Utilidad de la implementación de un método molecular automatizado en el diagnóstico de agentes causales de infecciones de transmisión sexual / Usefulness of the implementation of an automated molecular method in the diagnosis of causal agents of sexually transmitted infections / Utilidade da implementação de um método molecular automatizado no diagnóstico de agentes causais de infecções sexualmente transmissíveis

Resumen Se presentan los resultados obtenidos en los primeros meses desde la implementación del diagnóstico de agentes causales de infecciones de transmisión sexual (ITS) por PCR en tiempo real en un sistema automatizado. Se estudiaron 46 muestras endocervicales y 39 muestras de uretra masculina, por examen microscópico en fresco, coloración de Gram, cultivo en agar sangre, agar Thayer Martin, galerías miniaturizadas para investigar Ureaplasma spp. y Mycoplasma hominis (Mycoplasma IST3, bioMérieux, Francia) y PCR múltiple para Neisseria gonorrhoeae, Chlamydia trachomatis . Trichomonas vaginalis (BDMaxTM System, Becton Dickinson, EE.UU.). Mediante PCR múltiple se detectó un agente vinculado a ITS en el 48,7% de las muestras uretrales y en el 21,7% de las muestras endocervicales, además de 5 casos de coinfección. El 18,8% de las infecciones se diagnosticaron sólo por PCR. Estos datos demuestran que mediante PCR se optimizó el diagnóstico de ITS en personas de ambos sexos.

Abstract The results obtained in the first months after the implementation of the diagnosis of causative agents of sexually transmitted infections (STIs) by real- time PCR in an automated system are presented. Forty-six endocervical samples and 39 male urethral samples were studied by fresh microscopic examination, Gram staining, blood agar culture, Thayer Martin agar, miniaturised galleries to investigate Ureaplasma spp. and Mycoplasma hominis (Mycoplasma IST3, bioMérieux, France), and multiplex PCR for Neisseria gonorrhoeae, Chlamydia trachomatis and Trichomonas vaginalis (BDMaxTM System, Becton Dickinson, USA). Using multiplex PCR, an agent linked to STIs was detected in 48.7% of the urethral samples and in 21.7% of the endocervical samples, in addition to 5 cases of coinfection. A total of 18.8% of the infections were diagnosed only by PCR. These data show that PCR optimised the diagnosis of STIs in people of both sexes.

Resumo São apresentados os resultados obtidos nos primeiros meses a partir da implementação do diagnóstico de agentes causadores de infecções sexualmente transmissíveis (ISTs) por PCR em tempo real em um sistema automatizado. Quarenta e seis amostras endocervicais e 39 amostras uretrais masculinas foram estudadas por exame microscópico fresco, coloração de Gram, cultura de ágar sangue, ágar Thayer Martin, galerias miniaturizadas para investigar Ureaplasma spp. e Mycoplasma hominis (Mycoplasma IST3, bioMérieux, França) e PCR múltiplo para Neisseria gonorrhoeae, Chlamydia trachomatis e Trichomonas vaginalis (BDMaxTM System, Becton Dickinson, EUA). Utilizando PCR múltiplo, um agente ligado a IST foi detectado em 48,7% das amostras uretrais e em 21,7% das amostras endocervicais, além de 5 casos de coinfecção; 18,8% das infecções foram diagnosticadas apenas por PCR. Esses dados mostram que através do PCR foi otimizado o diagnóstico de IST em pessoas de ambos os sexos.

Orquiepididimitis por Haemophilus parainfluenzae: caso clínico y revisión de la literatura / Orchiepididymitis due to Haemophilus parainfluenzae: case report and review of the literature / Orquiepididimite por Haemophilus parainfluenzae: caso clínico e revisão da literatura

Resumen Haemophilus parainfluenzae forma parte de la microbiota normal de la cavidad oral y del tracto respiratorio superior. Es un reconocido agente causal de endocarditis y, con menor frecuencia, de enfermedades como neumonía, sepsis, osteomielitis, celulitis, meningitis y gastroenteritis aguda. Aquí se presenta un caso de orquiepididimitis en un joven adulto donde H. parainfluenzae, confirmado por espectrometría de masas (MALDI-TOF MS), fue el único patógeno detectado. Este caso contribuye a valorar el rol de H. parainfluenzae como patógeno humano, aislado a partir de sitios diferentes del torrente sanguíneo y las vías respiratorias.

Abstract Haemophilus parainfluenzae is part of the normal microbiota of the oral cavity and the upper respiratory tract. It is a recognised causal agent of endocarditis and, less frequently, of diseases such as pneumonia, sepsis, osteomyelitis, cellulitis, meningitis, and acute gastroenteritis. A case of orchiepididymitis in a young adult is reported, where H. parainfluenzae, confirmed by mass spectrometry (MALDI-TOF MS), was the only pathogen detected. This case contributes to assess the role of H. parainfluenzae as a human pathogen, isolated from sites other than the bloodstream and the respiratory tract.

Resumo Haemophilus parainfluenzae faz parte da microbiota normal da cavidade oral e do trato respiratório superior. É um reconhecido agente causal de endocardite e, menos frequentemente, de doenças como pneumonia, sepse, osteomielite, celulite, meningite e gastroenterite aguda. Aqui é relatado um caso de orquiepididimite em um adulto jovem onde H. parainfluenzae, confirmado por espectrometria de massa (MALDI-TOF MS), foi o único patógeno detectado. Este caso contribui para avaliar o papel do H. parainfluenzae como patógeno humano, isolado de outros locais que não sejam a corrente sanguínea e o trato respiratório.

Utilidad del panel FilmArray® BCID en la detección de bacteriemia: estudio multicéntrico latinoamericano / Usefulness of the FilmArray® BCID panel in the detection of bacteremia: Latin American multicenter study

Resumen Los objetivos de este estudio fueron determinar el desempeño del panel BCID de FilmArray® y establecer el impacto de estos resultados en el tratamiento antimicrobiano de pacientes con bacteriemia en 11 hospitales de Latinoamérica. Se incluyeron 397 episodios de bacteriemia y se documentaron 551 microorganismos aislados de hemocultivos. La identificación microbiana fue correcta en el 91,4% (504/551) de los aislados y en el 98,6% (504/511) si se consideran solo los microorganismos incluidos en el panel BCID. La sensibilidad en la detección de los genes mecA, vanA/B y blaKPC fue del 100% y la especificidad fue del 97%, 100% y 99,6% respectivamente. La notificación temprana del resultado permitió cambios terapéuticos en 242 episodios (60,9%). El panel BCID es un método confiable y rápido para la detección de mecanismos críticos de resistencia y de los microorganismos más frecuentemente aislados de bacteriemias y permite la optimización temprana del tratamiento antimicrobiano.

Abstract The objectives of this study were to determine the performance of the BCID panel and to establish the impact of these results on the antimicrobial treatment of patients with bacteremia in 11 hospitals in Latin America. Three hundred and ninety-seven episodes of bacteremia were included and 551 microorganisms isolated from blood cultures were documented. Microbial identification was correct in 91.4% (504/551) of the isolates and in 98.6% (504/511) if only the microorganisms included in the BCID panel are considered. The sensitivity in the detection of the genes mecA, vanA/B and blaKPC was 100% and the specificity was 97%, 100% and 99.6% respectively. Early notification of the outcome allowed therapeutic changes in 242 episodes (60.9%). The BCID panel is a reliable and rapid method for the detection of critical resistance mechanisms and of the microorganisms most frequently isolated from bacteremia and it enables early optimisation of antimicrobial treatment.

Resumo Os objetivos deste estudo foram determinar o desempenho do painel BCID do FilmArray® e estabelecer o impacto desses resultados no tratamento antimicrobiano de pacientes com bacteremia em 11 hospitais da América Latina. Trezentos e noventa e sete episódios de bacteremia foram incluídos e 551 microrganismos isolados de hemoculturas foram documentados. A identificação microbiana foi correta em 91,4% (504/551) dos isolados e em 98,6% (504/511) considerando apenas os microrganismos incluídos no painel BCID. A sensibilidade na detecção dos genes mecA, vanA/B e blaKPC foi de 100% e a especificidade foi de 97%, 100% e 99,6% respectivamente. A notificação precoce do desfecho permitiu mudanças terapêuticas em 242 episódios (60,9%). O painel BCID é um método confiável e rápido para a detecção de mecanismos críticos de resistência e dos microrganismos mais frequentemente isolados da bacteremia e permite a otimização precoce do tratamento antimicrobiano.

Implementación del método de FilmArrayTM en pacientes pediátricos con infecciones severas / Implementation of the FilmArrayTM method in pediatric patients with severe infections / Implementação do método de matriz do FilmArrayTM em pacientes pediátricos com infecções graves

Se evaluó la implementación del método de PCR múltiple FilmArrayTM (Biofire Diagnostics, LLC, EE.UU.) en 21 niños con infección respiratoria aguda baja, 3 con meningoencefalitis, y un caso de sepsis. Se registraron el tiempo de demora hasta obtener el resultado y adecuar el tratamiento, los días de internación, los patógenos detectados y el costo de la incorporación de esta metodología. En los niños estudiados con FilmArrayTM el resultado estuvo disponible a los 90 minutos desde la toma de la muestra. Se detectaron patógenos no demostrados por los métodos disponibles, como Rhinovirus, además de diagnosticar coinfección viral; el tiempo promedio de estadía resultó 5 días. Se estimó reducir un 40% el costo global de internación. La implementación de FilmArrayTM resultó sencilla y se pudo incorporar a la sistemática de trabajo. Si bien esta experiencia incluyó un bajo número de pacientes, aportó información que demuestra el potencial de esta metodología para un mejor manejo del paciente crítico.

The implementation of multiple PCR FilmArrayTM (Biofire Diagnostics, LLC, USA) for 21 children with low acute respiratory infection, 3 with meningoencephalitis, and 1 case of sepsis was evaluated. Delay time until the result was obtained and the treatment adapted, hospitalization days, pathogens detected and the cost of incorporating this methodology were all recorded. In the children studied with FilmArrayTM the result was available 90 minutes after the sample was taken. Pathogens were not demonstrated by the available methods, such as Rhinovirus, apart from diagnosing viral coinfection, the average length of stay was 5 days. It was estimated to reduce the overall cost of hospitalization by 40%. The implementation of FilmArrayTM was simple and could be incorporated into the work system. Although this experience included a low number of patients, it provided information that demonstrates the potential of this methodology for better management of the critical patient.

Foi avaliada a implementação do método de PCR múltiplo FilmArrayTM (Biofire Diagnostics, LLC, EUA) em 21 crianças com infecção respiratória aguda baixa, 3 com meningoencefalite e um caso de sepse. O tempo de atraso foi registrado até a obtenção do resultado e a adaptação do tratamento, dias de internação, patógenos detectados e o custo da incorporação dessa metodologia. Nas crianças estudadas com o FilmArrayTM, o resultado ficou disponível 90 minutos após a coleta da amostra; foram detectados os patógenos não demonstrados pelos métodos disponíveis, como o Rinovírus, além de diagnosticar a coinfecção viral; o tempo médio de permanência foi de 5 dias. Foi estimado reduzir o custo total da hospitalização em 40%. A implementação do FilmArrayTM foi simples e pôde ser incorporada à sistemática de trabalho. Embora essa experiência tenha incluído um número de pacientes baixo, forneceu informações que demonstram o potencial dessa metodologia para um melhor gerenciamento do paciente crítico.

Bacteriemia por Helicobacter cinaedi: presentación de los primeros casos descritos en Argentina / Helicobacter cinaedi bacteremia: Presentation of the first cases reported in Argentina

Se presentan 2 casos de bacteriemia por Helicobacter cinaedi. El primero se diagnosticó en un varón de 76 años y resultó secundario a la colocación de un acceso vascular; el segundo correspondió a un lactante febril de 37 días de vida, asociado a un cuadro de gastroen-terocolitis aguda. H. cinaedi es un microorganismo que presenta dificultad para desarrollarse en diferentes medios de cultivo y lograr su identificación a nivel de especie. En ambos casos fue fundamental la observación microscópica en fresco de las botellas de hemocultivo, la utilización de la espectrometría de masas y la posterior secuenciación del gen hsp60 para llegar a esa instancia. En los últimos anos se han informado infecciones por H. cinaedi con frecuencia creciente en otras partes del mundo. En este trabajo presentamos los primeros casos de bacteriemia por H. cinaedi documentados en Argentina.

Two cases of bacteremia caused by Helicobacter cinaedi are presented. The first case was diagnosed in a 76-year-old male patient, and was secondary to a vascular access device placement; the second case corresponded to a febrile infant of 37 days of life, and was associated with acute gastroenteritis. H. cinaedi is a microorganism difficult to grow in different culture media and also to identify to species level. In both cases, the microscopic observation of blood culture bottles, the use of mass spectrometry and the subsequent sequencing of the hsp60 gene were essential. In the recent literature, H. cinaedi infections are being reported more frequently. In this report we present the first documented cases of bacteremia caused by H. cinaedi in Argentina.

[Helicobacter cinaedi bacteremia: Presentation of the first cases reported in Argentina]. / Bacteriemia por Helicobacter cinaedi: presentación de los primeros casos descritos en Argentina.

Two cases of bacteremia caused by Helicobacter cinaedi are presented. The first case was diagnosed in a 76-year-old male patient, and was secondary to a vascular access device placement; the second case corresponded to a febrile infant of 37 days of life, and was associated with acute gastroenteritis. H. cinaedi is a microorganism difficult to grow in different culture media and also to identify to species level. In both cases, the microscopic observation of blood culture bottles, the use of mass spectrometry and the subsequent sequencing of the hsp60 gene were essential. In the recent literature, H. cinaedi infections are being reported more frequently. In this report we present the first documented cases of bacteremia caused by H. cinaedi in Argentina.

Training tuberculosis laboratory workers in LED-fluorescence microscopy: experience learned in Argentina.

OBJECTIVE: To assess a LED-fluorescence microscopy (LED-FM) capacitation program for the training of laboratory technicians without previous experience in FM. METHODS: We evaluated a teaching program that consists of a three-day course followed by an "in situ" two-month phase in which technicians acquired skills without the help of a FM expert; in order to gain confidence to recognize auramine-stained bacillus, during this phase, technicians examined duplicate slides stained by Ziehl Neelsen (ZN) and FM in a unblinded way. Technicians with acceptable performance, continued with a blinded-training period. Testing panels and rechecking process were used to evaluate proficiency after different length of experience. RESULTS: Post-course panel results showed that 70% of trainees made Low False Positive errors (LFPs). Analysis of two other panels showed that LFPs significantly decreased (Chi-squared test, p<0.05) as the "in situ" training phase progressed. Processing at least three slides/day was associated with acceptable performance. During the blinded-training period, results of the rechecking process showed that sensitivity (96.8%) and specificity (99.8%) levels were satisfactory. CONCLUSION: Moderate training (a three-day course) is not enough to make technicians proficient in LED-FM; however, great ability can be reached after a short "in situ" training phase even without the presence of experienced staff available in field to review doubtful results. Training was more effective in services with a minimum workload of 750 slides/year.

Genotypic diversity of Mycobacterium tuberculosis in Buenos Aires, Argentina.

Buenos Aires is an overpopulated port city historically inhabited by people of European descent. Together with its broader metropolitan area, the city exhibits medium tuberculosis rates, and receives migrants, mainly from tuberculosis highly endemic areas of Argentina and neighboring countries. This work was aimed to gain insight into the Mycobacterium tuberculosis population structure in two suburban districts of Buenos Aires which are illustrative of the overall situation of tuberculosis in Argentina. The Lineage 4 Euro-American accounted for >99% of the 816 isolates analyzed (one per patient). Frequencies of spoligotype families were T 35.9%, LAM 33.2%, Haarlem 19.5%, S 3.2%, X 1.5%, Ural 0.7%, BOV 0.2%, Beijing 0.2%, and Cameroon 0.2%. Unknown signatures accounted for 5.3% isolates. Of 55 spoligotypes not matching any extant shared international type (SIT) in SITVIT database, 22 fitted into 15 newly-issued SITs. Certain autochthonous South American genotypes were found to be actively evolving. LAM3, which is wild type for RDrio, was the predominant LAM subfamily in both districts and the RDrio signature was rare among autochthonous, newly created, SITs and orphan patterns. Two genotypes that are rarely observed in neighboring countries ̶ SIT2/H2 and SIT159/T1 Tuscany ̶ were conspicuously represented in Argentina. The infrequent Beijing patterns belonged to Peruvian patients. We conclude that the genotype diversity observed reflects the influence of the Hispanic colonization and more recent immigration waves from Mediterranean and neighboring countries. Unlike in Brazil, the RDrio type does not play a major role in the tuberculosis epidemic in Buenos Aires.

Epidemiología de la faringitis aguda bacteriana en un hospital general de agudos / Epidemiology of acute bacterial pharyngitis in a general acute care hospital / Epidemiologia da faringite bacteriana aguda em um hospital geral de cuidados agudos

La faringitis aguda bacteriana (FAB) representa entre un 20 a 30% de las faringitis. El principal agente causal es Streptococcus pyogenes. Se revisó retrospectivamente la información microbiológica disponible entre 2010 y 2016, para analizar la incidencia de la FAB, sus agentes causales, la incidencia de casos según el período estacional y rango etario. Se determinó el porcentaje de resistencia (R) de S. pyogenes a eritromicina y el fenotipo predominante. Se diagnosticó FAB en 21,5% de 3.246 cultivos, donde 89% fueron causados por S. pyogenes. El 35,3% de las FAB se presentó en niños de 7 a 10 años, seguido por el grupo de mayores de 15 años (31,3%). Se evidenció un aumento de faringitis por Streptococcus dysgalactiae subsp. equisimilis en 2011 y 2014; 56,2% de los casos se diagnosticaron en mayores de 15 años. Hubo 3 casos de FAB por Arcanobacterium haemolyticum. El 36,1% de las faringitis se presentaron en primavera, con un aumento significativo durante las correspondientes a 2010 y 2016. La R global a eritromicina en S. pyogenes fue del 16,6% con predominio del fenotipo M. El valor de R encontrado fue superior al ser comparado con otras estadísticas similares, lo que demostró la importancia de monitorear este dato, dado que es el antibiótico alternativo en pacientes alérgicos a penicilina.

Acute bacterial pharyngitis (ABP) accounts for 20 to 30% of acute pharyngitis. The most common bacterial agent is Streptococcus pyogenes. A retrospective, record-based study was carried out based on the microbiological records from 2010 to 2016, analyzing ABP's incidence, its causal agents and its incidence according to the season and age range. Erythromycin resistance and the main resistance phenotype were determined in S. pyogenes. Acute bacterial pharyngitis was diagnosed in 21.5% out of 3.246 cultures, 89% were due to S. pyogenes, 36.3% of patients were children between 7 to 10 years old and 31.3% were older than 15 years of age. There was a significant increase in pharyngitis due to Streptococcus dysgalactiae subsp. equisimilis since 2010 (p<0.05), 52.5% were detected in patients older than 15 years of age. There were only three cases of ABP produced by Arcanobacterium haemolyticum. A total of 36.1% of pharyngitis occurred during spring, with a significant increase during 2010 and 2016 springs. Global erythromycin resistance in S. pyogenes was 16.6% with predominance of the M phenotype. This resistance rate is higher than that described in other similar series, demonstrating the importance of continuously monitoring of macrolide R in S. pyogenes, since they are the antibiotics of choice to treat pharyngitis in patients allergic to penicillin.

A faringite bacteriana aguda (FAB) representa entre 20 e 30% das faringites. O principal agente causador é Streptococcus pyogenes. Foi revista retrospectivamente a informação microbiológica disponível entre 2010 e 2016, para analisar a incidência da FAB, seus agentes causadores, a incidência de casos de acordo com o período sazonal e a faixa etária. A percentagem de resistência (R) de S. pyogenes à eritromicina e ao fenótipo predominante foram determinadas. Foi diagnosticado FAB em 21,5% de 3,246 culturas, 89% das quais foram causadas por S. pyogenes. 35,3% das FAB se apresentou em crianças de 7 a 10 anos, seguidas pelo grupo de mais de 15 anos (31,3%). Houve aumento de faringite por Streptococcus dysgalactiae subsp. equisimilis em 2011 e 2014; 56,2% dos casos foram diagnosticados em jovens de mais de 15 anos. Houve 3 casos de FAB por Arcanobacterium haemolyticum. 36,1% das faringites ocorreu na primavera, com um aumento significativo em 2010 e 2016. A R global a eritromicina em S. pyogenes foi de 16,6% com prevalência do fenótipo M. O valor de R encontrado foi superior em comparação com outras estatísticas semelhantes, demonstrando a importância de monitorar esse dado, pois é o antibiótico alternativo em pacientes alérgicos à penicilina.

Training tuberculosis laboratory workers in LED-fluorescence microscopy: experience learned in Argentina / Capacitación de trabajadores de laboratorios de tuberculosis en microscopia de fluorescencia LED: experiencia en Argentina

ABSTRACT Objective To assess a LED-fluorescence microscopy (LED-FM) capacitation program for the training of laboratory technicians without previous experience in FM. Methods We evaluated a teaching program that consists of a three-day course followed by an "in situ" two-month phase in which technicians acquired skills without the help of a FM expert; in order to gain confidence to recognize auramine-stained bacillus, during this phase, technicians examined duplicate slides stained by Ziehl Neelsen (ZN) and FM in a unblinded way. Technicians with acceptable performance, continued with a blinded-training period. Testing panels and rechecking process were used to evaluate proficiency after different length of experience. Results Post-course panel results showed that 70% of trainees made Low False Positive errors (LFPs). Analysis of two other panels showed that LFPs significantly decreased (Chi-squared test, p<0.05) as the "in situ" training phase progressed. Processing at least three slides/day was associated with acceptable performance. During the blinded-training period, results of the rechecking process showed that sensitivity (96.8%) and specificity (99.8%) levels were satisfactory. Conclusion Moderate training (a three-day course) is not enough to make technicians proficient in LED-FM; however, great ability can be reached after a short "in situ" training phase even without the presence of experienced staff available in field to review doubtful results. Training was more effective in services with a minimum workload of 750 slides/year.(AU)

RESUMEN Objetivo Evaluar un programa de capacitación en microscopía de fluorescencia LED (MF-LED) para el entrenamiento de técnicos de laboratorio sin experiencia en MF. Métodos Se evaluó un programa de capacitación que consiste en un curso de tres días seguido de dos meses de entrenamiento «in situ¼, en donde los técnicos adquirieron habilidades sin presencia de un experto en la práctica diaria; para alcanzar confianza en el reconocimiento del bacilo, los técnicos, durante estos meses, examinaron en forma «no cegada¼ extendidos duplicados teñidos por Ziehl Neelsen (ZN) y MF. Aquellos laboratoristas que lograron rendimiento aceptable continuaron su entrenamiento «a ciegas¼. Su desempeño fue evaluado en distintos períodos del entrenamiento mediante paneles de láminas y relectura de extendidos. Resultados Los resultados de un panel posterior al curso mostraron que 70% de los participantes cometieron errores falsos positivos bajos (FPB). Dos paneles posteriores evidenciaron que los FPB disminuían significativamente (prueba de Chi cuadrado, p<0.05) a medida que el entrenamiento avanzaba. El procesamiento de al menos tres extendidos/ día se asoció con desempeño aceptable. Durante el período a ciegas, la relectura de láminas evidenció que la sensibilidad (96,8%) y especificidad (99,8%) fueron satisfactorias. Conclusiones Una capacitación moderada (curso de tres días) no es suficiente para adquirir competencia en MF-LED; sin embargo, se puede alcanzar habilidad después de una corta capacitación «in situ¼, incluso si no hay personal con experiencia disponible en el servicio para revisar los resultados dudosos diariamente. El entrenamiento fue más efectivo en servicios con carga de trabajo mínima de 750 extendidos/año.(AU)

Performance of LED fluorescence microscopy for the detection of acid-fast bacilli from respiratory samples in peripheral laboratories in Argentina / Rendimiento de la microscopía de fluorescencia LED para la detección de bacilos ácido-alcohol resistentes en muestras respiratorias en laboratorios periféricos de Argentina

ABSTRACT Introduction: Light-emitting diode fluorescence microscopy (LED-FM) has been endorsed by the World Health Organization (WHO) for tuberculosis diagnosis, but its accuracy in HIV-infected patients remains controversial, and only some few studies have explored procedural factors that may affect its performance. Objective: To evaluate the performance of LED-FM for tuberculosis diagnosis in patients with and without HIV infection using a newer, less expensive LED lamp. Materials and methods: We compared the performance of LED-FM and Ziehl-Neelsen (ZN) microscopy on respiratory specimen smears from tuberculosis (TB) suspects and patients on treatment examined by different technicians blinded for HIV-status and for the result of the comparative test. We analyzed the effect of concentrating specimens prior to microscopy using different examination schemes and user-appraisal of the LED device. Results: Of the 6,968 diagnostic specimens collected, 869 (12.5%) had positive Mycobacterium tuberculosis cultures. LED-FM was 11.4% more sensitive than ZN (p<0.01). Among HIV-positive TB patients, sensitivity differences between LED-FM and ZN (20.6%) doubled the figure obtained in HIV-negative patients or in those with unknown HIV status (9.3%). After stratifying by direct and concentrated slides, the superiority of LED-FM remained. High specificity values were obtained both with LED-FM (99.9%) and ZN (99.9%).The second reading of a sample of slides showed a significantly higher positive detection yield using 200x magnification (49.4 %) than 400x magnification (33.8%) (p<0.05). The LED-device had a very good acceptance among the technicians. Conclusion: LED-FM better performance compared with ZN in HIV-infected patients and user-appraisal support the rapid roll-out of LED-FM. Screening at 200x magnification was essential to achieve LED-FM increased sensitivity.

RESUMEN Introducción. La microscopía de fluorescencia con lámpara LED (MF-LED) ha sido recomendada por la Organización Mundial de la Salud (OMS) para el diagnóstico de la tuberculosis, pero su precisión en pacientes con HIV continúa siendo controversial y en pocos estudios se han explorado los factores metodológicos que pueden afectar su utilidad. Objetivo. Evaluar el rendimiento de la MF-LED en el diagnóstico de la tuberculosis en pacientes con HIV y sin él mediante un novedoso dispositivo LED. Materiales y métodos. Se comparó el rendimiento de la MF-LED y la microscopía en frotis de muestras respiratorias con tinción de Ziehl-Neelsen (M-ZN) examinados por técnicos cegados en cuanto al estado de HIV y el resultado de la prueba comparativa. Se analizó el efecto de concentrar muestras antes de la microscopía, usar diferentes esquemas de observación y la valoración con el dispositivo LED. Resultados. De las 6.968 muestras recolectadas, 869 (12,5 %) resultaron con cultivo positivo para Mycobacterium tuberculosis. La MF-LED fue 11,4 % más sensible que la M-ZN (p<0,01). Entre los pacientes con tuberculosis positivos para HIV, la diferencia de sensibilidad entre la MF-LED y la M-ZN (20,6 %) duplicó la cifra obtenida en pacientes negativos para HIV o con estatus desconocido (9,3 %). Al estratificar los frotis en directos y concentrados, se mantuvo la superioridad de la MF-LED. Las especificidades de la MF-LED (99,9 %) y la M-ZN (99,9 %) resultaron elevadas. La lectura de una muestra de frotis mostró una positividad significativamente mayor con un aumento de 200X (49,4 %) que con uno de 400X (33,8 %) (p<0,05). El dispositivo LED tuvo una buena aceptación entre los técnicos. Conclusión. Debido al mejor desempeño de la MF-LED comparada con la M-ZN en pacientes con HIV y su fácil utilización, se recomienda su adopción. La utilización del aumento de 200X fue esencial para el incremento de la sensibilidad de la MF-LED.

[Performance of LED fluorescence microscopy for the detection of acid-fast bacilli from respiratory samples in peripheral laboratories in Argentina].

INTRODUCTION: Light-emitting diode fluorescence microscopy (LED-FM) has been endorsed by the World Health Organization (WHO) for tuberculosis diagnosis, but its accuracy in HIV-infected patients remains controversial, and only some few studies have explored procedural factors that may affect its performance. OBJECTIVE: To evaluate the performance of LED-FM for tuberculosis diagnosis in patients with and without HIV infection using a newer, less expensive LED lamp. MATERIALS AND METHODS: We compared the performance of LED-FM and Ziehl-Neelsen (ZN) microscopy on respiratory specimen smears from tuberculosis (TB) suspects and patients on treatment examined by different technicians blinded for HIV-status and for the result of the comparative test. We analyzed the effect of concentrating specimens prior to microscopy using different examination schemes and user-appraisal of the LED device. RESULTS: Of the 6,968 diagnostic specimens collected, 869 (12.5%) had positive Mycobacterium tuberculosis cultures. LED-FM was 11.4% more sensitive than ZN (p;0.01). Among HIV-positive TB patients, sensitivity differences between LED-FM and ZN (20.6%) doubled the figure obtained in HIVnegative patients or in those with unknown HIV status (9.3%). After stratifying by direct and concentrated slides, the superiority of LED-FM remained. High specificity values were obtained both with LED-FM(99.9%) and ZN (99.9%).The second reading of a sample of slides showed a significantly higher positive detection yield using 200x magnification (49.4 %) than 400x magnification (33.8%) (p;0.05). The LEDdevice had a very good acceptance among the technicians. CONCLUSION: LED-FM better performance compared with ZN in HIV-infected patients and user-appraisal support the rapid roll-out of LED-FM. Screening at 200x magnification was essential to achieve LEDFM increased sensitivity.

Estudio de las infecciones por Staphylococcus aureus en un hospital general de agudos (2002-2013) / Study of Staphylococcus aureus infections in a general acute care hospital (2002-2013)

Se realizó una revisión retrospectiva durante 12 años de las infecciones por Staphylococcus aureus en pacientes adultos (PA) y pediátricos (PP) asistidos en el Hospital Interzonal General de Agudos Evita de Lanús, para conocer la incidencia, los focos y el origen de la infección, y analizar el perfil de resistencia antimicrobiana. Se documentaron 2125 casos de infección en PA y 361 en PP. La incidencia de casos en PA descendió significativamente en el último trienio (χ²; p < 0,05); en los PP aumentó significativamente durante los últimos 5 años (χ²; p < 0,0001). En ambas poblaciones se detectó un aumento significativo en las infecciones de piel y estructuras asociadas (PEA), de bacteriemias a punto de partida de un foco en PEA y de infecciones por S. aureus de inicio en el hospital (χ²; p < 0,005). La meticilino-resistencia (SAMR) aumentó del 28 al 78% en PP; en PA se mantuvo alrededor del 50%, con reducción significativa de la resistencia acompañante a antimicrobianos no betalactámicos en los SAMR de ambas poblaciones. En S. aureus documentados de infecciones de inicio en la comunidad (SAMR-CO) del último trienio, el porcentaje de meticilino-resistencia resultó del 57% en PP y 37% en PA; en infecciones con inicio en el hospital fue del 43 y el 63%, respectivamente. Si bien demostramos que S. aureus continúa siendo un patógeno asociado al ámbito nosocomial, hubo un aumento de infecciones por SAMR-CO, con compromiso en PEA en ambas poblaciones.

A twelve-year retrospective review of Staphylococcus aureus infections in adult and pediatric patients (AP and PP respectively) assisted in the Hospital Interzonal General de Agudos Evita in Lanús was performed to determine the incidence, foci of infection, the source of infection and to analyze the profile of antimicrobial resistance. An amount of 2125 cases of infection in AP and 361 in PP were documented. The incidence in AP decreased significantly in the last three years (χi²; p < 0.05); in PP it increased significantly during the last five years (χ²; p < 0.0001). In both populations was detected a notable increase in skin infections and associated structures (PEA) in bacteremia to the starting point of a focus on PEA, and in total S. aureus infections of hospital-onset (χ²; p < 0.005). Methicillin-resistance (MRSA) increased from 28 to 78% in PP; in AP it remained around 50%, with significant reduction in accompanying antimicrobial resistance to non-β-lactams in both groups of MRSA. In S. aureus documented from community onset infections (CO-MRSA) in the last three years, the percentage of methicillin-resistance was 57% in PP and 37% in AP; in hospital-onset infections it was 43% and 63% respectively. Although data showed that S. aureus remains a pathogen associated with the hospital-onset, there was an increase of CO-MRSA infections with predominance in PEA in both populations.

[Study of Staphylococcus aureus infections in a general acute care hospital (2002-2013)]. / Estudio de las infecciones por Staphylococcus aureus en un hospital general de agudos (2002-2013).

A twelve-year retrospective review of Staphylococcus aureus infections in adult and pediatric patients (AP and PP respectively) assisted in the Hospital Interzonal General de Agudos Evita in Lanús was performed to determine the incidence, foci of infection, the source of infection and to analyze the profile of antimicrobial resistance. An amount of 2125 cases of infection in AP and 361 in PP were documented. The incidence in AP decreased significantly in the last three years (χi2; p<0.05); in PP it increased significantly during the last five years (χ2; p<0.0001). In both populations was detected a notable increase in skin infections and associated structures (PEA) in bacteremia to the starting point of a focus on PEA, and in total S. aureus infections of hospital-onset (χ2; p < 0.005). Methicillin-resistance (MRSA) increased from 28 to 78% in PP; in AP it remained around 50%, with significant reduction in accompanying antimicrobial resistance to non-ß-lactams in both groups of MRSA. In S. aureus documented from community onset infections (CO-MRSA) in the last three years, the percentage of methicillin-resistance was 57% in PP and 37% in AP; in hospital-onset infections it was 43% and 63% respectively. Although data showed that S. aureus remains a pathogen associated with the hospital-onset, there was an increase of CO-MRSA infections with predominance in PEA in both populations.

LED fluorescence microscopy in the diagnosis of tuberculosis: Fading and restaining of smears for external quality assessment.

Blinded rechecking is a method proposed for external quality assurance (EQA) of auramine-stained acid-fast bacilli (AFB) smears using fluorescence microscopy (FM), however, this procedure is not well developed and slides fading over time could compromise its implementation. Since bleaching of fluorescent molecules involves temperature-dependent chemical reactions, it is likely that low temperatures could slow down this process. We stored auramine-stained slides under different environmental conditions, including -20°C, and examined them over time. The slides stored in all the environments faded. At -20°C, fading was not reduced in relation to room temperature. Restaining and re-examining smears after five months showed that the slides containing saliva and storage at -20°C were associated with failure in AFB reappearance. In conclusion, the practice of freezing slides until they are viewed should be discouraged as it has a negative effect on blinded rechecking by reducing reading concordance after restaining. Specimen quality should be considered when interpreting FM-EQA results.

MRSA Pediatric clone expressing ermC plus lnuA genes causing nosocomial transmission and healthcare workers colonization in a neonatal intensive care unit.

Methicillin-resistant Staphylococcus aureus (MRSA) is a major cause of both nosocomial and community-acquired infections. We describe an outbreak caused by the MRSA Pediatric clone expressing an unusual lincosamide resistant phenotype. Between January and May 2006, an MRSA outbreak was detected at the Neonatal Unit of Hospital Interzonal General de Agudos "Evita", Buenos Aires Province, Argentina that affected ten patients. Seven isolates from seven patients plus five MRSA recovered from health care workers (nasal carriage) were studied. Two phenotypes were observed: (i) ELCi (10), resistance to erythromycin and lincomycin and inducible resistance to clindamycin; (ii) ELiCi (2), resistance to erythromycin and inducible resistance to lincomycin and clindamycin. All 12 MRSA were resistant to oxacillin, erythromycin and gentamicin. Isolates expressing the ELCi-phenotype showed lincomycin MIC values between 16 and 32mg/L, while the remaining 2 isolates with ELiCi-phenotype presented a MIC value of 0.5mg/L. No differences were observed between the clindamycin MIC values in both phenotypes, ranging 0.25-0.5mg/L. Isolates showing ELCi-phenotype harbored ermC plus lnuA genes, and the other two only ermC gene. All 12 isolates were genetically related and belonged to the Pediatric clone (ST100) harboring a new variant of SCCmecIV. This is the first MRSA outbreak expressing an unusual ELCi phenotype due to a combination of ermC plus lnuA genes.

[Dissemination of blaIMP-8 among Enterobacteriaceae isolates from a Buenos Aires Hospital]. / Diseminación de blaIMP-8 en enterobacterias aisladas en un hospital de Buenos Aires.

From August 2008 to December 2011, six metallo-ß-lactamase-producing isolates, four Enterobacter cloacae, one Klebsiella oxytoca and one Citrobacter freundii, were detected at Hospital Interzonal General de Agudos "Evita" in Lanús. All six isolates showed multiresistant profiles and the presence of the blaIMP-8 gene. Five isolates also expressed PER-2 extended spectrum ß-lactamase. The blaIMP-8 gene was found as the first cassette in a class 1 integron. However, the 3´ conserved sequence could not be detected in three isolates. In all cases, blaIMP-8 was transferred by conjugation to azide-resistant Escherichia coli J53. PFGE analysis revealed that the four E. cloacae isolates were not genetically related. These are the first metallo-ß-lactamases detected in this institution and our results suggest a possible intra- and inter-species horizontal dissemination of blaIMP-8.

Diseminación de blaIMP-8 en enterobacterias aisladas en un hospital de Buenos Aires / [Dissemination of blaIMP-8 among Enterobacteriaceae isolates from a Buenos Aires Hospital].

From August 2008 to December 2011, six metallo-ß-lactamase-producing isolates, four Enterobacter cloacae, one Klebsiella oxytoca and one Citrobacter freundii, were detected at Hospital Interzonal General de Agudos "Evita" in Lanús. All six isolates showed multiresistant profiles and the presence of the blaIMP-8 gene. Five isolates also expressed PER-2 extended spectrum ß-lactamase. The blaIMP-8 gene was found as the first cassette in a class 1 integron. However, the 3´ conserved sequence could not be detected in three isolates. In all cases, blaIMP-8 was transferred by conjugation to azide-resistant Escherichia coli J53. PFGE analysis revealed that the four E. cloacae isolates were not genetically related. These are the first metallo-ß-lactamases detected in this institution and our results suggest a possible intra- and inter-species horizontal dissemination of blaIMP-8.

Outbreak of a cluster with epidemic behavior due to Serratia marcescens after colistin administration in a hospital setting.

Serratia marcescens causes health care-associated infections with important morbidity and mortality. Particularly, outbreaks produced by multidrug-resistant isolates of this species, which is already naturally resistant to several antibiotics, including colistin, are usually described with high rates of fatal outcomes throughout the world. Thus, it is important to survey factors associated with increasing frequency and/or emergence of multidrug-resistant S. marcescens nosocomial infections. We report the investigation and control of an outbreak with 40% mortality due to multidrug-resistant S. marcescens infections that happened from November 2007 to April 2008 after treatment with colistin for Acinetobacter baumannii meningitis was started at hospital H1 in 2005. Since that year, the epidemiological pattern of frequently recovered species has changed, with an increase of S. marcescens and Proteus mirabilis infections in 2006 in concordance with a significant decrease of the numbers of P. aeruginosa and A. baumannii isolates. A single pulsed-field gel electrophoresis (PFGE) cluster of S. marcescens isolates was identified during the outbreak. When this cluster was compared with S. marcescens strains (n = 21) from 10 other hospitals (1997 to 2010), it was also identified in both sporadic and outbreak isolates circulating in 4 hospitals in Argentina. In132::ISCR1::blaCTX-M-2 was associated with the multidrug-resistant cluster with epidemic behavior when isolated from outbreaks. Standard infection control interventions interrupted transmission of this cluster even when treatment with colistin continued in several wards of hospital H1 until now. Optimizing use of colistin should be achieved simultaneously with improved infection control to prevent the emergence of species naturally resistant to colistin, such as S. marcescens and P. mirabilis.

Diseminación de blaIMP-8 en enterobacterias aisladas en un hospital de Buenos Aires. / [Dissemination of blaIMP-8 among Enterobacteriaceae isolates from a Buenos Aires Hospital].

From August 2008 to December 2011, six metallo-ß-lactamase-producing isolates, four Enterobacter cloacae, one Klebsiella oxytoca and one Citrobacter freundii, were detected at Hospital Interzonal General de Agudos "Evita" in Lanús. All six isolates showed multiresistant profiles and the presence of the blaIMP-8 gene. Five isolates also expressed PER-2 extended spectrum ß-lactamase. The blaIMP-8 gene was found as the first cassette in a class 1 integron. However, the 3´ conserved sequence could not be detected in three isolates. In all cases, blaIMP-8 was transferred by conjugation to azide-resistant Escherichia coli J53. PFGE analysis revealed that the four E. cloacae isolates were not genetically related. These are the first metallo-ß-lactamases detected in this institution and our results suggest a possible intra- and inter-species horizontal dissemination of blaIMP-8.