosporin with potent in vitro activity against a broad spectrum of Gram-negative bacteria, including carbapenemase-producing Enterobacterales (CPE). However, the recent emergence of resistance in clinical settings raises important concerns regarding its long-term effectiveness. This study aims to investigate the genomic determinants associated with cefiderocol resistance in CPE isolates of human origin. Methods: Comparative genomic analyses were conducted between cefiderocol-susceptible and -resistant CPE isolates recovered from human clinical and epidemiological samples at a tertiary care hospital. Whole-genome sequencing, variant annotation, structural modelling, and pangenome analysis were performed to characterize resistance mechanisms. Results: A total of 59 isolates (29 resistant and 30 susceptible) were analyzed, predominantly comprising Klebsiella pneumoniae, Escherichia coli, and Enterobacter cloacae. The most frequent carbapenemase gene among the resistant isolates was blaNDM, which was also present in a subset of susceptible strains. The resistant isolates exhibited a significantly higher burden of non-synonymous mutations in their siderophore receptor genes, notably within fecR, fecA, fiu, and cirA. Structural modelling predicted deleterious effects for mutations such as fecR:G104S and fecA:A190T. Additionally, porin loss and loop 3 insertions (e.g., GD/TD) in OmpK36, as well as OmpK35 truncations, were more frequent in the resistant isolates, particularly in high-risk clones such as ST395 and ST512. Genes associated with toxin–antitoxin systems (chpB2, pemI) and a hypothetical metalloprotease (group_2577) were uniquely found in the resistant group. Conclusions: Cefiderocol resistance in CPE appears to be multifactorial. NDM-type metallo-β-lactamases and missense mutations in siderophore uptake systems—especially in those encoded by fec, fhu, and cir operons—play a central role. These may be further potentiated by alterations in membrane permeability, such as porin disruption and efflux deregulation. The integration of genomic and structural approaches provides valuable insights into emerging resistance mechanisms and may support the development of diagnostic tools and therapeutic strategies.

BACKGROUND:Livestock-associated (LA)-MRSA of CC398 lineage is related to the pig environment, although it also colonizes/infects humans. Tetracycline resistance (TETR) is a phenotypic marker of LA-MRSA-CC398. OBJECTIVES:To determine the prevalence and changing epidemiology of LA-MRSA-CC398 in seven Spanish hospitals (H1-H7) located in areas with different pig farming densities (PFDs) (extremely high, very high, medium, and very low: EH/VH/M/VL), and to identify other non-CC398-LA-MRSA clones. METHODS:MRSA-TETR isolates (n = 165) obtained from hospitals H1-H7 over 6 months in 2023 were characterized with respect to genetic lineages/antimicrobial resistance and virulence/immune evasion cluster of CC398 and non-CC398. Results were compared with a previous multicentre study from 2016. RESULTS:We identified 86/165 MRSA-TETR isolates (52.1%) as being MRSA-CC398. A significant difference in MRSA-CC398/MRSA prevalence was detected between hospitals located in EH-PFD areas (H1 and H2,  > 25%) and the one in a VL-PFD area (H7, 0%). Prevalences in the range 6.4%-12.2% were found in hospitals in M- and VH-PFD areas (H3-H6). Fourteen spa-types were identified among MRSA-CC398 isolates, with t011/t034 predominating (68.3%), followed by t1451/t1255 (13%). All but two MRSA-CC398 isolates were scn-negative (human adaptation marker). Among the 79 non-CC398-MRSA-TETR isolates, CC5, CC1 and CC8 predominated (45.6%, scn-negative). The prevalences of the scn gene among non-CC398-MRSA-TETR isolates in hospitals of EH-, VH-, M- and VL-PFD areas were 77.8%, 50%, 18.75% and 0%, respectively (significant correlation, P < 0.05). This study shows that MRSA-CC398 isolates are prevalent in hospitals located in EH-PFD areas but absent in the hospital of the VL-PFD area. Moreover, an increase in both PFD and prevalence of MRSA-CC398/MRSA in hospitals H3-H6 was observed in the 2023 study compared with the 2016 study. CONCLUSIONS:A significant increase and more genetic diversity of predominant lineages of CC398-MRSA-TETR were observed in hospitals located in M- to VH-PFD areas.

Background: Staphylococcus aureus is one of the most prevalent bacteria in skin and soft tissue infections (SSTIs). Multidrug-resistant strain emergence, particularly methicillin-resistant S. aureus (MRSA), highlights the need for alternative treatments. Objectives: This study investigates the antimicrobial properties of olive leaf extract (OLE) and describes an epidemiological profiling of patients with SSTI who may benefit from it. Methods: OLE was tested in two reference strains, methicillin-susceptible S. aureus (MSSA) ATCC 29213 and MRSA ATCC 700699, and in 126 clinical isolates from patients with SSTIs according to Clinical Laboratory Standards Institute guidelines. Results: The minimum bactericidal concentration (MBC) ranged from 3.12% to 6.25% w/v for MSSA and 1.56% to 3.12% for MRSA. The lethal curve showed a reduction of 6 log10CFU/mL after two hours of incubation. Most of the 126 clinical samples (103 MSSA and 23 MRSA) came from skin lesions, surgical wounds, and ulcers. Over 90% of MSSA strains were resistant to less than five antibiotics, while 82% of MRSA strains were resistant to more than six. Penicillins demonstrated the lowest susceptibility rate (19.8%), whereas linezolid, daptomycin, pristinamycin, trimethoprim–sulfamethoxazole, teicoplanin, vancomycin, and OLE exhibited 100% susceptibility. No growth was observed for all clinical strains with OLE at ≥6.25% w/v. Conclusions: The findings suggest that OLE could become a promising alternative treatment for skin infections, particularly in the context of increasing antibiotic resistance.

The genus Aeromonas comprises Gram-negative bacteria widely distributed in aquatic environments, with some species able to cause disease in humans, fish, and other aquatic animals. The dominating species in human infections are Aeromonas caviae, Aeromonas dhakensis, Aeromonas hydrophila, and Aeromonas veronii and the disease presentations gastroenteritis, bacteremia, and wound infections. Matrix-Assisted Laser Desorption Ionization Time of Flight Mass Spectrometry (MALDI-TOF MS) is an extremely rapid method used in clinical microbiology laboratories to identify bacterial isolates at the genus and species level. The present study aimed to evaluate the usefulness of the MALDI-TOF MS to identify 273 clinical isolates of Aeromonas that were also identified by rpoD gene. The latter recognized eight different species, but only 73.6% of the strains of six species were correctly identified with MALDI-TOF MS and results depended upon the species. The higher concordance was with A. veronii (92.8%), A. hydrophila (83.3%), A. caviae (73.1%), and Aeromonas media (60.0%). Our results and those of the reviewed literature corroborate that MALDI-TOF is a promising identifying method being the poorly updated database the main limitation. Improvement requires including a higher diversity of strains from all the described species which should be the responsibility of the provider.

Healthcare services and institutions are focused on providing the most appropriate medical service in terms of patient safety and satisfaction outcomes. According to Lean methodologies, effectiveness and efficiency can be improved by assuring value-added processes. This article presents a joint approach for the development and implementation of Lean techniques combined with Total Laboratory Automation (TLA) for serology diagnosis in a microbiology laboratory in a tertiary-level hospital. The results obtained show an improvement in the process efficiency and its key performance indicators. In particular, for the HIV and COVID tests, the process Turnaround Times (TAT) were decreased by up to 87.3% and 19.3%, having a direct effect in the diagnostic response time. The process added-value for HIV tests increased by 81%. This meant a cost reduction per test, a higher number of diagnostic tests and clinical samples processed and laboratory resource optimisation. The implementation of TLA also enabled the reallocation of skilled labour towards value-added tasks, increased the process quality and reduced sample waiting times. This work opens up new opportunities for their deployment in other laboratory areas and sample types, directly influencing the overall quality of patient diagnosis in the context of tertiary healthcare facilities.

Background/Objectives: Multidrug-resistant (MDR) infections pose a significant public health threat by worsening patient outcomes, contributing to hospital outbreaks, and increasing the health and economic burden. Advanced genomic tools enhance the detection of resistance genes, virulence factors, and high-risk clones, thus improving the management of MDR infections. In the Autonomous Community of Aragon, the carbapenemase-producing Enterobacteriaceae (CPE) diversity and incidence have increased during the last years. This study analyses CPE trends at a tertiary hospital in Spain from 2021-2023, aiming to optimize personalised medicine. Methods: CPE isolates were the first isolate per patient, year, species, and carbapenemase from January 2021 to December 2023. Antibiotic susceptibility testing was performed by broth microdilution method. Whole genome sequencing (WGS) was performed using Illumina. The genomes were thoroughly analysed to determine their complete taxonomic classification, resistoma and plasmidome composition, and sequence typing using a suite of bioinformatics approaches. Results: Between 2021 and 2023, 0.4% of all isolates were CPE. The CPE rate tripled in 2022 and doubled again in 2023. The most common species was Klebsiella pneumoniae (51.8%) and the most common carbapenemase was blaoxa-48. WGS revealed concordant species identification and detailed carbapenemase distribution. Resistance rates to critical antibiotics such as carbapenems were variable but in most cases above 70%. Genetic diversity was observed in WGS and phylogenetic analyses, with plasmids often mediating carbapenemase dissemination. Conclusions: The increasing rate of CPE in healthcare settings highlights a critical public health challenge, with limited treatment options. Genomic characterization is essential to understand resistance mechanisms, aid therapy, limit outbreaks and improve precision medicine.

Mediterraneibacter gnavus is a Gram positive, non-sporulated, obligate anaerobe diplococci. It was first described in 1974 by Moore et al. (under the name Ruminococcus gnavus) from faeces and contents of the gastrointestinal tract of humans. It is a relatively common member of the human gut microbiota, nevertheless its role as a pathogenic bacterium has not been completely elucidated yet and it seems to depend on numerous factors, including those of the host. Here we present a case of prosthetic joint infection following total knee arthroplasty by M. gnavus. A 74 years old patient was admitted to the emergency department presenting with acute onset of left knee pain and swelling 20 days after total left knee arthroplasty. Follow-up revealed erythema and oedema without signs of fluctuation or purulent discharge from the surgical wound and elevated inflammatory reactants. Synovial fluid was taken for bacterial culture and antibiotic treatment with ceftazidime and daptomycin was established. Examination of the synovial fluid revealed abundant polymorphonuclear leucocytes, without visualizing bacteria. After four days of incubation, anaerobic culture exhibit growth of small, grey, umbilicated colonies in pure culture on Schaedler agar. The microorganism was identified as R. gnavus by MALDI-TOF (Bruker Daltonics) and M. gnavus by 16S ribosomal bacterial sequencing. The isolated showed susceptibility to the most commonly used anaerobicidal antibiotics except for clindamycin. Surgical treatment and infection source control included DAIR (debridement, antibiotics, and implant retention) and vacuum assisted therapy. The patient was discharged after six weeks with a 3-month course of oral amoxicillin as consolidation therapy. Subsequent follow-up revealed adequate wound healing with no signs of infection. Mediterraneibacter gnavus have been reported as the causal microorganism in a range of human infections, nevertheless its identification remains challenging. Infection of prosthetic joints by anaerobic microorganisms is uncommon and is not considered in its empirical antibiotic treatment, thus, correct and swift identification of anaerobic bacteria in these cases is paramount.

In the frame of the I-MOVE-COVID-19 project, a cohort of 2050 patients admitted in two Spanish reference hospitals between March 2020 and December 2021 was selected and a range of clinical factor data were collected at admission to assess their impact on the risk COVID-19 severity outcomes through a multivariate adjusted analysis and nomograms. The need for ventilation and intensive care unit (ICU) admission were found to be directly associated with a higher death risk (OR 6.9 and 3.2, respectively). The clinical predictors of death were the need for ventilation and ICU, advanced age, neuromuscular disorders, thrombocytopenia, hypoalbuminemia, dementia, cancer, elevated creatin phosphokinase (CPK), and neutrophilia (OR between 1.8 and 3.5), whilst the presence of vomiting, sore throat, and cough diminished the risk of death (OR 0.5, 0.2, and 0.1, respectively). Admission to ICU was predicted by the need for ventilation, abdominal pain, and elevated lactate dehydrogenase (LDH) (OR 371.0, 3.6, and 2.2, respectively) as risk factors; otherwise, it was prevented by advanced age (OR 0.5). In turn, the need for ventilation was predicted by low oxygen saturation, elevated LDH and CPK, diabetes, neutrophilia, obesity, and elevated GGT (OR between 1.7 and 5.2), whilst it was prevented by hypertension (OR 0.5). These findings could enhance patient management and strategic interventions to combat COVID-19.

Background and Objectives: Neisseria gonorrhoeae and Chlamydia trachomatis can cause similar clinical syndromes and may coexist in infections. In emergency medicine, empirical treatment targeting both pathogens is often employed, potentially contributing to antibiotic resistance. Gonococcal resistance has emerged against first-line antimicrobials, necessitating prior testing for fluoroquinolone susceptibility. Certest Biotec developed two molecular diagnostic products for simultaneous detection: VIASURE C. trachomatis & N. gonorrhoeae Real-Time PCR Detection Kit and VIASURE Neisseria gonorrhoeae Ciprofloxacin-Resistant Real-Time PCR Detection Kit. To evaluate these products, clinical performance assessments were conducted at the Clinical Microbiology Laboratory of Miguel Servet University Hospital in Zaragoza, Spain. Results and Conclusions: Both VIASURE assays under study demonstrated high clinical sensitivity and specificity compared to reference molecular assays and Sanger sequencing. These kits offer an accurate diagnosis, facilitating appropriate treatment choices while addressing concerns about emerging antibiotic resistance. Methods: A total of 540 clinical samples from 540 patients already characterized as positive or negative for CT and NG by a molecular assay and by antibiotic susceptibility testing for ciprofloxacin using a concentration gradient diffusion method were used for the clinical evaluation. In cases where sensitivity results were unavailable, conventional PCR and Sanger sequencing were employed.

Purpose: Staphylococcus aureus is one of the most prevalent pathogens in cystic fibrosis (CF), being of special relevance those methicillin-resistant (MRSA). The livestock-associated (LA)-MRSA lineage CC398 is an emerging problem, specially related to pig-farming (PF) environments. The objective was to characterize the S. aureus isolates recovered from CF-patients in a Spanish hospital located in a region with high-PF activity. Methods: Forty-two isolates were obtained (January-October/2022) and characterised (one/patient). The antimicrobial resistance phenotype/genotype was evaluated by Microscan/PCR. The presence of virulence and Immune Evasion Cluster (IEC) genes as well as the agr type was determined by PCR. MLST and spa-typing were studied by PCR-sequencing. Results: Nine of the 42 isolates were MRSA (21.4 %), and 8 of them multidrug resistant (MDR). Among MRSA, 6 spa-types were detected, assigned to CC1, CC5, CC8, CC30, and CC398. Four MRSA isolates belonged to the lineage CC398-t011-IEC negative (animal adapted-clade, LA-MRSA). The remaining 33 isolates were methicillinsusceptible (MSSA), of 26 spa-types and associated with 11 CCs (predominant: CC5, CC30, and CC398). Seven MSSA isolates were of the lineage CC398 ( spa-types t034, t108, t571, t20352); four of them were IEC-positive and erm (T)-positive (t571, and t20352, human-adapted CC398 clade), being IEC-negative the remaining three. The tst and eta/etb genes were identified in 12 and 2 isolates, respectively (none CC398). Small-colony-variants were demonstrated in 9 isolates (two CC398, both MDR). Conclusion: The lineage CC398 was very frequent among CF-patients (26.2 %), both among MSSA and MRSA. The emergence of LA-MRSA-CC398 in CF-patients requires monitorization, especially in hospitals of high-PF-regions.

The genus Aspergillus consists of a vast number of medically and environmentally relevant species. Aspergillus species classified in series Versicolores are ubiquitous in the environment and include the opportunistic pathogen Aspergillus sydowii, which is associated with onychomycosis and superficial skin infections. Despite frequent clinical reports of A. sydowii and related series Versicolores species, antifungal susceptibility data are scarce, hampering optimal treatment choices and subsequent patient outcomes. Here, we employed antifungal susceptibility testing (AFST) based on microbroth dilution on a set of 155 series Versicolores strains using the common antifungals amphotericin B, itraconazole, voriconazole, posaconazole, isavuconazole and micafungin with the addition of luliconazole and olorofim. All strains were identified using partial calmodulin gene sequencing, with 145 being A. sydowii, seven A. creber and three A. versicolor, using the latest taxonomic insights. Overall, tested antifungals were potent against the entire strain collection. In comparison to A. fumigatus, azole and amphotericin B MICs were slightly elevated for some strains. AFST with luliconazole and olorofim, here reported for the first time, displayed the highest in vitro activity, making these antifungals interesting alternative drugs but clinical studies are warranted for future therapeutic use.

This paper targets the application of Lean Six Sigma (LSS) framework to microbiological diagnosis processes performed in a clinical microbiology laboratory of a tertiary level Spanish hospital.The increase in clinical samples and diagnostic requests during the COVID-19 pandemic led many microbiology laboratories to optimize their processes. Blood culture samples are one of the most critical cultures because a blood infection may endanger the individual's life. Not only the speed and quality of the diagnosis is important, but also the efficiency in the sample’s supply chain to the laboratory and its preservation conditions. To this end, the supply circuits of blood culture samples to the microbiology laboratory play a major role in the process, being the samples from intensive care units (ICUs) among the most critical. The work presents the analysis and optimization of the ordering and supply process of blood culture samples from an intensive care unit to a microbiology laboratory using LSS methodology and tools. The process starts with the blood culture’s order generation from the intensive care unit specialist and ends with the final reception of the blood culture sample in the pre-analytical area of the microbiology laboratory. The project is structured considering DMAIC problem-solving methodology. Main lean tools used for the supply process analysis and optimization were Gemba Walk, Value Stream Analysis (VSA) and mistake proofing. After project results validation with ICU and microbiology specialists, the next steps are the pilot implementation of the process improvement measures and their impact evaluation on the key performance indicators. The development of a lean concept, eliminating the waste and increasing the added-value of the blood culture samples ordering and supply process, improves the efficiency of the blood culture diagnosis process reducing the lead-time and impacting directly on the patient.