How a discerning cytological examination can aid in the diagnosis of infectious diseases: case reports

Infections caused by uncommon and resistant pathogens in unusual sites have been increasingly reported in medical literature. We describe four cases of rare cytological findings and clinical impact for patients. In the first case, Aspergillus sp and Pneumocystis jirovecii were observed in the bronchoalveolar lavage of a patient with severe systemic lupus. In the second and third cases, we describe the presence of Trichomonas sp and Strongyloides sp larvae in samples of pleural and peritoneal fluid, respectively. The fourth report is about a patient with a wrist subcutaneous nodule whose synovial aspiration and cytology revealed the presence of brown septate hyphae. The early identification of the infectious agent in the cytological examination was essential for the introduction and/or re-adaptation of therapy in the four cases described. Patients in this report were immunocompromised with severe comorbidities, conditions often associated with unfavorable clinical outcomes.

Genetic diversity of Mycoplasma hyopneumoniae in Mendoza province / Diversidad genética de Mycoplasma hyopneumoniae en la provincia de Mendoza

En Argentina, la neumonía enzoótica porcina (NEP) es altamente prevalente y se han identificado diferentes tipos genéticos de Mycoplasma hyopneumoniae. Sin embargo, se carece de información acerca de la prevalencia de NEP y de otros aspectos epidemiológicos de esta entidad en la provincia de Mendoza. En esta investigación se usó un análisis multilocus de regiones repetidas en tándem (MLVA) de los loci P97 R1, P97 R1A y P146 R3 para evaluar la diversidad genética de M. hyopneumoniae a partir de muestras clínicas de cerdos de cinco granjas localizadas en diferentes distritos de la provincia de Mendoza. M. hyopneumoniae pudo ser tipificado a partir de 27 muestras de lavado broncoalveolar (LBA) y se identificaron 8 diferentes MLVA-tipos. Este es el primer informe acerca de la diversidad genética de M. hyopneumoniae en Mendoza. Los resultados obtenidos permiten describir de manera más acabada la diversidad genética de este agente en nuestro país.

Accuracy of a rapid molecular test for tuberculosis in sputum samples, bronchoalveolar lavage fluid, and tracheal aspirate obtained from patients with suspected pulmonary tuberculosis at a tertiary referral hospital / Acurácia do teste rápido molecular para tuberculose em amostras de escarro, lavado broncoalveolar e aspirado traqueal obtidos de pacientes com suspeita de tuberculose pulmonar em um hospital de referência terciária

ABSTRACT Tuberculosis continues to be a major public health problem worldwide. The aim of the present study was to evaluate the accuracy of the Xpert MTB/RIF rapid molecular test for tuberculosis, using pulmonary samples obtained from patients treated at the Júlia Kubitschek Hospital, which is operated by the Hospital Foundation of the State of Minas Gerais, in the city of Belo Horizonte, Brazil. This was a retrospective study comparing the Xpert MTB/RIF test results with those of standard culture for Mycobacterium tuberculosis and phenotypic susceptibility tests. Although the Xpert MTB/RIF test showed high accuracy for the detection of M. tuberculosis and its resistance to rifampin, attention must be given to the clinical status of the patient, in relation to the test results, as well as to the limitations of molecular tests.

RESUMO A tuberculose permanece como um grave problema de saúde pública. O objetivo deste estudo foi avaliar a acurácia do teste rápido molecular Xpert MTB/RIF em amostras pulmonares no Hospital Júlia Kubitschek, Fundação Hospitalar do Estado de Minas Gerais, localizado em Belo Horizonte (MG). Trata-se de um estudo descritivo retrospectivo, considerando-se como método padrão a cultura para o bacilo da tuberculose e o teste de sensibilidade fenotípico. O teste Xpert MTB/RIF apresentou ótima acurácia para a detecção da tuberculose e resistência à rifampicina, mas é necessária a atenção a dados clínicos do paciente em relação ao resultado do exame e às limitações dos testes moleculares.

Optimización y validación de un método de PCR en tiempo real para el diagnóstico de Pneumocystis jirovecii en muestras respiratorias de pacientes pediátricos / Optimization and validation of a real-time PCR method for the diagnosis of Pneumocystis jirovecii in respiratory samples of pediatric patients

Pneumocystis jirovecii (PCP) es un hongo oportunista, que causa neumonía en pacientes con un sistema inmunitario seriamente comprometido. La prevalencia de la enfermedad disminuyó dramáticamente con la introducción de la terapia antirretroviral combinada (HAART) y actualmente las infecciones ocurren en aquellos pacientes que no reciben adecuada profilaxis o no la completan. Es una enfermedad grave con elevada tasa de mortalidad (30-60%) en pacientes oncohematológicos y receptores de trasplante de células progenitoras hematopoyéticas (HSCT), pero prevenible con profilaxis adecuada, por lo que el reconocimiento temprano de los pacientes en riesgo es crítico. El diagnóstico de certeza es de laboratorio ya que los hallazgos clínicos son inespecíficos y las imágenes no son patognomónicas de este agente. Actualmente las técnicas moleculares como la PCR en tiempo real son las metodologías recomendadas ya que poseen elevada sensibilidad, especificidad, rapidez y eficiencia. En el presente estudio se optimizó un método de PCR en tiempo real con iniciadores dirigidos al gen del ARNr de la subunidad grande mitocondrial, en formato dúplex junto con el gen constitutivo RNAsa P. El método demostró ser muy sensible y rápido para el diagnóstico clínico de PCP, con una concordancia қ: 0,789 con el método convencional de PCR anidada que emplea como target a la región espaciadora transcrita interna (ITS) del gen del ARNr de PCP, a la vez de ser mucho menos laborioso y con menor riesgo de contaminación, lo que permite el manejo de un alto número de muestras clínicas (AU)

Pneumocystis jirovecii (PCP) is an opportunistic fungus causing pneumonia in severely immunocompromised patients. Prevalence of the disease has dramatically decreased after the introduction of combined antiretroviral therapy (HAART) and currently these infections occur in patients who do not receive adequate prophylaxis or do not complete treatment. PCP is a severe disease with a high mortality rate (30-60%) in oncology-hematology patients and hematopoietic stem-cell transplantation (HSCT) recipients, but is preventable with adequate prophylaxis. Therefore, early recognition of at-risk patients is essential. Laboratory studies are the gold standard for the diagnosis as clinical findings are unspecific and imaging studies are not pathognomonic for this agent. Currently, molecular techniques, such as real-time PCR, are the methodology of choice because of their high sensitivity, specificity, speed, and efficiency. In this study, a real-time PCR method was optimized with primers targeting the gene of the mitochondrial large subunit rRNA in a duplex format together with the constitutive gene RNAsa P. The method showed to be very sensitive and fast for the clinical diagnosis of PCP, with a concordance of қ: 0.789 with the conventional nested PCR method targeting the internal transcribed spacer (ITS) region of the rRNA gene of PCP, and is much easier to perform with a lower contamination risk allowing a high through-put of clinical samples (AU)

Indications, clinical outcomes and complications of 1, 949 flexible bronchoscopies / Indicações, resultados clínicos e complicações de 1. 949 broncoscopias flexíveis

ABSTRACT Objective To describe indications, clinical outcomes and complications of flexible bronchoscopy. Methods A descriptive observational study of bronchoscopies performed at the endoscopy service of Hospital Israelita Albert Einstein . Demographic (age, gender and origin) and medical (indications and results of endoscopy and diagnostic tests, such as biopsy collection, lavage, cytology and culture) data were analyzed. Electronic medical records with incomplete data or reporting interventional procedures were excluded. Results Over a three-year period (2013 to 2016), a total of 1,949 bronchoscopies were performed by respiratory endoscopy team and anesthesia specialists of the hospital. The mean age of patients was 57.7±21.9 years (range of 3 days to 99 years), with prevalence of males (56.4%). The procedures were mostly (86.3%) elective and 30.7% were carried out in the intensive care unit. Major indications for bronchoscopy were infection or secretion (42.4%), followed by suspected neoplasm (10.8%). Endoscopic changes were reported in 91.9% of cases, with more than one change described in approximately 6.9% of patients. Positive results were obtained via direct testing or culture in 36.3% and 53.9% of 1,399 bronchoalveolar lavages, respectively. The overall diagnostic yield (bronchoalveolar lavage and biopsy) was 72.6%. Mild adverse event rate was 7.2%. The rate of severe adverse events requiring additional intervention was 0.5% (pneumothorax, 0.4%; severe bleeding with patient death, 0.1%). Conclusion Lower airway endoscopy is critical for respiratory disease assessment, diagnosis and treatment. Flexible bronchoscopy is associated with good diagnostic yield and minimal inherent risk.

RESUMO Objetivo Descrever as indicações, os resultados clínicos e as complicações associadas à broncoscopia flexível. Métodos Foi realizado um estudo observacional descritivo das broncoscopias realizadas no serviço de endoscopia do Hospital Albert Einstein. Foram analisados: informações demográficas, como idade, gênero e procedência; dados clínicos sobre a indicação do exame; e resultados endoscópicos e dos exames diagnósticos realizados, como biópsias, lavados, citologias e culturas. Os fatores de exclusão foram dados incompletos no sistema e procedimentos intervencionistas. Resultados No período de 3 anos, de 2013 a 2016, foram realizadas 1.949 broncoscopias no hospital pela equipe da endoscopia respiratória e anestesiologista. A média de idade dos pacientes foi de 57,7±21,9 anos, (variação: 3 dias a 99 anos), com prevalência do gênero masculino (56,4%). A maioria dos exames (86,3%) foi eletiva e 30,7% foram realizados na terapia intensiva. A indicação médica mais frequente para realização da broncoscopia foi infecção ou secreção (42,4%), seguida de suspeita de neoplasia (10,8%). Nas alterações endoscópicas das broncoscopias realizadas, obtivemos os dados em 91,9% dos exames. Cerca de 6,9% dos pacientes apresentaram mais de uma alteração endoscópica. Foram realizados 1.399 lavados, com positividade de 36,3% nas pesquisas diretas e 53,9% nas culturas. O rendimento geral com lavados broncoalveolares e biópsia foi de 72,6%. Em nossa série, a taxa de eventos adversos leves foi de 7,2%. Os eventos adversos graves, nos quais foi necessária alguma intervenção adicional, somaram 0,5%: 0,4% de pneumotórax e 0,1% de hemorragia grave com óbito. Conclusão A endoscopia das vias aéreas inferiores é imprescindível para avaliação, diagnóstico e tratamento de doenças respiratórias. A broncoscopia flexível tem bom rendimento diagnóstico e risco mínimo associado.

Preanalytical conditions can interfere with M. tuberculosis detection by PCR in respiratory samples

OBJECTIVES: Tuberculosis is one of the most prevalent infections in humans. Although culture is the reference for diagnosis, its sensitivity is compromised, especially in paucibacillary samples. Because polymerase chain reaction (PCR) amplifies mycobacterial DNA, it is more sensitive than culture for the diagnosis of Mycobacterium tuberculosis (Mtb). However, its performance can be affected by intrinsic sample inhibitors and by the extraction/detection techniques used. METHODS: We evaluated the influence of preanalytical conditions on Mtb detection in samples of sputum (SPU), bronchoalveolar lavage (BAL), and pleural fluid (PF) using combinations of extraction/detection methods. Respiratory samples were prepared to contain different concentrations of red blood cells and nucleated cells to which increasing amounts of Mtb colonies were inoculated and submitted to PCR. RESULTS: Up to 102 CFU/ml of Mtb were detected in the SPU in all methods, except for the Roche extraction/detection method, regardless of the preanalytical sample condition. In BAL samples, medium and high concentrations of cells and high concentrations of red blood cells contributed to a lower Mtb detection, regardless of the extraction method used. In PF, red blood cells were the variable that most interfered with Mtb detection, with better recovery (102 CFU/ml) observed with the Qiagen/Nanogen combination. CONCLUSION: The choice of Mtb extraction and detection method is of fundamental importance for PCR analytical sensitivity, especially when paucibacillary samples and/or samples containing potential PCR inhibitors are analyzed.

Evaluación del Aspergillus lateral flow device para el diagnóstico de aspergilosis invasora, experiencia en un hospital universitario / Evaluation of the lateral flow Aspergillus assay for the diagnosis of invasive aspergillosis, experience in a university hospital

Resumen Introducción: El diagnóstico de aspergilosis invasora (AI) se realiza mediante criterios clínicos y microbiológicos los que incluyen marcadores séricos. Recientemente, el test inmunocromatográfico Aspergillus lateral flow device (LFD), ha sido evaluado como método para diagnóstico de AI. Objetivo: Evaluar el desempeño de este test para el diagnóstico de AI. Material y Método: Estudio transversal en que se evaluaron muestras de suero y lavado bronco-alveolar (LBA) procesadas para galactomanano provenientes de pacientes adultos con sospecha de AI, atendidos en el Hospital Clínico de Red de Salud UCCHRISTUS. Resultados: Se procesó un total de 142 muestras de 98 pacientes, correspondientes a AI probada 5,6%, AI probable 41,5%, AI posible 12,7% y ausencia de AI 40,1%. Al confrontar los resultados con las categorías diagnósticas según criterios EORTC/MSG se obtuvo una sensibilidad y especificidad de LFD para diagnóstico de AI de 70,9 y 53,5% para muestras de suero y 83,3 y 38,5% para muestras de LBA. La concordancia entre galactomanano y LFD fue de 62,4% (54,1-69,9) con un índice Kappa de 0,202 (0,03682-0,3669). Conclusiones: Aspergillus LFD presentó una adecuada sensibilidad; sin embargo, la especificidad fue baja por lo que un resultado positivo requiere ser confirmado.

Background: The incidence of invasive aspergillosis is increasing. Its diagnosis is based on clinical and microbiological criteria which include the determination of serological markers such as galactomannan. Recently, the Aspergillus lateral flow device, an inmunocromatograph assay has been described for its diagnosis. Aim: To evaluate the performance of the lateral flow device for the diagnosis of invasive aspergillosis (IA) in adult patients. Material and Method: In this cross-sectional study, frozen samples that had been previously evaluated for galactomannan from patients classified with proven/probable/possible or no AI according to the EORTC/MSG criteria were selected. Results: A total of 142 samples from 98 patients were processed, corresponding to proven AI 5.6%, probable IA 41.5%, possible IA 12.7% and no-IA 40.1%. The sensitivity and specificity of the Aspergillus lateral flow was 70.9% and 53.5% for serum samples and 83.3% and 38.5% for BAL samples. The concordance between the galactomannan and Aspergillus lateral flow was 62.4% (54.1 - 69.9) with a Kappa index of 0.202 (0.03682 - 0.3669). Conclusions: We observed a good sensitivity but low specificity, a positive result need a confirmatory test.

Análise da produção de biofilme por isolados clínicos de Pseudomonas aeruginosa de pacientes com pneumonia associada à ventilação mecânica / Analysis of biofilm production by clinical isolates of Pseudomonas aeruginosa from patients with ventilator-associated pneumonia

RESUMO Objetivo: Avaliar fenotipicamente a produção de biofilme por isolados clínicos de Pseudomonas aeruginosa de pacientes com pneumonia associada à ventilação mecânica. Métodos: Foram analisados 20 isolados clínicos de P. aeruginosa, sendo 19 provenientes de amostras clínicas de aspirado traqueal e uma de lavado broncoalveolar. A avaliação da capacidade de P. aeruginosa em produzir biofilme foi verificada por duas técnicas, sendo uma qualitativa e outra quantitativa. Resultados: A técnica qualitativa mostrou que apenas 15% dos isolados foram considerados produtores de biofilme, enquanto que a quantitativa demonstrou que 75% dos isolados foram produtores de biofilme. Os isolados produtores de biofilme apresentaram o seguinte perfil de suscetibilidade: 53,3% eram multidroga-resistentes e 46,7% eram multidroga-sensíveis. Conclusão: A técnica quantitativa foi mais eficaz para detecção da produção de biofilme em comparação com a qualitativa. Para a população bacteriana analisada, a produção de biofilme independeu do perfil de suscetibilidade das bactérias, demonstrando que a falha terapêutica pode estar relacionada com a produção de biofilme, por impedir a destruição das bactérias presentes nesta estrutura, ocasionando complicações da pneumonia associada à ventilação mecânica, incluindo infecções extrapulmonares, e dificultando o tratamento da infecção.

ABSTRACT Objective: To phenotypically evaluate biofilm production by Pseudomonas aeruginosa clinically isolated from patients with ventilator-associated pneumonia. Methods: Twenty clinical isolates of P. aeruginosa were analyzed, 19 of which were from clinical samples of tracheal aspirate, and one was from a bronchoalveolar lavage sample. The evaluation of the capacity of P. aeruginosa to produce biofilm was verified using two techniques, one qualitative and the other quantitative. Results: The qualitative technique showed that only 15% of the isolates were considered biofilm producers, while the quantitative technique showed that 75% of the isolates were biofilm producers. The biofilm isolates presented the following susceptibility profile: 53.3% were multidrug-resistant, and 46.7% were multidrug-sensitive. Conclusion: The quantitative technique was more effective than the qualitative technique for the detection of biofilm production. For the bacterial population analyzed, biofilm production was independent of the susceptibility profile of the bacteria, demonstrating that the therapeutic failure could be related to biofilm production, as it prevented the destruction of the bacteria present in this structure, causing complications of pneumonia associated with mechanical ventilation, including extrapulmonary infections, and making it difficult to treat the infection.

Rapid detection of Candida species in bronchoalveolar lavage fluid from patients with pulmonary symptoms

Abstract Candida species, especially C. albicans, are commensals on human mucosal surfaces, but are increasingly becoming one of the important invasive pathogens as seen by a rise in its prevalence in immunocompromised patients and in antibiotic consumption. Thus, an accurate identification of Candida species in patients with pulmonary symptoms can provide important information for effective treatment. A total of 75 clinical isolates of Candida species were obtained from the bronchoalveolar lavage fluid of both immunocompromised and immunocompetent patients with pulmonary symptoms. Candida cultures were identified based on nuclear ribosomal Internal Transcribed Spacer (ITS1-ITS2 rDNA) sequence analysis by polymerase chain reaction–restriction fragment length polymorphisms (PCR-RFLP). Molecular identification indicated that the isolates belonged predominantly to C. albicans (52%), followed by C. tropicalis (24%), C. glabrata (14.7%), C. krusei (5.3%), C. parapsilosis (1.3%), C. kefyr (1.3%) and C. guilliermondii (1.3%). Given the increasing complexity of disease profiles and their management regimens in diverse patients, rapid and accurate identification of Candida species can lead to timely and appropriate antifungal therapy.

Infección pulmonar no invasora por Scedosporium apiospermum / Pulmonary non invasive infection by Scedosporium apiospermum

We reported a case of non-invasive pulmonary infection by Scedosporium apiospermum in 67 years old female with bronchiectasis and caverns secondary to tuberculosis. Diagnosis was made with lung CT and bronchial lavage cultures. The patient was initially treated with itraconazole for six weeks without success and then voriconazole for 16 weeks, with good clinical response.

Reportamos el caso clínico de una infección pulmonar no invasora por Scedosporium apiospermum en una mujer de 67 años de edad, con bronquiectasias y cavernas pulmonares secundarias a una tuberculosis. El diagnóstico se realizó con la TAC pulmonar y cultivos de lavado bronquial. La paciente fue tratada inicialmente con itraconazol oral por seis semanas sin respuesta y luego voriconazol vía oral por 16 semanas, con una buena respuesta clínica.

Quantitative culture of endotracheal aspirate and BAL fluid samples in the management of patients with ventilator-associated pneumonia: a randomized clinical trial / Cultura quantitativa de aspirado traqueal e lavado broncoalveolar no manejo de pacientes com pneumonia associada à ventilação mecânica: um ensaio clínico randomizado

OBJECTIVE: To compare 28-day mortality rates and clinical outcomes in ICU patients with ventilator-associated pneumonia according to the diagnostic strategy used. METHODS: This was a prospective randomized clinical trial. Of the 73 patients included in the study, 36 and 37 were randomized to undergo BAL or endotracheal aspiration (EA), respectively. Antibiotic therapy was based on guidelines and was adjusted according to the results of quantitative cultures. RESULTS: The 28-day mortality rate was similar in the BAL and EA groups (25.0% and 37.8%, respectively; p = 0.353). There were no differences between the groups regarding the duration of mechanical ventilation, antibiotic therapy, secondary complications, VAP recurrence, or length of ICU and hospital stay. Initial antibiotic therapy was deemed appropriate in 28 (77.8%) and 30 (83.3%) of the patients in the BAL and EA groups, respectively (p = 0.551). The 28-day mortality rate was not associated with the appropriateness of initial therapy in the BAL and EA groups (appropriate therapy: 35.7% vs. 43.3%; p = 0.553; and inappropriate therapy: 62.5% vs. 50.0%; p = 1.000). Previous use of antibiotics did not affect the culture yield in the EA or BAL group (p = 0.130 and p = 0.484, respectively). CONCLUSIONS: In the context of this study, the management of VAP patients, based on the results of quantitative endotracheal aspirate cultures, led to similar clinical outcomes to those obtained with the results of quantitative BAL fluid cultures. .

OBJETIVO: Comparar a mortalidade em 28 dias e desfechos clínicos em pacientes com pneumonia associada à ventilação mecânica (PAVM) internados em UTI conforme a estratégia diagnóstica utilizada. MÉTODOS: Ensaio clínico randomizado prospectivo. Dos 73 pacientes incluídos no estudo, 36 e 37, respectivamente, foram randomizados para a realização de LBA ou aspiração traqueal (AT). A antibioticoterapia inicial baseou-se em diretrizes e foi ajustada de acordo com os resultados das culturas quantitativas. RESULTADOS: A taxa de mortalidade em 28 dias foi semelhante nos grupos LBA e AT (25,0% e 37,8%, respectivamente; p = 0,353). Não houve diferenças entre os grupos em relação a duração da ventilação mecânica, antibioticoterapia, complicações secundárias, recidiva de PAVM ou tempo de permanência hospitalar e na UTI. A antibioticoterapia inicial foi considerada adequada em 28 (77,8%) e 30 (83,3%) dos pacientes nos grupos LBA e AT, respectivamente (p = 0,551). A mortalidade em 28 dias não se associou com a adequação da antibioticoterapia inicial nos grupos LBA e AT (tratamento apropriado: 35,7% vs. 43,3%; p = 0,553; e tratamento inapropriado: 62,5% vs. 50,0%; p = 1,000). O uso prévio de antibióticos não interferiu no rendimento das culturas nos grupos AT e LBA (p = 0,130 e p = 0,484, respectivamente). CONCLUSÕES: No contexto deste estudo, o manejo dos pacientes com PAVM, baseado nos resultados da cultura quantitativa do aspirado traqueal, resultou em desfechos clínicos semelhantes aos obtidos com os resultados da cultura quantitativa do LBA. (Registro Brasileiro de Ensaios ...

Antibiotic Sensitivity Pattern of Gram Negative Bacilli Isolated from the Lower Respiratory Tract of Ventilated Patients in the Intensive Care Unit.

The causative organisms vary according to the patients’ demographics in the ICU, methods of diagnosis, the durations of hospital and ICU stay. It is necessary to study different etiological organisms and their antimicrobial susceptibility pattern for generating local antibiotic policy. Aims: To study the causative organisms and determine the antibiotic susceptibility pattern of the lower respiratory tract isolates from patients admitted to ICU. Methods and Material: Endotracheal aspirates from 200 patients admitted to the ICU were cultured, identified and antimicrobial susceptibility testing was performed by standard methods. Results: From 200 specimens, 69(34.5%) were culture positive. Total 96 isolates were recovered, from these 92 (96.87%) isolates were gram negative bacilli (GNB). In 34.78% specimens, two isolates were recovered. The most common Gram- negative organism being Acinetobacter spp. (31.25%) followed by Klebsiella spp. (21.87%), E-coli (21.87%) and Pseudomonas Spp. (17.7%). All GNBs were 100% sensitive to polymyxin B and colistin and resistant to piperacillin, ceftazidime and cotrimoxazole. 50% E-coli and 38% of Klebsiella pneumoniae strains were ESBL (extendedspectrum b-lactamase) producers. Conclusions: This study demonstrates the trend in antimicrobial susceptibility pattern of gram negative bacilli in intensive care unit. It is the most important for specific treatment of ventilator associated pneumonia patients and to generate local data periodically to decide empiric antimicrobial therapy.

Stenotrophomonas maltophilia pseudo-outbreak at a University Hospital Bronchoscopy unit in Turkey / Pseudo-brote de Stenotrophomonas maltophilia en una unidad de Broncoscopia del Hospital Universitario en Turquía

OBJECTIVE: Stenotrophomonas maltophilia is an opportunistic pathogen found predominantly in the enviroment and hospital setting. Invasive procedures and treatment methods, instruments used for diagnosis and irrational antibiotic use play major roles in the spread of this pathogen. The study aimed to evaluate consecutive S maltophilia isolation from bronchoalveolar lavage samples during bronchoscopy procedure during a week. METHODS: Four patients consecutively had S maltophilia isolated during bronchoscopy between September 8 and 15, 2012. The identification of the isolates and their antibiotic susceptibility were studied by automated Vitek version 2.0 (Biomerieux, France) system. The clonal relationship between the isolates was studied by enterobacterial repetitive intergenic consensus (ERIC) polymerase chain reaction (PCR). RESULTS: Four consecutive S maltophilia isolates had identical band patterns and showed clonal relatedness. CONCLUSION: Bronchoscopy is a common invasive procedure that is utilized in chest diseases departments and intensive care units (ICUs). Contamination may take place due to inappropiate use and cause spread of infectious pathogens. In the current study, we detected consecutive S maltophilia strains with identical band patterns isolated within a week. After appropiate disinfection and cleaning procedures, no further isolation was detected.

OBJETIVO: Stenotrophomonas maltophilia es un patógeno oportunista que se encuentra predominantemente en el medio ambiente y entorno de los hospitales. Los procedimientos invasivos y los métodos de tratamiento, los instrumentos utilizados para el diagnóstico y tratamiento, así como el uso irracional de antibióticos, desempeñan un importante papel en la propagación de este patógeno. Este estudio persigue evaluar el aislamiento consecutivo de S maltophilia de las muestras de lavado broncoalveolar durante el procedimiento broncoscópico en el período de una semana. MÉTODOS: A cuatro pacientes se les aisló S maltophilia consecutivamente en broncoscopias realizadas entre el 8 y el 15 de septiembre de 2012. La identificación de los aislamientos y su sensibilidad a los antibióticos fueron estudiados por el sistema automatizado Vitek 2 (Biomerieux, Francia). La relación clonal entre los aislamientos fue estudiada mediante consenso intergénico repetitivo enterobacteriano (ERIC) en conjunción con la reacción en cadena de la polimerasa (PCR). RESULTADOS: Cuatro aislados consecutivos de S maltophilia tenían patrones de banda idénticos y exhibían conexidad clonal. CONCLUSIÓN: La broncoscopia es un procedimiento invasivo común que se aplica en los departamentos de enfermedades torácicas, y las unidades de cuidados intensivos (UCI). La contaminación puede ocurrir debido a usos inapropiados y a la propagación de agentes patógenos infecciosos. En el presente estudio, hemos detectado cepas de S maltophilia consecutivas con idénticos patrones de banda aislados en una semana. Después de los procedimientos de limpieza y desinfección adecuada, no se detectó ningún otro aislamiento.

Assessment of the Quantitative Ability of AdvanSure TB/NTM Real-Time PCR in Respiratory Specimens by Comparison with Phenotypic Methods

Accurate quantification of mycobacterial load is important to evaluate disease severity and to monitor the course of treatment in tuberculosis (TB). We evaluated the quantitative capability of the AdvanSure TB/NTM real-time PCR kit (LG Life Science, Korea) to determine the cycle threshold (Ct) for mycobacterial burden. We retrospectively analyzed data from 108 patients whose respiratory specimens (sputums and bronchoalveolar lavage fluids) were positive for Mycobacterium tuberculosis complex (85 culture-positive and 23 culture-negative specimens). We compared Ct values with grades of acid-fast bacilli (AFB) staining, semi-quantitative colony count on solid medium, and time to positivity (TTP) in liquid and solid media. We also investigated the cutoff Ct value for predicting stain-positive status. Ct value showed significant reverse correlation with AFB staining grade (r(s)=-0.635, P<0.01). Ct value significantly decreased as the semi-quantitative counts on the solid medium increased (P<0.001), and the mean Ct value of each of the groups 1+, 2+, 3+, and 4+ were 29.0, 30.0, 27.1, and 25.5, respectively. A weak correlation between Ct value and TTP in liquid and solid media was observed (r(s)=0.468 and 0.365, respectively). A cutoff Ct value of <33.2 best predicted stain positivity, with a sensitivity of 95.0% and a specificity of 32.0%. Our findings suggest the potential use of AdvanSure TB/NTM real-time PCR kit for quantitatively determining bacterial burden, albeit with some enhancements.

Recent Trends in Clinically Significant Nontuberculous Mycobacteria Isolates at a Korean General Hospital

Lung disease caused by nontuberculous mycobacteria (NTM) represents an increasing proportion of all mycobacterial diseases. We investigated recent occurrences of NTM and evaluated the clinical significance of NTM isolates from 752 respiratory specimens collected from patients at National Health Insurance Service Ilsan Hospital between January 2007 and May 2011. Specimens were incubated on solid and liquid media (BACTEC MGIT 960, BD, USA) for 6-8 weeks, and PCR and reverse blot hybridization were performed (REBA Myco-ID, Molecules & Diagnostics, Korea). Clinical features of the patients were reviewed through medical records. The most frequently isolated organism was Mycobacterium avium (46.7%), followed by M. intracellulare (14.8%), M. fortuitum (7.2%), and M. abscessus (6.6%). The most common mycobacteria among definitive cases of NTM lung disease were M. avium (42/351, 12.0%), M. intracellulare (19/111, 17.1%), M. abscessus (11/50, 22.0%), M. massiliense (4/13, 30.8%), and M. fortuitum (4/54, 7.4%). Clinically significant cases of NTM lung disease increased from 4 patients in 2007 to 32 in 2011. The mean patient age was 64 yr (range: 35-88 yr), and 58 (64%) patients were women. Patients suffered from cough, productive sputum, and hemoptysis. In summary, the most common mycobacteria causing NTM lung disease were M. avium and M. intracellulare; however, cases of M. massiliense and M. abscessus infection are on the rise in Korea.

Co-existence of HIV, active tuberculosis and aspergilloma in a single individual - A case report.

Summary: Tuberculosis (TB) is a disease as old as mankind, whereas in India the first case of Human Immunodeficiency Virus (HIV) was reported in 1986. HIV and TB are so closely connected that their relationship is often described as a coepidemic. Aspergilloma (Fungal Ball, Mycetoma) represents a saprophytic growth of aspergillus that colonizes in the preformed cavities commonly due to pulmonary tuberculosis (PTB). We report a case of HIV, active pulmonary tuberculosis and aspergilloma occurring in the same patient. Despite our best efforts, we could not lay our hands on any similar case in the medical literature.

The Combination of Real-Time PCR and HPLC for the Identification of Non-Tuberculous Mycobacteria

We used HPLC and AdvanSure real-time PCR (LG Life Sciences, Korea) to retrospectively analyze non-tuberculous mycobacteria (NTM) in 133 clinical specimens. The specimens were culture-positive for NTM and the HPLC method identified 130 strains of mycobacteria from the cultures (97.7%) at the species level. Among the isolates, 48 Mycobacterium. kansasii (36.1%), 39 M. intracellulare (29.3%), 17 M. avium (12.8%), 16 M. abscessus (12.0%), 6 M. fortuitum (4.5%), 2 M. szulgai (1.5%), 2 M. gordonae (1.5%), and 3 unclassified NTM strains (2.3%) were identified. The real-time PCR assay identified 60 NTM-positive specimens (45.1%), 65 negative specimens (48.9%), and 8 M. tuberculosis (TB)-positive specimens (6.0%). The real-time PCR assay is advantageous because of its rapid identification of NTM. However, in our study, the real-time PCR assay showed relatively low sensitivity (45.1%) when using direct specimens including sputum and bronchoalveolar lavage (BAL) fluid. HPLC is useful as it discriminates NTM at the species level, although it is time-consuming and requires specific equipment and technical expertise. A combination of both methods will be helpful for the rapid and accurate identification of mycobacteria in clinical laboratories.

Rothia mucilaginosa Pneumonia Diagnosed by Quantitative Cultures and Intracellular Organisms of Bronchoalveolar Lavage in a Lymphoma Patient

Rothia mucilaginosa is a gram-positive coccus of the family Micrococcaceae. R. mucilaginosa is considered a part of the normal flora of the human oropharynx and upper respiratory tract and lower respiratory tract infections attributable to R. mucilaginosa are not frequent. We present a case of pneumonia, in which the R. mucilaginosa infection was diagnosed by quantitative cultures of a bronchoalveolar lavage (BAL) specimen. A 46-yr-old woman with B lymphoblastic lymphoma was admitted to the hospital for scheduled chemotherapy. Her chest computed tomography (CT) scan revealed bilateral multifocal nodular and patchy consolidation in both lungs. Investigation of the BAL specimen revealed that 7% of leukocytes had intracellular gram-positive cocci. The quantitative cultures of the BAL specimen grew mucoid, non-hemolytic, and grayish convex colonies on blood agar at a count of approximately 200,000 colony-forming units/mL. The colonies were identified as R. mucilaginosa. The patient was empirically treated with levofloxacin for 7 days, after which findings on the chest radiograph and CT scan improved. She was discharged with improvement on hospital day 46. To our knowledge, this is the first report of R. mucilaginosa pneumonia diagnosed in Korea. Quantitative culture of BAL specimen and examination of intracellular organisms are crucial for assessing the clinical significance of R. mucilaginosa recovered from the lower respiratory tract.