Diagnóstico por estudio bacteriológico cuantitativo de la infección en la herida por quemadura / Diagnosis for quantitative bacteriological study of the infection in the injury by burn

RESUMEN Fundamento: los procesos infecciosos son una de las causas de morbilidad y mortalidad en pacientes quemados, por lo que el diagnóstico temprano de la infección a través del estudio bacteriológico cuantitativo representa un paso de avance para el tratamiento oportuno. Objetivo: determinar mediante el estudio bacteriológico cuantitativo de la herida por quemadura el diagnóstico de infección en pacientes quemados. Métodos: se realizó un estudio descriptivo, de corte transversal para determinar mediante el estudio bacteriológico cuantitativo de la herida por quemadura el diagnóstico de infección en los pacientes quemados ingresados en el servicio de Caumatología del Hospital Universitario Manuel Ascunce Domenech de la provincia Camagüey, desde septiembre de 2015 a noviembre de 2017. Se estudiaron 34 pacientes y se evaluaron las variables: edad, sexo, índice de gravedad, positividad o no de los estudios cualitativos y cuantitativos en relación con las manifestaciones clínicas de infección. Resultados: el sexo femenino fue el más representado con 70,59 % de casos, predominaron las edades entre 48-67 años, el 38,23 % de los lesionados estaban clasificados como muy grave, la colonización fue la predominante sobre la infección en el cultivo cuantitativo con un 26,47 %. En los pacientes con manifestaciones clínicas de infección, el cultivo bacteriológico cuantitativo fue positivo en 11 de ellos para un 32,35 %. Se encontró en el 44,12 % la presencia de gérmenes a una concentración de más de 105 gérmenes por gramo de tejido. Conclusiones: los factores determinantes en la aparición de infección en la herida por quemadura son la edad, la extensión y profundidad de las lesiones. Existió una correlación entre la positividad del estudio bacteriológico cuantitativo y la presencia de manifestaciones clínicas de infección en los pacientes. Se documentó mayor número de cultivos cuantitativos con resultados positivos y su correlación con la presencia de gérmenes en los cultivos cualitativos.

ABSTRACT Background: the infectious processes are one of the main causes of morbidity and mortality in the burned patients, which is why the early diagnosis of the infection through the bacteriological quantitative study represents a forward-motion step for the opportune treatment of these patients. Objective: to determine the diagnosis of infection in the burned patients by means of the bacteriological quantitative study of the injury by burn. Methods: a descriptive, cross-section study was carried out to determine by means of the bacteriological quantitative study of the injury by burn the diagnosis of infection in the burned patients entered in the service of Burn at Manuel Ascunce Domenech Universitary Hospital of the province Camagüey, from September 2015 to November 2017. 34 patients were studied in those who were evaluated the following variables: age, sex, severity rate, positivity or no of the qualitative and quantitative study relating to the clinical public demonstrations of infection. Results: in this study the feminine sex became represented by 70.59 %, predominating ages between 48- 67 years, the 38.23 % of injured persons were classified as very grave, and colonization was the predominant on the infection in the quantitative cultivation with a 26.47 %. In patients with clinical demonstrations of infection, the quantitative culture was positive in 11 of them for a 32.35 %. It was found in 44.12 % of patients, the presence from germs to a concentration of over 105 germen per gram of fabric. Conclusions: determining factors in the appearing of infection in the injury by burn were age, extension and depth of the injuries. There was a correlation between the positivity of the bacteriological quantitative study and the presence of clinical demonstrations of infection in these patients. Greater number of quantitative cultivations with positive results and their correlation with the presence of germs in the qualitative cultivations were documented.

Comparison of four screening methods for group B streptococcus / 中华检验医学杂志

Objective@#To evaluate diagnostic performance of Todd-Hewitt (T-H) broth culture method, direct culture method, liquid chromogenic culture method, and loop-mediated isothermal amplification (LAMP) method for screening group B streptococcus (GBS) during late pregnancy.@*Methods@#In the retrospective study, the rectal vaginal secretions samples were collected from pregnant women at 35 to 37 weeks at the obstetrics clinic of Guangzhou Women and Children′s Medical Center affiliated to Guangzhou Medical University during October 2016 to April 2018. For the purposes of clinical evaluation, T-H broth culture was used as the standard reference method, and double-blind trials were used to evaluate diagnostic performance of direct culture method, liquid chromogenic culture method, and LAMP method for screening group B streptococcus during late pregnancy in three research stages. Sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV), coincidence rate and Yoden index for each method were calculated. Also, the level of agreement between each method and T-H broth was assessed using the kappa (k) coefficient.@*Results@#A total of 969 specimens were detected by the T-H enrichment culture method, and 90 were positive (9.3%). The sensitivities from high to low were LAMP method [100% (25/25)], direct culture method [81.5% (22/27), 95%CI:65.8%-97.1%], and liquid color culture method [71.1% (27/38), 95%CI:55.9%-86.2%]. Specificities were direct culture method [100% (282/282)], liquid color culture method [98.1% (455/464), 95%CI:96.8%-99.3%], and LAMP method [94.0% (125/133), 95%CI: 89.9%-98.1%]. The coincidence rates were direct culture method [98.4% (22+282)/309], liquid color culture method [96.0% (27+455)/502], and LAMP method [94.9% (25+125)/158]. The Kappa values of the direct culture method (0.889), LAMP method (0.832) and the enrichment culture method were all ≥0.75, and that of the liquid color culture method was 0.708. The false negative rate of direct culture method was 18.5% (5/27), and no false negative case by LAMP method, but its false positive rate was 6.0% (8/133). The false negative rate and false positive rate of liquid color culture method were 28.9% (11/38) and 1.9% (9/464), respectively.@*Conclusions@#Of the three screening methods compared in this study, only the LAMP method has the advantages in sensitivity, specificity, and coincidence rate compared with T-H enriched culture method, while the others have a certain degree of false negatives rate. The clinical laboratory can introduce these methods based on laboratory facilities and staffing, or refer to the European and American guidelines and combine the recommended antenatal GBS screening method with intrapartum nucleic acid amplification tests to best meet the clinical demands.

Diphtheria diagnostic capacity in the Western Pacific Region

@#Diphtheria is an acute infectious disease affecting the upper respiratory tract and occasionally the skin and is caused by the action of diphtheria toxin produced by Corynebacterium diphtheriae, Corynebacterium ulcerans and Corynebacterium pseudotuberculosis. Corynebacterium infections are usually difficult to control due to their epidemic patterns, the emergence of new strains, novel reservoirs and their dissemination to susceptible human and animal populations.1 Although C. diphtheriae is largely controlled through mass immunization programmes, diphtheria escalated to epidemic proportions within the Russian Federation and the former Soviet Republics in the 1990s, highlighting the potential for this disease to cause morbidity and mortality when immunization programmes are disrupted.2 A recent review of global diphtheria epidemiology, which included an analysis of cases and information about age, showed age distribu­tion shifts and found that the majority of cases occur in adoles­cents and adults.3 Shifts in age distribution, from children to adolescents and adults, were observed from countries in the Western Pacific Region such as the Lao People’s Democratic Republic,4 the Philippines3 and Viet Nam.5

Study on clinical separation characteristics of L-form of mycobacterium tuberculosis / 中国医师杂志

Objective To explore the laboratory culture and identification of Mycobacterium tuberculosis L forms (MTB-L),isolation rate and drug resistance in smear-positive tuberculosis patients,and to improve clinical attention to MTB-L.Methods 222 smear-positive pulmonary tuberculosis patients treated in our hospital from September 2017 to December 2017 were randomly selected for MGIT 960 and 92-3TB-L liquid culture.After MGIT 960 was reported positive,acid-fast staining was performed on the precipitated smears of 92-3TB-L liquid medium for preliminary screening.The suspected L-positive strain culture was transformed into improved TSA-L solid medium to observe the colony characteristics and microscopic characteristics.The properties of the strain were confirmed by acid-fast staining and tuberculosis DNA amplification.Drug susceptibility and mutation sites of drug resistance genes were analyzed in MTB-L.Results Ⅰ-dentification of MTB-L:after the positive strain has been cultured,the colonies have the characteristics of "fried egg sample","particle-like" and " filament-like".MTB confirmed by tubercle DNA amplification experiments.Isolation rate:after cultured by MGIT 960 and Modified 92-3TB-L medium,the positive rate of single bacterial type was 50.90％,(113/222) the positive rate of both bacterial type and L type was 15.32％ (34/222),and the positive rate of MTB-L type was 2.25％ (5/222).Drug resistance:MTB-L was resistant to Streptomycin,Isoniazid,Rifampin,and Ethanol butylamine.No mutation was found in the drug resistance gene loci.Conclusions Clinical laboratory should routinely develop the culture of L-form of Mycobacterium tuberculosis bacteria,and increase the clinical attention to MTB-L.

Evaluación de una técnica de PCR en tiempo real para determinar colonización por Streptococcusagalactiae en mujeres gestantes de Medellín que consultan en Dinamica IPS / Evaluation of a real-time PCR technique to determine colonization by Streptococcus agalactiae in pregnant women of Medellín who consult in Dinamica IPS

Objetivo: Evaluar una técnica de PCR en tiempo real para determinar colonización por Streptococcus agalactiae en mujeres gestantes de Medellín. Materiales Y Métodos: Se realizó un estudio descriptivo prospectivo, en 150 mujeres gestantes, seleccionadas de forma aleatoria, en una IPS en el periodo comprendido entre Enero-Julio 2016. Criterio de inclusión: Ser gestante entre la semana 35-37, declaración voluntaria de participación en el estudio y de exclusión el uso de antibióticos. A las pacientes, se les tomó muestra con hisopo de lintroito vaginal y de la región anal. Las muestras se procesaron para qPCR, cultivo en caldo selectivo con posterior siembra en agar sangre de carnero y medio cromogénico para S. agalactiae STRB (ChromIDTMStrepto,BioMérieuxSA.). Resultados: La prevalencia de colonización por S. agalactiae en las gestantes fue de 20,9% y 22,3% en agar sangre y agar cromogénico STRB respectivamente, mientras que mediante PCR en tiempo real la prevalencia fue de 36%. Al comparar la qPCR con la prueba de oro se encontró: sensibilidad 79,31% (ICdel95%:0,61-0,90), especificidad 75,45% (IC del 95%: 0,66-0,82), valor predictivo positivo 46% (IC del 95%:0,32-0,59) y negativo 93,2% (IC del 95%: 0,86-0,96). Discusión: Elempleo de la qPCR permitió aumentar la sensibilidad y la oportunidad diagnostica (Eltiempo requerido empleando elcultivo fue de 24-48 Horas y por qPCR 6 horas) ,impactando la reducción de riesgos de transmisión neonata lde S.agalactiae, lo cualpodría representar una Disminución en días de estancia y costos hospitalarios por una infección prevenible.

Materials and Methods: A prospective and descriptive study was conducted in 150 pregnant women, randomly selected, at an IPS between January and July 2016. Inclusion criteria: gestation period week 35-37, voluntary declaration of participation in the study. Exclusion criteria: the use of antibiotics. Samples were taken from the vaginal introitus and the anal region using a hyssop and processed for qPCR as well as the gold standard test [selective broth culture with subsequent culture in blood agar and chromogenic medium for S. agalactiae STRB (ChromIDTMStrepto, BioMérieux SA)]. Results: The prevalence of colonization by S. agalactiae in pregnant women was 20.9% and 22.3% in blood agar and chromogenic agar STRB respectively, where as using qPCR the prevalence was 36.0%. The time required using the culture was 24-48h compared to 6h for qPCR. Our data comparing qPCR with the gold standard test showed: sensitivity 79.31% (95% CI: 0.61-0.90), specificity 75.45% (95% CI: 0.66-0.82), positive predictive value 46.0% (95% CI: 0.32-0.59) and negative 93.2%(95% CI: 0.86-0.96). Discussion: The use of the qPCR increased the sensitivity and the diagnostic opportunity (4x to 8x faster using qPCR), which can lead to a decrease in the risk of neonatal transmission of S. agalactiae and result in a reduction in the length of hospital stay and costs induced by a preventable infection.

Application of SAT technique in detection of Chlamydia trachomatis and Ureaplasma urealyticum / 中华检验医学杂志

Objective To study the Chlamydia trachomatis(CT)and Ureaplasma urealyticum (UU)infection in Guangzhou area, and analyze the consistency of simultaneous amplification and testing (SAT)and conventional methods(CT was detected by latex immunochromatography, UU was detected by liquid culture method).Methods A total of 12 120 samples of urogenital secretions or urine samples were collected from Sun Yat-Sen Memorial Hospital of Sun Yat-Sen University from January 2015 to December 2016.CT-RNA and UU-RNA were detected by the SAT technique, a part of samples were tested by conventional methods at the same time.The positive rates of CT and UU by SAT and the conventional methods between different gender and age groups were analyzed by χ2test, the consistencies between different detection methods were analyzed by Kappa test.Results The positive rate of CT was 4.05%(356/8 781), UU 33.69%(1 125/3 339)in Guangzhou from 2015 to 2016.The positive rate of UU was significantly higher than that of CT(χ2=1 981,P<0.01).Of 145 specimens for CT test,the coincidence rate between SAT and latex immunochromatographic method was 96.55%(140/145), which showed good consistency(Kappa=0.65).Of 186 specimens for UU test,the coincidence rate of the results between the SAT method and liquid culture was 92.47%(172/186),which showed strong consistency(Kappa=0.81). Conclusions The positive rate of UU was significantly higher than that of CT in Guangzhou.The SAT method and conventional methods to detect CT and UU show high consistency, which can provide the evidence for clinical diagnosis of CT and UU infection.

Evaluation of clinical application of different microbial automated inoculation systems / 中华检验医学杂志

Objectives To study the performance of different microbial automated inoculation systems and to evaluate the performance of the Probact microbial automated inoculation and incubation system ( Probact system) and its applications in clinical microbiology laboratory.Methods A total of 160 clinical specimens, including respiratory secretions ( n=61 ) , urine ( n=49 ) , and feces ( n=50 ) , that were submitted to the Clinical Microbiology Laboratory in Peking Union Medical College Hospital of Chinese Academy of Medical Sciences from February 2015 to April 2015 were evaluated.These specimens were processed with conventional manual method, the Probact automated inoculation system, and PREVI Isola Inoculator.The quantity of bacterial species recovery, number of effectively isolated colonies, total number of colonies recovery per plate, and time of processing the 160 specimens by the three methods were evaluated. Wilcoxon signed-rank test and Kruskal-Wallis rank sum test were used for statistical analysis.Results The Probact system had significantly higher quantity of bacterial species recovery (respiratory specimens 3.41 ±1.40, urine 1.92 ±0.86, and feces 1.16 ±0.79) than those by the Isola Inoculator (respiratory specimens 3.75 ±1.29, urine 2.24 ±0.97, and feces 1.92 ±0.72), (P=0.006, 0.011, <0.001).Compared to the manual method, Probact performed less quantity of bacterial species recovery for respiratory specimens(3.85 ±1.38), but higher in feces(0.80 ±0.81)( P<0.001).There is no significant differences for urine ( 1.84 ±1.23 ) ( P=0.266 ) .As for number of isolated colony, the Probact system ( respiratory specimens 12.16 ±7.72, urine 2.71 ±4.24, and feces 5.40 ±5.04 ) had significant smaller numbers than that of Isola Inoculator (respiratory specimens 16.56 ±5.76, urine 4.35 ± 4.89, and feces 8.40 ±3.70) (P<0.001,0.007,0.003).However, both system had larger numbers of isolated colonies than those by the manual method (respiratory specimens 11.30 ±8.42, urine 2.67 ±4.34, and feces 1.90 ±3.90) and the difference was significant for fecal specimens(P<0.001).Regarding the total number of colonies recovery, larger number was found by Isola Inoculator than that by the Probact system for fecal specimens, however, there were no significant differences for respiratory or urine specimens (P=0.524,0.738).Compared with manual method, the Probact system had significantly more numbers of colonies recovery for respiratory and fecal specimens ( P<0.001 ) . The total time for processing 160 specimens was shortest for manual method (281 min), followed by Probact system (419 min) and Isola Inoculator (495 min) .Conclusions The performance of the Probact system is better than the manual method but no superior to the Isola Inoculator.The Probact system can meet the clinical need in terms of full automation and standardization of specimen inoculation and prevention of bias of processing by laboratory staffs using manual method.

The value of smear test in clinical microbiology / 中华检验医学杂志

Smear is the classic method for pathogen dectection,and it is the most direct,simplest and the fastest identification method.The conscientious and standardized operation of smears can effectively determine the quality of specimen,the proportion of bacteria,and even identify the suspicious pathogen directly,which provide valuable information for clinical diagnosis and treatment In addition,morphological examination can also become supplementary approach for the validation,calibration of modern equipment The staff working in the laboratory of microbiology should notice the important value of smear test.

Application of rapid detection of MRSA ST239 clones in bloodstream infections / 中华检验医学杂志

Objective To evaluation the method of rapid detection of Methicillin-resistant Stphylococcus aureus (MRSA) ST239 clones with multiplex PCR assay and investigation of the epidemic status of MRSA blood stream infections in Hefei area.Methods Antibiotic susceptibility testing were applied to MRSA isolates from bloodstream infection,rapid screening and confirmation of MRSA ST239 clones by using multiplex PCR,Multilocus Sequence typing (MLST) and Staphyloccoccal Cassette Chromosome mec(SCCmec) typing.Results 51 of 106 clinic isolates Staphylococcus aureus were identified as MRSA,accounting for 48.1％.The resistance rate of MRSA to erythromycin,aminoglycosides and quinolone were significantly higher than Methicillin Sensitive Staphylococcus aureus (MSSA).Both MRSA and MSSA had a high sensitivity to cotrimoxazole,the sensivity rates were 86.3％ and 94.5％,respectively; 47 of 51 isolates of MRSA that detected by MRSA ST239 rapid screening were ST239 clones.Randomly selected 20 positive screening stains were confirmed as MRSA-ST239-SCCmec Ⅲ by MLST and SCCmectyping.Conclusions In Hefei area,nearly half of MRSA bloodstream infections in clinical isolates are MRSA-ST239-SCCmeclⅢ type and serious multidrug-resistance.The rapid detection of ST239 clones by multiplex PCR is a reliable and effective method for large-scale screening in laboratory.

Detection of Mycobacterium tuberculosis growth by direct microscopic observation has a lower cost, higher sensitivity and higher diagnostic value than liquid and solid media cultures.

Background: Culture is needed to confirm tuberculosis but results are generally obtained after several weeks. Objectives: We compared a direct microscopic observation technique for detection of mycobacterial culture positivity (MODS) with the classic solid and MB/BacT cultures in terms of sensitivity, contamination rate, speed and cost on 488 samples. Results: The sensitivity of the MODS technique - 99,2% (162 positive samples) was higher than MB/BacT 78,4% (125 positive samples) and solid culture 69,6% (113 positive samples) P < 0.005 for all comparisons. The median times to positivity were 21, 13.3 and 3 days on solid media, B/BacT and MODS respectively. Conclusions: The MODS technique is faster and more sensitive than both solid media and MB/BacT culture.

A novel molecular point-of-care testing detection technology for infectious pathogens：GeneXpert / 中华检验医学杂志

GeneXpert is an advanced molecular biological detection system developed in recent years, it integrates and automates the sample preparation , nucleic acid purification, gene amplification, results report of the fluorescent quantitative PCR process.As, an accurate, rapid, biosafe technology , GeneXpert has become one of the novel molecular POCT detection technology platforms for diagnosis of infectious pathogens and especially declared it a major milestone for global TB diagnosis .This paper introduces the detection principle , clinical application in the detection of infectious pathogens , limitation and prospect of GeneXpert.

Isolation and identification of Malassezia furfur from lower respiratory tract secretions / 中华检验医学杂志

Objectives To culture and isolate Malassezia furfur which were suspected in sputum smears,and then to identify the isolates systematically.Methods From March 2010 to September 2011,133 lower respiratory tract secretion samples were collected in Beijing Hospital of the Ministry of Health which were suspected containing Malassezia furfur by Parker ink staining.The samples were inoculated and cultured on CHROMagar mediums containing 1％ Tween 60 ( which was modified from Candida chromogenic media) in the atmospheric environment at 35 ℃.The suspected colonies were selected and cultured on both SDA plates containing 0.5％ Tween 40 and SDA plates containing 0.5％ Tween 60.The traditional phenotypic identification methods were used to identify the pure colonies,including staining,growth on Sabouraud agar plates,Tween test,decomposing Escalin,Catalase test,Tween precipitation test and castor oil test.Then the suspected colonies were identified by analyzing the sequences of 18s rRNAs.Results The isolation rate of Malasseziafurfur from lower respiratory tract secretions was 47.4％ (63/133) and 63 strains were also identified as Malassezia furfur by traditional phenotypic identification methods combined with the 18s rRNA gene sequence analysis.Malassezia can be significantly distinguished from Candida by direct smear of the specimen with Parker ink staining.Conclusions Malassezia furfur was found as pink colonies on modified Candida chromogenic media which were significantly different from other Candida colonies.The isolation rate of lower respiratory tract secretions can be improved by modified Candida chromogenic media.The identification accuracy can be further improved by combining traditional phenotypic identification methods with 18s rRNA sequence analysis.

Evaluación de predictores clínicos de bacteriemia en adultos hospitalizados por neumonía adquirida en la comunidad / Evaluation of clinical predictors of bacteremia in adults hospitalized for community-acquired pneumonia

OBJETIVO: Identificar elementos clínicos sencillos que hagan posible determinar adecuadamente los casos con mayor probabilidad de presentar aislamientos bacterianos en los hemocultivos. MÉTODOS: Estudio de casos y controles con pacientes internados por neumonía adquirida en la comunidad entre 1998 y 2009, definiéndose como casos a los pacientes que presentaron hemocultivos positivos y como controles a aquellos con hemocultivos negativos. Se registraron variables demográficas y clínicas y se las sometió a un análisis bivariado. Las que presentaron diferencias estadísticamente significativas entre los grupos fueron introducidas en un modelo de regresión logística para definir predictores independientes y generar un modelo de predicción clínica. RESULTADOS: De los 322 pacientes estudiados, 15,2 por ciento tuvo hemocultivos positivos. Diez variables mostraron diferencias significativas, pero solo tres (temperatura <38°C, sodio <135 mEq/L y puntaje CURB-65) fueron seleccionadas para el análisis multivariado. El modelo desarrollado mostró escasa capacidad para predecir el resultado de los hemocultivos (R² = 0,176; Hosmer-Lemeshow: P = 0,338). CONCLUSIONES: Los datos obtenidos en esta serie no evidenciaron elementos clínicos con capacidad suficiente para predecir el resultado de los hemocultivos.

OBJECTIVE: Identify simple clinical elements that can be used to adequately determine the cases with the highest probability of presenting bacterial isolates in blood cultures. METHODS: Case-control study with patients hospitalized for community-acquired pneumonia from 1998-2009. Patients with positive blood cultures were defined as cases, and patients with negative blood cultures were defined as controls. The demographic and clinical variables were recorded and a bivariate analysis was conducted. The variables with statistically significant differences between the groups were introduced in a logistic regression model in order to define the independent predictors and generate a clinical prediction model. RESULTS: A total of 15.2 percent of the 322 patients studied had positive blood cultures. Ten variables showed significant differences, but only three variables (temperature <38°C, sodium <135 mEq/L and CURB-65 score) were selected for the multivariate analysis. The model developed showed limited capacity to predict the result of the blood cultures (R² = 0.176; Hosmer-Lemeshow: P = 0.338). CONCLUSIONS: The data obtained in this series did not demonstrate clinical elements with sufficient capacity to predict the result of the blood cultures.

Rapid detection of clinical common bacteria using DNA microarray / 中华检验医学杂志

Objective To detect eight kinds of clinical common pathogenic bacteria by DNA microarray.Methods Eight kinds of common pathogenic bacteria,including Staphylococcus aureus,Pseudomonas aeruginosa,Klebsiella pneumoniae,Escherichia coli,Proteus mirabilis,Enterobacter aerogenes,Pseudomonas fluorescens,Shigella sonnei were collected.Universal primers were designed to amplify 16S rRNA gene fragment from the genomic DNA of the eight bacteria,and probes were designed in the highly variable regions.DNA microarray detection system was established and used for detection of colleted bacteria.A total of 50 samples were collected from the Zhongnan Hospital of Wuhan University,including 6 blood samples,32 sputum samples,9 feces samples and 3 bronchoscope lavage samples.DNA were extracted and detected by the established DNA microarray system.Results The desired fragments were well amplified by the self-designed universal primers.The selected probes had good detection results according to repeated detection.Of the 50 samples detected,pathgenic bacteria were accurately detected in 47 samples.Other three samples were not detected as those bacteria were not included in the chip.By optimizing the detection process,the results could be reported within 8 hours.Observation of probe signal attenuation indicated that even attenuated after 60 days,but the attenuation did not affect the results.Conclusion A microarray system was established for detection of clinical common bacteria accurately and quickly,which provided foundation for its clinical application.

Evaluation of effectiveness of MGIT liquid medium culture manual method for Mycobacterium detection / 中华检验医学杂志

Objective To evaluate the effectiveness of MGIT liquid medium fluorescence instrument manual interpretation method for rapid detection of Mycobacterium. Methods Two hundred sputa with newly diagnosed tuberculosis patients were collected from October 2008 to January 2009 in the district hospitals in Shanghai. Of these 200 sputa, 67 sputa were positive AFB, 133 were negative. All the sputa were isolated by L-J, BacT / Alert 3D system and MGIT liquid medium methods. Results Of the 200 sputa specimens,105(52. 5% ) were isolated as Mycobacterium strains. The positive culture rate of the MGIT, BacT/Alert 3D and L-J method was 49. 5% ( 99/200 ), 48. 0% (96/200) and 45.0% ( 90/200), respectively. The MGIT culture positive rate was significantly higher than that of L-J method (x2 = 5.40, P = 0. 020 1 ). Of the 133 sputa with negative AFB, the positive culture rate was 24. 8% ( 33/133 ), 23. 3% ( 31/133 ) and 18. 8% (25/133) with MGIT, BacT/Alert 3D and L-J method, respectively. The MGIT culture positive rate with the AFB negative sputum was significantly higher than that of L-J method (x2 = 5. 33, P = 0. 020 9 ).The median time of detection with MGIT, BacT/Alert 3D system and L-J method was 11 days, 15 days and 22 days, respectively. Comparing the median time of detection of MGIT with BacT/Alert 3D, the difference was statistically significant ( Z = 3.414 ,P ＜ 0. 01 ). Comparing the median time of detection of MGIT with L-J method, the difference was statistically significant (Z =7.083,P＜0. 01).Conclusions MGIT liquid medium manual method is a rapid detection method of Mycobacterium with a high positive detection rate, and do not need expensive equipment This method may suitable to resource limited medical institutions due to its low cost and short round time.

Rapid identification of common clinical organism by matrix-assisted laser desorption/ionization time of-fright mass spectrometry / 中华检验医学杂志

Objective To estimate the application for the rapid identification of common clinical bacteria by MALDI-TOF MS. MethodsFour hundred and twenty-six bacteria, including Salmonella spp strains collected from Zhejiang center for disease control and prevention were collected from blood, sputum,secretion and urine in 2nd Affiliated Hospital of Zhejiang University during December 2008 to August 2009. The isolates included 76 gram positive coccus and 350 gram negative bacilli. Species identification was performed with the Vitek system, and serotypes of Salmonella and Shigella were determined by serum agglutination test. 16s rDNA gene of 91 bacteria were amplified by PCR. The RCP products were sequenced. Then the results were compared with the reported sequences from GenBank. All strains were identified by MALDI-TOF MS. Results of three identification methods were compared with each other. Results Among 426 tested isolates, identification results from Vitek system and MALDI-TOF MS for gram positive coccus and 323 out of 350 gram negative bacilli (exception for Salmonella and Shigella spp.),were identical. For 23 Salmonella and Shigella spp. , only 2 Salmonella enterica subsp, enterica serovar Typhimurium were identified the same results by the three methods. Besides, results from Vitek system and serum agglutination test for 1 Salmonella enterica subsp, enterica serovar Typhi, 3 Salmonella enterica subsp.enterica serovar Paratyphi A, 1 Salmonella enterica subsp, enterica serovar Paratyphi B, 1 Salmonella enterica subsp, enterica serovar Enteritidis, and 1 Salmonella enterica subsp, enterica serovar Bovis-morbificans were consistent with that from 16S rDNA gene sequence. Four isolates which were confirmed as S. flexneri by Vitek system and serum agglutination test were identified as Escherichia coli by both 16S rDNA gene sequence and MALDI-TOF MS. ConclusionMALDI-TOF MS could be used for rapid and accurate identification of common clinical bacteria with good repeatabihty, excepting for the Salmonella and Shigella spp.

Análise e validação do protocolo de pesquisa de atividade antimicrobiana frente a cepas bacterianas estudadas no LABioMol / UFF - RJ / Analysis and validation of a protocol research of antimicrobial activity against bacterial strains studied in LABioMol / UFF - RJ

A resistência dos microrganismos aos antibióticos e quimioterápicos é um problema de ordem mundial que demanda a descoberta de novas moléculas para o tratamento de infecções causadas por microorganismos multirresistentes. Este trabalho visa a análise e a validação de um protocolo de pesquisa de atividade antimicrobiana frente a cepas bacterianas estudadas no LABioMol/UFF-RJ. Descreve a construção e validação de um instrumento de percepção de suporte biológico. Trata-se de uma pesquisa que visa identificar indicadores biológicos objetivos de ações conceituais e empíricas. A resistência dos microrganismos aos antibióticos e quimioterápicos é um problema de ordem mundial que demanda a descoberta de novas moléculas para o tratamento de infecções causadas por microorganismos multirresistentes. O Laboratório de Antibióticos, Bioquímica e Modelagem Molecular (LABioMol) do Instituto de Biologia da Universidade Federal Fluminense tem desenvolvido um protocolo para pesquisa de atividade antimicrobiana com cepas bacterianas, inclusive com cepas de importância odontológica. Por isso, é importante a divulgação desta metodologia utilizada como meio de informação destas etapas laboratoriais. O protocolo se encontra resumido na tabela abaixo e descrito no texto.

The resistance of microorganisms to antibiotics and chemotherapeutics agents is a problem of world order that demands the discovery of new molecules to the treatment of infections caused by multidrug-resistant bactéria. This work concerns the analysis and validation of a research protocol antimicrobian activity front of the bacterial strains studied in LABioMol / UFF-RJ. This paper describes the construction and validation of an instrumento f perception of biological support. This is a survey that aims to identify biological indicators of actions goals conceptual and empirical. The Laboratory of Antibiotics, Biochemistry and Molecular Modeling (LABioMol) Institute of Biology, Fluminense Federal University hás developed a protocol for the detection of antimicrobial activity in bacterial strains, including strains of dental importance. Therefore, it is important to show this methodology from these steps. The protocol is summarized in the table below and describeb in the test.

Study on blood culture in blood of paratyphoid fever A patients / 中华检验医学杂志

Objective To compare the yield and speed of detection of Salmonella subsp, enterica serotypo Paratyphi A from the blood of patients with suspected paratyphoid fever A. Methods With the BacT/ALERT 3D system and paired aerobic and anaerobic bottles (AEB, ANB) that were each filled with 5 ml of blood, the blood culture of 13 500 suspected paratyphoid fever A patients were performed. Results A total of 4 060 isolates were detected. About 3 149 were detected from both AEB and ANB. Four hundred and sixty-one isolates were detected only from the AEB and 450 were only from the ANB. The detection rates of the AEB and ANB were all 26.7% (χ<'2>=0.023, P=0.880). The increased detection rate attributed to the additional blood volume in the AEB and ANB were 11.3% and 11.1%, respectively. The detection speed differed between the two medium formulations. The time to detection was (23.66±15.89) h and (25.48±16.92) h for3 149 isolates, respectively (t=7.007, P<0.01).The mean time to detection was 31.80±20. 97 for 461 isolates discovered with AEB and (33.45±20.72) h for 450 isolates discovered with ANB. Conclusion The blood volume is an important factor in determining the detection rate of blood culture. Although no statistical difference for positive rate was found between the AEB and ANB, more isolates was detected earlier in AEB.

Screening of LoxP-positive recombinant clones by self-primer colony polymerase chain reaction / 中华检验医学杂志

Objective To investigate the screening and evaluating methods of positive recombinant clones for small fragments such as LoxP sequence. Methods Synthesized LoxP and vector complementary sequence were used as the upper and lower primer respectively, and colonies were used directly as the templates of polymerase chain reaction (PCR). The presence of 784 bp strap in electrophoresis was seen as positive. The positive recombinant clones screened by PCR were evaluated contrastively by restriction endonuclease digestion and verified by DNA sequence analysis. Results Among the six colonies randomly screened by PCR, three showed positive straps and one was verified by DNA sequence analysis. However, the electrophoresis only showed unclear and clouding straps when the three positive recombinant clones were evaluated by restriction endonuclease digestion. Conclusion Self-primer colony PCR is a high-speed, convenient, economic and effective method for screening and evaluating of positive clones recombinated by small fragments such as LoxP sequence.

Analysis of 246 cases of positive blood culture / 中华检验医学杂志

Objective To investigate the kinds of isolates and the causes of bloodsream infection.Methods A total of 4 459 blood samples were cultured by BacT/Alert 3D240. The isolates were identified by API system.Results 247 strains were isolated from 246 patients. The isolates included Gram-negative bacteria (66.0%), Gram-positive bacteria (30.4%) and fungi(3.6%). E. coli, Salmonella paratyphi A , Salmonella typhi and Staphylococcus sp. not-Staphy. aureus were the main pathogen. The causes of bloodstream infection resulted from non-surgical disease, surgical disease, and others were 82.5%, 4.1% and 13.4%, respectively. The major symptom of blood infection was fever, which was presented in 41.5% of positive blood culture cases. 77.6% of Salmonella sp. was isolated from the patients of the department of respiratory and the department of emergency. E. coli was mainly isolated from the patients of the department of nephrolgy, haemotology and surgical department. Staphycoccus sp.not-Staphy. aureus was mainly isolated from the patients of the department of respiratory and pediatrics. The positive blood culture rate in 12,24,36,48,72,96 h were 24.4%,74.0%,87.4%,93.1%,97.2%, and 99.2% respectively.The coincidence rate of positive blood culture detected under microscope and identified by API system was ~99.6%. Conclusion Automated blood culture systems were important apparatus for diagnosis of bloodstream infection.