Single case of lymphogranuloma venereum on the tongue.

Traditionally, lymphogranuloma venereum (LGV) has been associated with disease of the genital area. However, atypical presentations and proctitis are increasingly observed. We report a case of LGV affecting the dorsum of the tongue, which presented as a very painful ulcer. The response to doxycycline (100 mg two times per day for 21 days) was satisfactory. This case may represent a paradigm shift in the differential diagnosis of lingual ulcers.

Comparison of gradient diffusion and molecular methods using Allplex™ NG&DR assay (Seegene®) for macrolide and fluoroquinolone screening resistance in Neisseria gonorrhoeae.

Antimicrobial resistance in Neisseria gonorrhoeae (NG) is increasing worldwide. Second-line treatments with macrolides or fluoroquinolones are an option for NG infections in some cases following the STI guideline recommendations. In our study, we compared the gradient diffusion test using EUCAST 2024 breakpoints with a new molecular method using the Allplex™ NG&DR assay (Seegene®) including A2059G/C2611 mutations (23S rRNA) associated with high/moderate-level macrolide resistance and S91F mutation (gyrA) relationship with fluoroquinolone resistance in NG isolates (n = 100). We calculated the sensitivity, specificity, and correlation of the molecular test for fluoroquinolone using the gradient diffusion as the reference method. In twenty-three strains was not detected any mutation associated with macrolides or fluoroquinolone resistance. No A2059G/C2611T mutations were detected, and the S91F mutations were detected in 77 out of the 100 isolates screened. Twenty-three NG isolates were reported to be resistant to azithromycin (ECOFF: >1 mg/L), and 78 NG isolates were resistant to ciprofloxacin (MIC: >0.06 mg/L). The molecular method showed a sensitivity of 96.1% and, a specificity of 90.9% for fluoroquinolone susceptibility, but the statistical analysis between the molecular test and gradient diffusion test was not statistically significant for fluoroquinolone resistance (p = 1). Statistical analysis was not performed for macrolides because of the absence of positive RT-PCR results. According to our data, Allplex™ assay cannot replace the gradient diffusion test for macrolide resistance. However, the assay could be used to test fluoroquinolone resistance in NG isolates as a replacement for phenotypic methods.

Fluconazole-resistant Candida parapsilosis genotypes from hospitals located in five Spanish cities and one in Italy: Description of azole-resistance profiles associated with the Y132F ERG11p substitution.

BACKGROUND: Fluconazole-resistant Candida parapsilosis is a matter of concern. OBJECTIVES: To describe fluconazole-resistant C. parapsilosis genotypes circulating across hospitals in Spain and Rome and to study their azole-resistance profile associated with ERG11p substitutions. PATIENTS/METHODS: We selected fluconazole-resistant C. parapsilosis isolates (n = 528 from 2019 to 2023; MIC ≥8 mg/L according to EUCAST) from patients admitted to 13 hospitals located in five Spanish cities and Rome. Additionally, we tested voriconazole, posaconazole, isavuconazole, amphotericin B, micafungin, anidulafungin and ibrexafungerp susceptibility. RESULTS: Of the 53 genotypes found, 49 harboured the Y132F substitution, five of which were dominating city-specific genotypes involving almost half the isolates. Another genotype involved isolates harbouring the G458S substitution. Finally, we found two genotypes with the wild-type ERG11 gene sequence and one with the R398I substitution. All isolates were fully susceptible/wild-type to amphotericin B, anidulafungin, micafungin and ibrexafungerp. The azole-resistance patterns found were: voriconazole-resistant (74.1%) or voriconazole-intermediate (25.2%), posaconazole-resistant (10%) and isavuconazole non-wild-type (47.5%). Fluconazole-resistant and voriconazole non-wild-type isolates were likely to harbour substitution Y132F if posaconazole was wild type; however, if posaconazole was non-wild type, substitution G458S was indicated if isavuconazole MIC was >0.125 mg/L or substitution Y132F if isavuconazole MIC was ≤0.125 mg/L. CONCLUSIONS: We detected a recent clonal spread of fluconazole-resistant C. parapsilosis across some cities in Spain, mostly driven by dominating city-specific genotypes, which involved a large number of isolates harbouring the Y132F ERG11p substitution. Isolates harbouring substitution Y132F can be suspected because they are non-susceptible to voriconazole and rarely posaconazole-resistant.

Trends in antifungal resistance in Candida from a multicenter study conducted in Madrid (CANDIMAD study): fluconazole-resistant C. parapsilosis spreading has gained traction in 2022.

We previously conducted a multicenter surveillance study on Candida epidemiology and antifungal resistance in Madrid (CANDIMAD study; 2019-2021), detecting an increase in fluconazole-resistant Candida parapsilosis. We here present data on isolates collected in 2022. Furthermore, we report the epidemiology and antifungal resistance trends during the entire period, including an analysis per ward of admission. Candida spp. incident isolates from blood cultures and intra-abdominal samples from patients cared for at 16 hospitals in Madrid, Spain, were tested with the EUCAST E.Def 7.3.2 method against amphotericin B, azoles, micafungin, anidulafungin, and ibrexafungerp and were molecularly characterized. In 2022, we collected 766 Candida sp. isolates (686 patients; blood cultures, 48.8%). Candida albicans was the most common species found, and Candida auris was undetected. No resistance to amphotericin B was found. Overall, resistance to echinocandins was low (0.7%), whereas fluconazole resistance was 12.0%, being higher in blood cultures (16.0%) mainly due to fluconazole-resistant C. parapsilosis clones harboring the Y132F-R398I ERG11p substitutions. Ibrexafungerp showed in vitro activity against the isolates tested. Whereas C. albicans was the dominant species in most hospital wards, we observed increasing C. parapsilosis proportions in blood. During the entire period, echinocandin resistance rates remained steadily low, while fluconazole resistance increased in blood from 6.8% (2019) to 16% (2022), mainly due to fluconazole-resistant C. parapsilosis (2.6% in 2019 to 36.6% in 2022). Up to 7 out of 16 hospitals were affected by fluconazole-resistant C. parapsilosis. In conclusion, rampant clonal spreading of C. parapsilosis fluconazole-resistant genotypes is taking place in Madrid.

Evidence of Fluconazole-Resistant Candida parapsilosis Genotypes Spreading across Hospitals Located in Madrid, Spain and Harboring the Y132F ERG11p Substitution.

We have been monitoring the antifungal resistance in Candida parapsilosis isolates collected from inpatients at Madrid metropolitan area hospitals for the last 3 years. The study aimed to elucidate the presence of fluconazole-resistant C. parapsilosis genotypes in Madrid. From January 2019 to December 2021, a total of 354 C. parapsilosis isolates (n = 346 patients) from blood (76.6%) or intraabdominal samples were collected and genotyped using species-specific microsatellite markers. Antifungal susceptibilities to amphotericin B, the triazoles, micafungin, anidulafungin, and ibrexafungerp were performed according to EUCAST E.Def 7.3.2; the ERG11 gene was sequenced in fluconazole-resistant isolates. A total of 13.6% (n = 48/354) isolates (one per patient) were found to be resistant to fluconazole and non-wild-type to voriconazole but fully susceptible to ibrexafungerp. Resistant isolates were mostly sourced from blood (n = 45/48, 93.8%) and were detected in five hospitals. Two hospitals accounted for a high proportion of resistant isolates (n = 41/48). Resistant isolates harbored either the Y132F ERG11p amino acid substitution (n = 43) or the G458S substitution (n = 5). Isolates harboring the Y132F substitution clustered into a clonal complex involving three genotypes (one genotype accounted for n = 39/43 isolates) that were found in four hospitals. Isolates harboring the G458S substitution clustered into another genotype found in a fifth hospital. C. parapsilosis genotypes demonstrating resistance to fluconazole have been spreading across hospitals in Madrid, Spain. Over the last 3 years, the frequency of isolation of such isolates and the number of hospitals affected is on the rise.

Blood and intra-abdominal Candida spp. from a multicentre study conducted in Madrid using EUCAST: emergence of fluconazole resistance in Candida parapsilosis, low echinocandin resistance and absence of Candida auris.

OBJECTIVES: We prospectively monitored the epidemiology and antifungal susceptibility of Candida spp. from blood cultures and intra-abdominal samples in patients admitted to hospitals in the Madrid area. METHODS: Between 2019 and 2021, we prospectively collected incident isolates [one per species, patient and compartment (blood cultures versus intra-abdominal samples)] from patients admitted to any of 16 hospitals located in Madrid. We studied the antifungal susceptibilities to amphotericin B, triazoles, micafungin, anidulafungin and ibrexafungerp following the EUCAST E.Def 7.3.2 procedure. RESULTS: A total of 2107 Candida spp. isolates (1895 patients) from blood cultures (51.7%) and intra-abdominal samples were collected. Candida albicans, the Candida glabrata complex, the Candida parapsilosis complex, Candida tropicalis and Candida krusei accounted for 96.9% of the isolates; in contrast, Candida auris was undetected. Fluconazole resistance in Candida spp. was higher in blood cultures than in intra-abdominal samples (9.1% versus 8.2%; P > 0.05), especially for the C. parapsilosis complex (16.6% versus 3.6%, P < 0.05), whereas echinocandin resistance tended to be lower in blood cultures (0.5% versus 1.0%; P > 0.05). Resistance rates have risen, particularly for fluconazole in blood culture isolates, which increased sharply in 2021. Ibrexafungerp showed in vitro activity against most isolates. Species distributions and resistance rates varied among hospitals. CONCLUSIONS: Whereas no C. auris isolates were detected, fluconazole-resistant C. parapsilosis isolates have been spreading across the region and this has pulled up the rate of fluconazole resistance. In contrast, the rate of echinocandin resistance continues to be low.

A matrix-assisted laser desorption/ionization time of flight mass spectrometry reference database for the identification of Histoplasma capsulatum.

The isolation of the pathogenic fungus Histoplasma capsulatum from cultures together with the visualization of typical intracellular yeast in tissues are the gold standard methods for diagnosis of histoplasmosis. However, cultures are time-consuming, require level 3 containment and experienced personnel, and usually call for an additional confirmation test. Matrix-Assisted Laser Desorption Ionization Time of Flight Mass Spectrometry (MALDI-ToF MS) has been established as a suitable tool for microbial identification in several clinical laboratories. A reference database has been constructed for the identification of H. capsulatum by MALDI-ToF MS by using six H. capsulatum strains previously identified by molecular methods. For validation, 63 fungal strains belonging to the Collection of the Spanish National Centre for Microbiology were tested against the new reference database combined with other commercial and in-house databases. In a blind assay, all H. capsulatum strains (n = 30) were correctly identified by the database and 86.6% had scores above 1.7. Considering both phases of the fungus for the same strain, the most reliable results were obtained with the mycelial phase, with only 13.3% of isolates having scores below 1.7. The new database was able to identify both morphological phases of the fungus. MALDI-ToF technology yields a prompt and simple identification from H. capsulatum yeast forms and early mycelial cultures. It allows for reducing response time and decreasing risk in fungus manipulation.

[Application of mass spectrometry in mycology]. / Aplicación de la espectrometría de masas en micología.

MALDI-TOF (matrix-assisted laser desorption ionization time-of-flight) mass spectrometry (MS) is becoming an essential tool in most microbiology laboratories. At present, by using a characteristic fungal profile obtained from whole cells or through simple extraction protocols, MALDI-TOF MS allows the identification of pathogenic fungi with a high performance potential. This methodology decreases the laboratory turnaround time, optimizing the detection of mycoses. This article describes the state-of-the-art of the use of MALDI-TOF MS for the detection of human clinical fungal pathogens in the laboratory and discusses the future applications of this technology, which will further improve routine mycological diagnosis.

Increase in gonococcal arthritis in Madrid, Spain, 2022-2023.

In recent years, there has been an increase in Neisseria gonorrhoeae infections in Europe and Spain. Disseminated gonococcal infection is an uncommon clinical presentation that includes gonococcal arthritis. Improved antibiotic treatment has reduced the incidence of gonococcal arthritis. However, the increase in gonococcal infections may have increased the frequency of this clinical entity in recent times. We report five cases of gonococcal arthritis in patients in a tertiary-care hospital in the northern area of Madrid (Spain) from October 2022 to October 2023. Major cases occurred in male patients with unprotected sex and polyarticular symptoms requiring hospital admission and treatment with ceftriaxone and cefixime. The use of molecular techniques has allowed the detection of a greater number of culture-negative cases of gonococcal arthritis, as well as the detection of mutations associated with resistance to fluoroquinolone for switching to oral treatment.

Pneumonia in Children With Complex Chronic Conditions With Tracheostomy: An Emerging Challenge.

BACKGROUND: Despite respiratory infections being a leading cause of hospitalization in children with tracheostomy tubes, there are no published guidelines for their diagnosis and management. This study aims to outline the clinical, laboratory and microbiological aspects of pneumonia in these children, along with the antibiotics used and outcomes. Additionally, it seeks to determine pneumonia incidence and associated risk factors. METHODS: We conducted a retrospective study using the medical records of tracheostomized children at La Paz University Hospital in Madrid from 2010 to 2021. RESULTS: Thirty-three pneumonia cases were observed in 25 tracheostomized children. Pseudomonas aeruginosa was the predominant bacterium (52%), followed by Escherichia coli, Staphylococcus aureus and Serratia marcescens. The same microorganism isolated in the tracheal aspirate culture during pneumonia was previously isolated in 83% of cases that had a similar culture, with some growth obtained within 7-30 days prior. Multiplex respiratory PCR detected respiratory viruses in 73% of cases tested. Antibiotic treatment was administered in all cases except 1, mostly intravenously (81%), with piperacillin/tazobactam and meropenem being commonly used. Only 1 of the described episodes had a fatal outcome. CONCLUSIONS: It is advisable to include coverage for P. aeruginosa, E. coli, S. aureus, and S. marcescens in the empirical antibiotic treatment for pneumonia in tracheostomized children, along with the microorganisms identified in tracheal cultures obtained within 7-30 days prior, if available. A positive PCR for respiratory viruses is often discovered in bacterial pneumonia in tracheostomized children.

Mechanisms of linezolid resistance among Staphylococci in a tertiary hospital.

The mechanisms of linezolid resistance among 86 staphylococcal isolates from two intensive care units were investigated. The most frequent was the G2576T mutation in the 23S rRNA (82%). The cfr gene was found in 17% of the isolates, seven S. aureus and eight S. epidermidis isolates. Four of the S. epidermidis isolates had the G2576T mutation and the cfr gene. In four S. haemolyticus isolates, the mechanism could not be identified.

Isolation of filamentous basidiomycetes from respiratory samples in a tertiary care Spanish hospital.

BACKGROUND: The clinical significance of the filamentous basidiomycetes isolated from clinical samples is not always clear. Thus, these fungi have been considered environmental contaminants traditionally. AIMS: To review those clinical cases in which filamentous basidiomycetes from respiratory samples had been isolated. METHODS: The retrospective study was carried out in a single tertiary care hospital. We recovered all culture-confirmed isolations of filamentous basidiomycetes from respiratory samples (bronchial aspirate [BAS], bronchoalveolar lavage [BAL] and sputum) analyzed between the years 2020 and 2023. Isolates were identified by ITS region sequencing. RESULTS: In six patients a filamentous basidiomycete had been isolated from a respiratory sample. The species identified were all different: Fomitopsis sp. (BAS), Trametes ljubarskyi (BAL), Stereum gausapatum (BAS), Porostereum spadiaceum (BAS), Phlebia subserialis (sputum) and Inonotus levis (BAL). All the patients were immunosuppressed or had an underlying disease with pulmonary involvement. None of them received any specific antifungal treatment (in relation with the fungus isolated) and all six improved clinically and were discharged. CONCLUSIONS: The isolation of filamentous basidiomycetes in these patients had uncertain clinical significance. However, the isolation of any filamentous basidiomycete in respiratory samples from immunosuppressed patients or patients with chronic pulmonary disease is an emerging situation that should be carefully assessed in the context of chronic allergic episodes or suspicion of invasive fungal infections.

Sexually transmitted infections and clinical features in monkeypox (mpox) patients in Madrid, Spain.

Since the beginning of the monkeypox (mpox) virus outbreak in May 2022, there has been an increase in the number of cases worldwide in the setting of sexual transmission. We have tested by real-time PCR 187 mpox patients, of which 157 patients were screened for sexually transmitted infections (STI) in 245 samples. Thirty-six pathogens were detected in 30 patients: herpes simplex virus (HSV-I/II, 12/36, 33.3%), Neisseria gonorrhoeae (NG, 9/36, 25%), Chlamydia trachomatis (CT, 5/36, 13.8%), Chlamydia trachomatis-lymphogranuloma venereum (CT-LGV, 3/36, 8.3%), Treponema pallidum (TP, 4/36, 11.1%) and Mycoplasma genitalium (MG, 3/36, 8.3%). Screening of STI is recommended in mpox patients for the differential diagnosis of the main infections of sexual tract especially in patients with rectal involvement.

Correction: Díaz-García et al. Candida Genotyping of Blood Culture Isolates from Patients Admitted to 16 Hospitals in Madrid: Genotype Spreading during the COVID-19 Pandemic Driven by Fluconazole-Resistant C. parapsilosis. J. Fungi 2022, 8, 1228.

In the original publication [...].

Antifungal resistance in Candida spp within the intra-abdominal cavity: study of resistance acquisition in patients with serial isolates.

OBJECTIVES: Antifungal susceptibility testing is mostly conducted on blood-cultured Candida spp isolates. Because the intra-abdominal cavity has been highlighted as a hidden echinocandin-resistant C. glabrata reservoir, we assessed whether testing sequential isolates from a given patient might increase the chances of detecting antifungal resistance. METHODS: Intra-abdominal initial and sequential isolates from the same species from patients included in the CANDIdaemia in MADrid study (January 2019 to June 2022) were studied. We assessed antifungal susceptibility to amphotericin B, azoles, anidulafungin, micafungin, and ibrexafungerp using European Committee on Antimicrobial Susceptibility Testing (EUCAST) methodology and molecularly characterized resistant isolates. RESULTS: We collected 308 isolates (C. albicans [n = 179/308; 58.1%], C. glabrata [n = 101/308; 32.8%], C. tropicalis [n = 17/308; 5.5%], and C. parapsilosis [n = 11/308; 3.6%]) from 112 patients distributed as incident (n = 125/308) and sequential (n = 183/308). Per patient resistance rates of fluconazole (13.4% [15/112] vs. 8% [9/112]); 5.4% proportions difference (95% CI, -2.7% to 13.5%, p 0.09) and echinocandins (8.9% [10/112] vs. 1.8% [2/112]); 7.1% proportions difference (95% CI; 1.2-12.9%; p 0.01) were higher when considering all available isolates than only incident isolates. Resistance was detected in 18 of 112 patients and would have been overlooked in 11 of 18 (61.1%) patients if only incident isolates had been studied. Of the patients who harboured fluconazole or echinocandin-resistant isolates, 14 of 15 and 8 of 10 had received or were receiving fluconazole or echinocandins, respectively. DISCUSSION: Testing sequential Candida isolates from intra-abdominal samples is required to detect antifungal resistance, particularly to echinocandins, in patients whose incident isolates turned out to be susceptible. Furthermore, patients with echinocandin-resistant infections had frequently used echinocandins and had common secondary resistance acquisition.

Candida Genotyping of Blood Culture Isolates from Patients Admitted to 16 Hospitals in Madrid: Genotype Spreading during the COVID-19 Pandemic Driven by Fluconazole-Resistant C. parapsilosis.

BACKGROUND: Candidaemia and invasive candidiasis are typically hospital-acquired. Genotyping isolates from patients admitted to different hospitals may be helpful in tracking clones spreading across hospitals, especially those showing antifungal resistance. METHODS: We characterized Candida clusters by studying Candida isolates (C. albicans, n = 1041; C. parapsilosis, n = 354, and C. tropicalis, n = 125) from blood cultures (53.8%) and intra-abdominal samples (46.2%) collected as part of the CANDIMAD (Candida in Madrid) study in Madrid (2019-2021). Species-specific microsatellite markers were used to define the genotypes of Candida spp. found in a single patient (singleton) or several patients (cluster) from a single hospital (intra-hospital cluster) or different hospitals (widespread cluster). RESULTS: We found 83 clusters, of which 20 were intra-hospital, 49 were widespread, and 14 were intra-hospital and widespread. Some intra-hospital clusters were first detected before the onset of the COVID-19 pandemic, but the number of clusters increased during the pandemic, especially for C. parapsilosis. The proportion of widespread clusters was significantly higher for genotypes found in both compartments than those exclusively found in either the blood cultures or intra-abdominal samples. Most C. albicans- and C. tropicalis-resistant genotypes were singleton and presented exclusively in either blood cultures or intra-abdominal samples. Fluconazole-resistant C. parapsilosis isolates belonged to intra-hospital clusters harboring either the Y132F or G458S ERG11p substitutions; the dominant genotype was also widespread. CONCLUSIONS: the number of clusters-and patients involved-increased during the COVID-19 pandemic mainly due to the emergence of fluconazole-resistant C. parapsilosis genotypes.

[Systemic antifungal drugs]. / Antifúngicos de uso sistémico.

Invasive fungal infections have increased over the last decades and the therapeutic choices to treat them are limited. The antifungal agents currently available are useful and have optimal in vitro activity; however, their activity can be lowered due to the development of fungal resistance. The increase in primary or secondary resistance to some antifungal drugs has led to the search of alternatives such as the combination of drugs or the development of new antifungals. In this paper, the activity of the main families of antifungal drugs, polyenes, azoles, echinocandins, 5-fluorocytosine and other new antifungal drugs, are reviewed. The main resistance mechanisms developed by fungi are also described.

Is an Antenatal Screening for Chlamydia trachomatis Necessary in the Current Society?

A screening of Chlamydia trachomatis infection in young pregnant women (≤25 years old) and their newborns was conducted. A total of 136 women were tested with urine samples in the immediate postpartum period. The prevalence was 18.4% (95% confidence interval [CI]: 11.9-24.9%) (25/136) and the rate of perinatal transmission was 35% (7/20). These results support the need for antenatal screening programs in high-risk women in Madrid (Spain).

Evaluation of two real time PCR assays for the detection of bacterial DNA in amniotic fluid.

PURPOSE: The aim of this study was to evaluate two non-commercial Real-Time PCR assays for the detection of microorganisms in amniotic fluid followed by identification by pyrosequencing. METHODS: We collected 126 amniotic fluids from 2010 to 2015 for the evaluation of two Real-Time PCR assays for detection of bacterial DNA in amniotic fluid (16S Universal PCR and Ureaplasma spp. specific PCR). The method was developed in the Department of Microbiology of the University Hospital La Paz. RESULTS: Thirty-seven samples (29.3%) were positive by PCR/pyrosequencing and/or culture, 4 of them were mixed cultures with Ureaplasma urealyticum. The Universal 16S Real-Time PCR was compared with the standard culture (81.8% sensitivity, 97.4% specificity, 75% positive predictive value, 98% negative predictive value). The Ureaplasma spp. specific Real-Time PCR was compared with the Ureaplasma/Mycoplasma specific culture (92.3% sensitivity, 89.4% specificity, 50% positive predictive value, 99% negative predictive value) with statistically significant difference (p=0.005). CONCLUSIONS: Ureaplasma spp. PCR shows a rapid response time (5h from DNA extraction until pyrosequencing) when comparing with culture (48h). So, the response time of bacteriological diagnosis in suspected chorioamnionitis is reduced.