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.

Aspergillus granulosus femoral osteomyelitis in a cardiac transplant patient: first reported case and literature review.

Aspergillus osteomyelitis is a rare complication of extrapulmonary invasive aspergillosis, which usually presents as spondylodiscitis. The clinical picture is usually paucisymptomatic and of long evolution, which leads to diagnostic difficulties, especially in immunosuppressed patients presenting a delayed systemic host response. We report a case of femoral osteomyelitis caused by Aspergillus granulosus in a heart transplant recipient successfully treated with a combined surgical and antifungal approach. A 65-year-old heart transplant male presented with left knee pain lasting 3 months. X-ray and magnetic resonance imaging identified a lesion with aggressive characteristics at the distal third of the left femur, due to which the patient underwent excisional surgery. Aspergillus granulosus was cultured from the removed material and antifungal treatment with oral isavuconazole was started. Chest imaging excluded pulmonary aspergillosis, while the positron emission tomography/computed tomography (PET/CT) identified a remnant of a prosthetic vascular graft sewn to the proximal third of the right axillary artery, through which a catheter-based micro-axial left ventricular assist device was implanted previously as bridge to transplant therapy. The patient presented a rapid clinical improvement with complete functional recovery following the surgical treatment and the antifungal therapy and finally underwent surgical removal of the residual vascular graft. This is the first reported episode of long bone osteomyelitis due to A. granulosus that occurred in a heart transplant recipient without pulmonary infection and was successfully treated with isavuconazole. The PET/CT was useful in supporting the diagnostic process and follow-up. Cryptic fungal species can cause invasive infections, particularly in immunocompromised patients. Molecular methods are crucial in fungal identification.

Severe Impairment of T-cell Immunity and Pulmonary GvHD Are Major Risk Factors for Nontuberculous Mycobacterial Infection After Pediatric Allogeneic Hematopoietic Stem Cell Transplantation.

Hematopoietic stem cell transplant recipients are prone to infectious complications. Infections caused by nontuberculous mycobacteria have increased in adults but literature in children is scarce. We report 6 episodes of disseminated or pulmonary nontuberculous mycobacteria infection among 5 pediatric hematopoietic stem cell transplant recipients. All but one were caused by Mycobacterium avium complex. Four patients died, 2 related to nontuberculous mycobacteria infection.

Prevalence of mutations associated with macrolide and fluoroquinolone resistance in Neisseria gonorrhoeae with Allplex™ NG&DR Assay (Seegene®) in a tertiary hospital from Madrid, Spaina / Prevalencia de mutaciones asociadas a resistencia a macrólidos y fluoroquinolonas en Neisseria gonorrhoeae con el kit Allplex™ NG&DR Assay (Seegene®) en un hospital terciario de la Comunidad de Madrid, España

Background. The prevalence of drug-resistant Neisseria gonorrhoeae (NG) infections is increasing. Studies report the prevalence of NG strains presenting A2059G/C2611T (rRNA 23S) and S91F (parC) mutations conferring resistance to azith romycin and ciprofloxacin. Material and methods. We conducted a prospective cohort study evaluating first void-urine urines, rectal, and oropharyngeal swabs collected from a cohort of patients in a tertiary hospital in Madrid between October 2022 and January 2023. Samples were screened by Allplex™ 7-STI Essential As say (Seegene®). Drug resistances were performed by Allplex™ NG&DR Assay (Seegene®). Results. A total of 1,415 patients were included, of which 112 had a positive sample for NG infection. One patient had a C2611T mutation (0.9%) and neither patient showed A2059G mutation. We found 67 (59.8%) S91F-positive patients. For ty-four patients (39.3%) not had any mutations. Conclusions. We report a low-prevalence of mutations A2059G/C2611T to macrolides and a high-prevalence to S91F in NG infections. Molecular methods for the detection of NG resistance could be useful in direct non-culturable samples (AU)

Introducción. La infección por Neisseria gonorrhoeae (NG) resistente está aumentando. Se ha descrito la prevalencia de cepas de NG con mutaciones A2059G/C2611T (rRNA 23S) y S91F (parC) que confieren resistencia a azitromicina y cipro floxacino. Material y métodos. Realizamos un estudio prospecti vo evaluando orinas de primera micción, hisopos anales y fa ríngeos recogidos de una cohorte de pacientes en un hospital terciario de Madrid entre octubre de 2022 y enero de 2023. El cribado de las muestras se realizó mediante Allplex™ 7-STI Es sential Assay (Seegene®). Las resistencias a macrólidos y fluo roquinolonas se realizaron mediante Allplex™ NG&DR Assay (Seegene®). Resultados. Se incluyeron 1.415 pacientes, de los cua les 112 fueron positivos para NG. Un paciente presentaba una mutación C2611T (0,9%) y en ningún paciente se detec tó A2059G. Encontramos 67 pacientes (59,8%) positivos pa ra S91F. Cuarenta y cuatro pacientes (39,3%) no presentaban mutaciones. Conclusiones. Reportamos una baja prevalencia de mu taciones A2059G/C2611T a macrólidos y una alta prevalencia de S91F en NG. Los métodos moleculares para la detección de resistencias en NG podrían ser útiles en muestras directas no cultivables (AU)

Epidemiology and prevalence of mutations associated with resistance to macrolides and fluoroquinolones in Mycoplasma genitalium in a tertiary hospital from Madrid, Spain / Epidemiología y prevalencia de mutaciones asociadas a la resistencia a macrólidos y fluoroquinolonas en Mycoplasma genitalium en un hospital terciario de Madrid, España

Objectives: Mycoplasma genitalium causes persistent sexually transmitted infections. The aims of this study were to estimate the prevalence of resistances to macrolides and fluoroquinolones in M. genitalium and the sexually transmitted coinfections in patients at Hospital Universitario La Paz (Madrid, Spain). Material and methods: Patients attended between January and October 2021 were studied. Screening for sexually transmitted pathogens and detection of 23S rRNA and parC genes mutations were performed by real-time PCR (Allplex,SeegeneTM). Results: A total of 1,518 females and 1,136 males were studied. The prevalence of M. genitalium was 2.1%. The macrolides resistance rate was 51.8%. The mutations found were A2059G, A2058T and A2058G. The rate of resistance to fluoroquinolones was 17.8% being the G248T mutation (S83I) the most frequent. Seven males had some sexual transmitted coinfection. Conclusions: Although the percentage of M. genitalium infections is low, the high rate of resistance to macrolides makes it necessary to revise the protocols for diagnosis and empirical treatment of sexually transmitted infections. The use of fluoroquinolones is appropriate after screening of macrolide resistance profile. (AU)

Objetivos: Mycoplasma genitalium causa infecciones de transmisión sexual persistentes. Los objetivos de este trabajo fueron estimar la prevalencia de resistencias a macrólidos y fluoroquinolonas en M. genitalium así como las coinfecciones de transmisión sexual en pacientes del Hospital Universitario La Paz (Madrid, España). Material y métodos: Se estudiaron pacientes atendidos entre enero y octubre de 2021. El cribado de patógenos de transmisión sexual y la detección de mutaciones de los genes ARNr 23S y parC se realizaron por PCR en tiempo real (Allplex, SeegeneTM). Resultados: Se estudiaron 1.518 mujeres y 1.136 hombres. La prevalencia de M. genitalium fue del 2,1%. La tasa de resistencia a macrólidos fue del 51.8%. Las mutaciones encontradas fueron A2059G, A2058T y A2058G. La tasa de resistencias a fluoroquinolonas fue del 17.8% siendo la mutación G248T (S83I) la más frecuente. Siete hombres presentaron alguna coinfección de transmisión sexual. Conclusiones: Aunque el porcentaje de infecciones por M. genitalium es bajo, la elevada tasa de resistencias frente a macrólidos hace necesario modificar los protocolos de diagnóstico y tratamiento empírico de las infecciones de transmisión sexual. El uso de fluoroquinolonas es adecuado tras testar previamente el perfil de resistencia a macrólidos. (AU)

Optimized identification of microorganisms directly from positive blood cultures by MALDI-TOF to improve antimicrobial treatment / Optimización en el proceso de identificación directamente de hemocultivos positivos por MALDI-TOF para mejorar el tratamiento antimicrobiano

Introduction. Bacteriemia is a major cause of morbidityand mortality among hospitalized patients worldwide. Earlyidentification of microorganisms from blood culture can leadto improvement of treatment and outcomes.Methods. The study was divided into two phases. Thefirst phase when a comparison of the methods was made tocheck the concordance between them, using as a reference thestandard method implemented in the laboratory. In a secondphase, both methods are combined. We used the rapid identification method and when it could not identify we used thestandard method. The microorganisms that were not identifiedby either of the two methods were identified from colony at24 hoursResults. A total of 589 microbial positive blood cultures have been included in the present study. With the rapidmethod we obtained 96% and 88% identification results forGram-negative bacilli (GNB) and Gram-positive cocci (GPC)respectively. In this study we observed that the combinationof the rapid and standard method achieved identifications of98% and 97% for GNB and GPC respectively.Conclusions. The data analysed shows that both methodscombined perform better than individually. We achieved anoptimization of the identification of microorganisms directlyfrom positive blood cultures by MALDI-TOF. This combinationidentified 98% of the microorganisms in between ten minutesto one hour and a half since the blood culture flagged positive (AU)

Introducción. La bacteriemia es una de las principalescausas de morbilidad y mortalidad entre los pacientes hospitalizados de todo el mundo. La identificación temprana de losmicroorganismos que están en la sangre, permite optimizar lostratamientos y conseguir mejores resultados.Material y métodos. El estudio se dividió en dos fases. Enla primera fase se realizó una comparación de los dos métodospara comprobar la concordancia entre ambos, tomando comoreferencia el método estándar implementado en el laboratorio.La segunda fase combinó ambos métodos para la identificación de hemocultivos positivos. Se utilizó el método de identificación rápida como primera opción y el método estándarsolo cuando no se consiguió identificar por la primera opción.Los microorganismos que no fueron identificados por ningunode los dos métodos, se identificaron directamente de la coloniacrecida a las 24 horas.Resultados. Se analizaron un total de 589 hemocultivospositivos en este estudio. Con el método rápido obtuvimos un96% y 88% de identificación de bacilos gramnegativos y cocosgrampositivos respectivamente. En este estudio observamosque la combinación del método rápido y el método estándarconsiguió identificaciones del 98% y 97% para bacilos gramnegativos y cocos grampositivos respectivamente.Conclusiones. Los datos analizados muestran que ambosmétodos combinados consiguen mejores resultados que utilizados de forma individual. Logramos una optimización de laidentificación de microorganismos directamente a partir dehemocultivos positivos por MALDI-TOF. Con esta combinaciónse identificó el 98% de los microorganismos entre los primeros10 minutos y hora y media de hemocultivo positivo. (AU)