Bacteriological Profile and Antibiotic Susceptibility Patterns in Neonatal Septicaemia in Tertiary Care Hospital

Introduction: Neonatal septicaemia has great role in morbidityand mortality among neonates. Neonatal mortality rate hasbeen reported in India as 17 per 1000 live births as per 2016-17 data. Neonatal septicaemia may be of early onset or lateonset depending of the age of the neonates. The mostcommon bacterial agents involved are Group B Streptococcus,Klebsiella pneumoniae, CoNS, Streptococcus pneumoniae,Haemophilus influenzae etc. Diagnosis is done by manymethods but the most important and absolute mode ofdiagnosis is blood culture.Aims and Objectives: The present study is done for thedetection of bacteriological profile and their antibioticsusceptibility pattern in case of neonatal septicaemia. Earlydiagnosis and specific treatment can save the lives of manyneonates who are suffering from neonatal septicaemia.Materials and Methods: The material used for the diagnosis isvenous blood of the suspected neonates. Blood culture methodis used for the diagnosis of Neonatal septicaemia. Repeatedsubculture is done on Blood agar, Nutrient agar, andMacConkey agar plates. Confirmation of organism is donethrough different biochemical tests. The antibiotic susceptibilitytesting was performed on Muller Hinton agar (MHA) by KirbyBauer disc diffusion method for bacterial isolates, as perclinical and laboratory standards institute (CLSI) guideline.Results: Total 206 cases of suspected neonatal septicaemiawere investigated in which 142 cases are found positive. Mostcommon organism isolated was Klebsiella pneumoniae(39.44%) than Staphylococcus aureus (33.8%), otherorganisms are Escherichia coli (9.86%), CoNS (8.48%),Pseudomonas (5.63%), Enterococcus (2.82%) etc. overallincidence of Gram negative organism (54.93%) was more thanGram positive organism (45.07%). As far as antibioticsensitivity pattern was concerned most of the organism were100% sensitive to imipenem, meropenem and colistin B andresistant to Ampicillin.Conclusion: Gram negative isolates were more common thanGram positive as the causative agents of neonatal sepsis. Themost common causative organism was Klebsiella pneumoniae.The other organisms isolated were Pseudomonas aeruginosa,Staphylococcus aureus, CoNS, etc. Most of the Gram negativeisolates were sensitive to Amikacin, Gentamycin, Ofloxacin andCiprofloxacin but were highly susceptible to Meropenem,Imipenem and Collistin-B. The Gram positive isolates werebetter sensitive to Amikacin, Cephalosporin, Ciprofloxacin andClindamycin but were less sensitive or resistant to Ampicillinand Erythromycin. They showed high susceptibility toTicoplanim, Linezolid, Vancomycin and Methicillin.

Effect of Sophora flavescens alkaloid on the growth of vaginal lactobacilli:an in vitro study / 中国实用妇科与产科杂志

OBJECTIVE: To study the effects of Kushen gel on the growth of six common vaginal lactobacilli in vitro.METHODS: This study was conducted at Beijing Tsinghua Changgung Hospital from June 2018 to March 2019.Six different vaginal lactobacilli isolated from the healthy women of reproductive age in China were purified and cultured in vitro. The effect of different concentrations of Sophora flavescens alkaloid,which was the main active ingredient of Kushen gel,on the growth of various clinical lactobacillus isolates was observed by the agar dilution method.RESULTS: The minimum inhibitory concentration(MIC)of Sophora flavescens alkaloid to Lactobacillus rhamnosus was 40 mg/mL. The MICs for Lactobacillus crispatus,Lactobacillus jensenii,Lactobacillus gasseri and Lactobacillus reuteri were all 20 mg/mL. The MIC for Lactobacillus vaginalis was 10 mg/mL.When the concentration of Sophora flavescens alkaloid was lower than the MIC value,it had a certain effect on promoting proliferation of both Lactobacillus gasseri and Lactobacillus reuteri,but had no effect on Lactobacillus crispatus or Lactobacillus vaginalis.CONCLUSION: Under the clinical dosage,Kushen gel does not inhibit the growth of common vaginal lactobacilli,which is conducive to the recovery of the vaginal microecosystem.

Development of Statistical Software for the Korean Laboratory Accreditation Program Using R Language: LaboStats

BACKGROUND: In Korea, the Korean Laboratory Accreditation Program (KLAP) has set minimum standards for verification of clinical test performance. This verification process is time-consuming and labor-intensive when performed manually. We developed a free, statistical software program for KLAP, using the R language (R Foundation for Statistical Computing, Vienna, Austria). METHODS: We used CLSI guidelines for the algorithm. We built graphic user interfaces, including data input, with Embarcadero Delphi EX4 (Embarcadero Technologies, Inc., Texas, USA). The R Base Package and MCR Package for Method Comparison Regression were used to implement statistical and graphical procedures. RESULTS: Our program LaboStats has six modules: parallel test, linearity, method comparison, precision, reference interval, and cutoff. Data can be entered into the field either manually or by copying and pasting from an MS Excel worksheet. Users can print out precise reports. CONCLUSIONS: LaboStats can be useful for evaluating clinical test performance characteristics and preparing documents requested by KLAP.

Protocol for Evaluation of the Automated Coagulation Analyzer / 现代检验医学杂志

When the device is installed,it needs to be verified by some special tests to evaluate its performance and to determine whether the device is in accordance with the requirements of the relevant inspection specifications.The projects of the automated coagulation analyzer are particularity different from other tests,especially the biochemistry tests.So the clinical laboratory should establish and implement the matching performance validation protocol.This article refers to the CLSI related documents to summarize a feasible and applicable evaluation protocol,helping for evaluating automated coagulation analyzer.

Establishment and application of external quality assessment system for microbial morphology and detection of special drug-resistance in clinical laboratory / 临床检验杂志

Objective To investigate the establishment,operation and performance of external quality assessment(EQA) system for microbial morphology and detection of special drug-resistance in clinical laboratory,and explore the value of the developed system in clinical application.Methods The pictures of known bacteria and fungi colony,gram staining and acid-fast staining from clinical microbiology were distributed to the participating laboratories in Gansu province twice a year at regular intervals.The pictures of standard knowledge points from CLSI,such as special drug resistance were distributed simultaneously.All the participating laboratories were required to complete the interpretation for the pictures and report their resuhs in a scheduled time.Then the resuhs were summarized and analyzed as 3 modes:complete consistency,general consistency and non-consistency.Results During the 2 years when the EQA system for microbial morphology and detection of special drug-resistance were performed for 24 times,the rate of annual complete consistency increased year by year and reached to 91.3％ in 2015.Conclusion The EQA system based on the examinations of microbial morphology and CLSI standard knowledge points for clinical laboratory may supervise the staff of clinical microbiology laboratories in the hospitals at second grade or above to master the skills of morphological identification and learn CLSI knowledge points,so their professional skills of clinical microbiology could be comprehensively improved.

A Comparison of the Performance of Soyagreentec Ampulab EDTA and Sodium Citrate Tubes with That of BD Vacutainer Tubes

BACKGROUND: Because blood collection tube can affect the results of various laboratory tests, it is necessary before using a newly developed product to verify its performance stringently and objectively. We compared the performance of Ampulab EDTA and sodium citrate tubes (Soyagreentec, Korea) with that of Vacutainer tubes (BD, USA) in accordance with international guidelines. METHODS: This study was performed as a multicenter study of Chung-Ang University Hospital and Seoul National University Hospital to evaluate the performance of two different instrument platforms. We performed the precision test according to CLSI GP34-A, the accuracy test according to CLSI EP09-A2-IR, and the vacuum test according to CLSI H1-A5 as well as stability, and aseptic condition tests. We evaluated 3 lots of Ampulab tubes for their precision, accuracy, vacuum, and aseptic condition. RESULTS: In precision test, the total precision levels of Ampulab tubes in most measurands were desirable or allowable. The results of mean corpuscular hemoglobin concentration, platelet distribution width, basophil, and reticulocyte counts for Ampulab tubes showed imprecision beyond allowable limits, but were similar to those of Vacutainer tubes. In the accuracy test, the bias in most measurands, except for the mean platelet volume, was within allowable limits. In the stability test, Ampulab showed similar performance to Vacutainer. In tests of the vacuum and aseptic conditions, Ampulab fulfilled both requirements. CONCLUSIONS: The performance of Ampulab EDTA and sodium citrate tubes was equivalent to that of Vacutainer tubes.

A Comparison of the Performance of Soyagreentec Ampulab EDTA and Sodium Citrate Tubes with That of BD Vacutainer Tubes

BACKGROUND: Because blood collection tube can affect the results of various laboratory tests, it is necessary before using a newly developed product to verify its performance stringently and objectively. We compared the performance of Ampulab EDTA and sodium citrate tubes (Soyagreentec, Korea) with that of Vacutainer tubes (BD, USA) in accordance with international guidelines. METHODS: This study was performed as a multicenter study of Chung-Ang University Hospital and Seoul National University Hospital to evaluate the performance of two different instrument platforms. We performed the precision test according to CLSI GP34-A, the accuracy test according to CLSI EP09-A2-IR, and the vacuum test according to CLSI H1-A5 as well as stability, and aseptic condition tests. We evaluated 3 lots of Ampulab tubes for their precision, accuracy, vacuum, and aseptic condition. RESULTS: In precision test, the total precision levels of Ampulab tubes in most measurands were desirable or allowable. The results of mean corpuscular hemoglobin concentration, platelet distribution width, basophil, and reticulocyte counts for Ampulab tubes showed imprecision beyond allowable limits, but were similar to those of Vacutainer tubes. In the accuracy test, the bias in most measurands, except for the mean platelet volume, was within allowable limits. In the stability test, Ampulab showed similar performance to Vacutainer. In tests of the vacuum and aseptic conditions, Ampulab fulfilled both requirements. CONCLUSIONS: The performance of Ampulab EDTA and sodium citrate tubes was equivalent to that of Vacutainer tubes.

Molecular Identification and Amphotericin B Susceptibility Testing of Clinical Isolates of Aspergillus From 11 Hospitals in Korea

BACKGROUND: We investigated the species distribution and amphotericin B (AMB) susceptibility of Korean clinical Aspergillus isolates by using two Etests and the CLSI broth microdilution method. METHODS: A total of 136 Aspergillus isolates obtained from 11 university hospitals were identified by sequencing the internal transcribed spacer (ITS) and beta-tubulin genomic regions. Minimal inhibitory concentrations (MICs) of AMB were determined in Etests using Mueller-Hinton agar (Etest-MH) and RPMI agar (Etest-RPG), and categorical agreement with the CLSI method was assessed by using epidemiological cutoff values. RESULTS: ITS sequencing identified the following six Aspergillus species complexes: Aspergillus fumigatus (42.6% of the isolates), A. niger (23.5%), A. flavus (17.6%), A. terreus (11.0%), A. versicolor (4.4%), and A. ustus (0.7%). Cryptic species identifiable by beta-tubulin sequencing accounted for 25.7% (35/136) of the isolates. Of all 136 isolates, 36 (26.5%) had AMB MICs of > or =2 microg/mL by the CLSI method. The categorical agreement of Etest-RPG with the CLSI method was 98% for the A. fumigatus, A. niger, and A. versicolor complexes, 87% for the A. terreus complex, and 37.5% for the A. flavus complex. That of Etest-MH was < or =75% for the A. niger, A. flavus, A. terreus, and A. versicolor complexes but was higher for the A. fumigatus complex (98.3%). CONCLUSIONS: Aspergillus species other than A. fumigatus constitute about 60% of clinical Aspergillus isolates, and reduced AMB susceptibility is common among clinical isolates of Aspergillus in Korea. Molecular identification and AMB susceptibility testing by Etest-RPG may be useful for characterizing Aspergillus isolates of clinical relevance.

SUSCEPTIBILITY OF Candida spp. ISOLATED FROM BLOOD CULTURES AS EVALUATED USING THE M27-A3 AND NEW M27-S4 APPROVED BREAKPOINTS / Suscetibilidade de Candida spp. isoladas de hemocultivos, avaliadas pelos breakpoints dos documentos M27-A3 e M27-S4 do CLSI

The high mortality rates associated with candidemia episodes and the emergence of resistance to antifungal agents necessitate the monitoring of the susceptibility of fungal isolates to antifungal treatments. The new, recently approved, species-specific clinical breakpoints (SS-CBPs)(M27-S4) for evaluating susceptibility require careful interpretation and comparison with the former proposals made using the M27-A3 breakpoints, both from CLSI. This study evaluated the susceptibility of the different species of Candida that were isolated from candidemias based on these two clinical breakpoints. Four hundred and twenty-two isolates were identified and, among them, C. parapsilosis comprised 46.68%, followed by C. albicans (35.78%), C. tropicalis (9.71%), C. glabrata (3.55%), C. lusitaniae (1.65%), C. guilliermondii (1.65%) and C. krusei (0.94%). In accordance with the M27-A3 criteria, 33 (7.81%) non-susceptible isolates were identified, of which 16 (3.79%) were resistant to antifungal agents. According to SS-CBPs, 80 (18.95%) isolates were non-susceptible, and 10 (2.36%) of these were drug resistant. When the total number of non-susceptible isolates was considered, the new SS-CBPs detected 2.4 times the number of isolates that were detected using the M27-A3 interpretative criteria. In conclusion, the detection of an elevated number of non-susceptible species has highlighted the relevance of evaluating susceptibility tests using new, species-specific clinical breakpoints (SS-CBPs), which could impact the profile of non-susceptible Candida spp. to antifungal agents that require continuous susceptibility monitoring.

As elevadas taxas de mortalidade associadas com episódios de candidemia e a emergência da resistência aos antifúngicos, requerem o monitoramento da suscetibilidade de Candida spp., isoladas das candidemias, frente aos agentes antifúngicos. Os novos breakpoints, chamados “espécie-específicos,” foram recentemente aprovados (M27-S4) requerendo, pois, cuidadosa interpretação e comparações com aqueles até agora utilizados (M27-A3); ambos são propostos pelo Clinical Laboratory Standard Institute (CLSI). O presente estudo avaliou a suscetibilidade de espécies de Candida isoladas de candidemias baseando-se nestes dois breakpoints. Quatrocentos e vinte e dois isolados de Candida foram identificados e assim distribuídos: C. parapsilosis (48,68%), C. albicans (35,78%), C. tropicalis (9,71%), C. glabrata (3,55%), C. lusitaniae (1,65%), C. guilliermondii (1,65%), C. krusei (0,94%). Com base nos critérios do M27-A3, um total de 33 (7,81%) isolados foram julgados não-sensíveis, dos quais 16 (3,79%) como resistentes aos antifúngicos. De acordo com os breakpoints espécie-específicos (M27-S4) um total de 80 (18,95%) isolados foram considerados não-sensíveis, dos quais 10 (2,36%) resistentes a algum dos antifúngicos testados. Com base nos novos breakpoints espécie-específicos, o número de isolados não-sensíveis foi 2,4 vezes maior do que o número de não-sensíveis detectado pelos breakpoints do documento M27-A3. A detecção de um elevado número de isolados não-sensíveis através dos breakpoints propostos pelo M27-S4 destaca a importância dos testes de suscetibilidade, os quais trarão impactos no reconhecimento de isolados de Candida spp. não-sensíveis em episódios de candidemias, requerendo, portanto, continua avaliação.

Use of the VITEK 2 system to identify and test the antifungal susceptibility of clinically relevant yeast species

Eleven quality control isolates (Candida albicans ATCC 64548, C. tropicalis ATCC 200956, C. glabrata ATCC 90030, C. lusitaniae ATCC 200951, C. parapsilosis ATCC 22019, C. krusei ATCC 6258, C. dubliniensis ATCC 6330, Saccharomyces cerevisiae ATCC 9763, Cryptococcus neoformans ATCC 90012, C. gattii FIOCRUZ-CPF 60, and Trichosporon mucoides ATCC 204094) and 32 bloodstream isolates, including C. albicans, C. tropicalis, C. parapsilosis, C. glabrata, C. krusei, C. guilliermondii, C. pelliculosa (Pichia anomala), C. haemulonii, C. lusitaniae, and C. kefyr were identified at the species level by the VITEK 2 system. A set of clinical isolates (32 total) were used as challenge strains to evaluate the ability of the VITEK 2 system to determine the antifungal susceptibility of yeasts compared with the CLSI and EUCAST BMD reference standards. The VITEK 2 system correctly identified 100% of the challenge strains. The identification of yeast species and the evaluation of their susceptibility profiles were performed in an automated manner by the VITEK 2 system after approximately 15 h of growth for most species of Candida. The VITEK 2 system ensures that each test is performed in a standardized manner and provides quantitative MIC results that are reproducible and accurate when compared with the BMD reference methods. This system was able to determine the MICs of amphotericin B, flucytosine, voriconazole, and fluconazole in 15 h or less for the most common clinically relevant Candida species. In addition, the VITEK 2 system could reliably identify resistance to flucytosine, voriconazole, and fluconazole and exhibits excellent quantitative and qualitative agreement with the CLSI or EUCAST broth microdilution reference methods.

Validación del inóculo por densitometría para las pruebas de susceptibilidad a los antifúngicos en especies del género Fusarium / Validation of the inoculum by densitometry for antifungal susceptibility testing in Fusarium species

El objetivo de este trabajo fue validar la preparación del inóculo por densitometría para las pruebas de susceptibilidad a los antifúngicos en especies del género Fusarium. Se emplearon 15 aislamientos clínicos de Fusarium spp. para preparar los inóculos por espectrofotometría y contaje de unidades formadoras de colonias en cámara de Neubauer, siguiendo los protocolos establecidos por los documentos de referencia M38-A2 del Instituto de Estándares Clínicos y de Laboratorio (CLSI) y E.DEF 9.1 del Comité Europeo para Pruebas de Susceptibilidad a los Antimicrobianos (EUCAST), respectivamente. En paralelo se determinaron las lecturas por densitometría para ambos procedimientos. Se estableció un rango de 0,5-0,7 unidades McFarland para la preparación del inóculo por densitometría según el CLSI, y un rango de 0,2-0,8 unidades McFarland para la metodología descrita por el EUCAST. Con este estudio, se logró validar la preparación del inóculo para las pruebas de susceptibilidad en Fusarium spp., utilizando la densitometría como método alternativo de los procedimientos descritos internacionalmente, con considerables ventajas para ser implementado en los laboratorios de microbiología clínica. La variabilidad en cuanto a la capacidad de esporulación y tamaño de las conidias, sobre todo en especies poco frecuentes de Fusarium, sugiere la necesidad de validar el inóculo por especie.

The purpose of this work was to validate the preparation of the inoculum by densitometry for antifungal susceptibility testing in Fusarium species. Fifteen clinical isolates of Fusarium spp. were used to prepare the inocula by spectrophotometry and counting of colony forming units in a Neubauer chamber, according to the protocols established by the reference documents M38-A2 of the Clinical and Laboratory Standards Institute (CLSI), and E.DEF 9.1 of the European Committee on Antimicrobial Susceptibility Testing (EUCAST), respectively. Densitometry readings were determined in parallel for both procedures. A range of 0.5-0.7 McFarland units was established for inocula preparation by densitometry according to the CLSI, and a range of 0.2-0.8 McFarland units was established for the methodology described by EUCAST. This study allowed validating the preparation of the inocula for antifungal susceptibility testing in Fusarium spp., using densitometry as an alternative method for other procedures described internationally, with considerable advantages that can be implemented at clinical microbiology laboratories. The variability regarding sporulation capacity and conidia size, especially in less frequent Fusarium species, suggests the need of validating inocula per species.

In Vitro Fluconazole and Voriconazole Susceptibilities of Candida Bloodstream Isolates in Korea: Use of the CLSI and EUCAST Epidemiological Cutoff Values

BACKGROUND: At present, the clinical breakpoints (CBPs) of both fluconazole and voriconazole are available only for 3 common Candida species in the Clinical and Laboratory Standards Institute (CLSI) and the European Committee on Antimicrobial Susceptibility Testing (EUCAST) methods. Epidemiological cutoff values (ECVs) were recently applied to both methods to detect the emergence of acquired resistance (i.e., non-wild-type isolates) among 5 common Candida species. METHODS: We performed a nationwide study to determine the fluconazole and voriconazole susceptibility of Candida bloodstream isolates (BSIs) using both the CLSI and EUCAST methods. A total of 423 BSIs of 5 Candida species were collected from 8 hospitals. The azole susceptibilities were assessed on the basis of the species-specific CBPs and ECVs. RESULTS: Of the 341 BSIs of 3 common Candida species (i.e., C. albicans, C. tropicalis, and C. parapsilosis), 0.3% and 0.9%, 0.0% and 1.5% of isolates were categorized as fluconazole and voriconazole resistant according to the CLSI and EUCAST CBPs, respectively. Of 423 total BSIs, 1.4% and 2.6% had fluconazole minimum inhibitory concentrations (MICs) exceeding the ECVs according to the CLSI and EUCAST, respectively; 1.0% and 2.1% had voriconazole MICs exceeding the ECVs according to the CLSI and EUCAST, respectively. Categorical agreement between the methods using ECVs was 98.3% for fluconazole and 98.3% for voriconazole. CONCLUSIONS: The EUCAST and CLSI methods using ECVs provide highly concordant results. Moreover, non-wild-type isolates with possibly acquired azole resistance were rare among the BSIs of 5 common Candida species in Korea.

In Vitro Fluconazole and Voriconazole Susceptibilities of Candida Bloodstream Isolates in Korea: Use of the CLSI and EUCAST Epidemiological Cutoff Values

BACKGROUND: At present, the clinical breakpoints (CBPs) of both fluconazole and voriconazole are available only for 3 common Candida species in the Clinical and Laboratory Standards Institute (CLSI) and the European Committee on Antimicrobial Susceptibility Testing (EUCAST) methods. Epidemiological cutoff values (ECVs) were recently applied to both methods to detect the emergence of acquired resistance (i.e., non-wild-type isolates) among 5 common Candida species. METHODS: We performed a nationwide study to determine the fluconazole and voriconazole susceptibility of Candida bloodstream isolates (BSIs) using both the CLSI and EUCAST methods. A total of 423 BSIs of 5 Candida species were collected from 8 hospitals. The azole susceptibilities were assessed on the basis of the species-specific CBPs and ECVs. RESULTS: Of the 341 BSIs of 3 common Candida species (i.e., C. albicans, C. tropicalis, and C. parapsilosis), 0.3% and 0.9%, 0.0% and 1.5% of isolates were categorized as fluconazole and voriconazole resistant according to the CLSI and EUCAST CBPs, respectively. Of 423 total BSIs, 1.4% and 2.6% had fluconazole minimum inhibitory concentrations (MICs) exceeding the ECVs according to the CLSI and EUCAST, respectively; 1.0% and 2.1% had voriconazole MICs exceeding the ECVs according to the CLSI and EUCAST, respectively. Categorical agreement between the methods using ECVs was 98.3% for fluconazole and 98.3% for voriconazole. CONCLUSIONS: The EUCAST and CLSI methods using ECVs provide highly concordant results. Moreover, non-wild-type isolates with possibly acquired azole resistance were rare among the BSIs of 5 common Candida species in Korea.

ESBLs — A Challenge to the Clinicians.

Introduction: Treatment of extended spectrum betalactamase (ESBL) producing strains of Enterobacteriaceae has emerged as a major challenge in hospitalized critical as well as community based patients. Infections due to ESBL producers range from uncomplicated urinary tract infection to life threatening sepsis. Methods: We conducted a study to detect the presence of these enzymes in isolates in tertiary care hospital. A total 318 non-repetitive isolates were screened for resistance to any of five screening agents by CLSI (formerly NCCLS) disc diffusion method. Those with suspicious profile were checked for ESBL production by phenotypic confirmation test as recommended by CLSI Disc potentiation method. Various cephalosporin- b-lactamase inhibitor combinations were also tested. Results: Of the 269(84.59%) screen-positive isolates, only 219(81.41%) were identified as ESBL producers. From 219, only 136(62.10%) of Escherichia coli and 83(37.89%) Klebsiella pneumoniae were ESBL producers. Conclusion: Tests for the detection of ESBL producing Escherichia coli and Klebsiella pneumoniae should be carried out in all diagnostic centres routinely because drug resistance patterns are constantly changing.

Susceptibility tests of oropharyngeal Candida albicans from Egyptian patients to fluconazole determined by three methods

Candida albicans frequently cause oropharyngeal candidiasis in immunocompromised patients. As some of these isolates show resistance against azoles, the clinician is wary of initiating therapy with fluconazole (FZ) until a final susceptibility report is generated. We aimed to evaluate the efficacy of rapid flow cytometry (FCM) and disc diffusion (DD) methods in comparison to reference microdilution (MD) of Clinical and Laboratory Standards Institute (CLSI) method for FZ. Thirty seven Candida albicans isolates were tested by the three methods. By both MD and FCM, 26/37 (70.3%) were sensitive with minimal inhibitory concentration (MIC) ¡Ü 8¦Ìg/ml, 5/37 (13.5%) were susceptible dose dependant (S-DD) with MIC 16-32 ¦Ìg/ml and 6/37 (16.2%) were resistant with MIC ¡Ý64¦Ìg/ml. More than 92% of isolates susceptible to FZ by the MD were susceptible by the DD methods with good agreement (81.08%, P = 0.000). However, 4/5 isolates diagnosed as S-DD by MD were resistant by DD. Interestingly, the MIC by FCM at 4 h showed excellent agreement (95.59%, P = 0.000) to that obtained by MD method at 24 h. Overall, FCM antifungal susceptibility testing provided rapid, reproducible results that are valuable alternative to MD. The DD test is recommended as a simple and reliable screening test for the detection of susceptible Candida albicans isolates to FZ.

Estudio comparativo de medición entre lecturas visuales y espectrofotométricas en pruebas de susceptibilidad in vitro de aislamientos de candida / Comparative study of measure visual and spectrophotometer lectures in a in vitro antifungal susceptibility testing in candida isolates

Se realizó un estudio de susceptibilidad in vitro frente a itraconazol, ketoconazol y clotrimazol de 144 cepas de Candida, conservadas y previamente identificadas, aisladas de la cavidad oral de pacientes infectados por el virus de inmunodeficiencia humana (VIH) con cuadros clínicos de candidiasis orofaríngea (COF). El estudio se llevó a cabo mediante dos metodologías; la primera, utilizando los requerimientos del Clinical and Laboratory Standard Institute (CLSI), en el cual está establecida la lectura visual para determinar los patrones de susceptibilidad; y la segunda, mediante la propuesta de la European Committee on Antibiotic Susceptibility Testing, el cual tiene fijado la lectura espectrofotométrica para eliminar las posibles subjetividades de la metodología del CLSI. Los resultados obtenidos mediante ambas lecturas no mostraron diferencias mayores a dos diluciones en los valores de concentración mínima inhibitoria, y demostraron que ambos métodos se correlacionan y es importante para aquellos laboratorios de pocos recursos económicos.

A study of in vitro susceptibility was realized opposite to itraconazol, ketoconazol and clotrimazol of 144 strains of Candida, conserved and previously identified, isolated of the oral cavity of patients infected by the virus of human immunodeficiency (VIH) with clinical pictures of oropharingeal candidiasis (COF). The study was realized by means of two methodologies; the first one, using the requests of the Clinical and Laboratory Standard Institute (CLSI), in which the visual reading is established to determine the patterns of susceptibility; and the second one, by means of the proposal of the European Committee on Antibiotic Susceptibility Testing, which has fixed the spectrophotometric to eliminate the possible subjectivities of the methodology of the CLSI. The results obtained by means of both readings did not show differences bigger than two dilutions in the values of minimal inhibitory concentration, demonstrating that both methods are correlated and it is important for those laboratories of few economic resources.

Comparison of Clinical and Laboratory Standards Institute and European Committee on Antimicrobial Susceptibility Testing Breakpoints for beta-Lactams in Enterobacteriaceae Producing Extended-Spectrum beta-Lactamases and/or Plasmid-Mediated AmpC beta-Lacta / 대한임상미생물학회지

BACKGROUND: In 2010, the Clinical and Laboratory Standards Institute (CLSI) revised breakpoints for cephalosporins and carbapenems and indicated that extended-spectrum beta-lactamase (ESBL) testing is no longer necessary for Enterobacteriaceae. We compared the results of the CLSI 2010 and the European Committee on Antimicrobial Susceptibility Testing (EUCAST) MIC breakpoints for Enterobacteriaceae producing ESBL and/or plasmid-mediated AmpC beta-lactamase (PABL). METHODS: A total of 94 well-characterized clinical isolates of Escherichia coli, Klebsiella oxytoca, Klebsiella pneumoniae, Proteus mirabilis, Salmonella spp., Shigella spp., Citrobacter freundii, Enterobacter aerogenes, Enterobacter cloacae, and Serratia marcescens were analyzed. Of them, 57 were ESBL producers, 24 were PABL producers, and 13 were ESBL plus PABL co-producers. Broth microdilution MIC tests were performed for cefotaxime, ceftazidime, aztreonam, cefepime, and imipenem. RESULTS: Among the 94 isolates containing ESBL and/or PABL, the number of isolates that were susceptible to cefotaxime, ceftazidime, aztreonam, cefepime, and imipenem according to the CLSI 2010 vs. the EUCAST breakpoints were 4 (4.3%) vs. 4 (4.3%); 26 (27.7%) vs. 8 (8.5%); 37 (39.4%) vs. 14 (14.9%); 71 (75.5%) vs. 31 (33.0%); and 76 (80.9%) vs. 90 (95.7%), respectively. Of the 18 isolates that were not susceptible to imipenem according to the CLSI 2010 breakpoints, 13 isolates (72.2%) were P. mirabilis. CONCLUSION: The CLSI 2010 MIC breakpoints without tests to detect ESBL and/or PABL for Enterobacteriaceae could be unreliable. Thus, special tests for ESBLs and AmpC beta-lactamases are required to detect the resistance mechanisms involved.

Gestão de qualidade na fase pré-analítica parte I: análise crítica do CLSI H3-A6 / Quality management in pre analytical phase part I: critical analyze of CLSI H3-A6

A fase pré analítica, segundo dados da literatura é responsável por mais de dois terços de todos os erros atribuídos aos laboratórios de análises clínicas e há apenas alguns procedimentos de rotina para detecção de não conformidades neste domínio de atividades. Nosso objetivo foi fazer uma análise crítica do CLSI H3-A6 Procedimentos para coleta do espécime diagnóstico sanguíneo por punção venosa para ajudar flebotomista e o gestor da qualidade a garantir a segurança do paciente. Nós confrontamos os detalhes do CLSI A3-H6 com as publicações internacionais mais recentes e concluímos que os procedimentos relacionados à punção venosa necessitam de pequenas mudanças para garantir a segurança do paciente.

The pre analytical phase is responsible for more than two-thirds of all errors attributed to the clinical laboratory and there are only a few routine procedures for the detection of nonconformities in this field of activity. Our aim was make a critical analyze of CLSI H3-A6 Procedures for the Collection of Diagnostic Blood Specimens by Venipuncture to help the phlebotomists and quality manager guarantee the patient safety. We contrast details from CLSI H3-A6 with up to date from literature and conclude that the procedures related venipuncture needs little changes to guarantee the patient safety.

Antifungal susceptibility profile of trichosporon isolates: correlation between CLSI and etest methodologies

The aim of the present study was to evaluate the antifungal susceptibility profile of Trichosporon species isolated from different sources employing the Clinical and Laboratory Standards Institute (CLSI) method and E-test method. Thirty-four isolates of Trichosporon spp. and six CBS reference samples were tested for their susceptibility to Amphotericin B, 5-flucytosine, Fluconazole, Itraconazole, Voriconazole and Terbinafine. All species showed high Minimun Inhibitory Concentrations (MIC) for Itraconazole and susceptibility to Fluconazole, The comparison among the results obtained by the CLSI method and E-test revealed larger discrepancies among 5-flucytosine and Itraconazole. The present work provides epidemiological data that could influence therapeutic choices. Furthermore, the comparison between different methodologies could help to analyze results obtained by different laboratories.

Estandarización del inóculo por desintometría para las pruebas de susceptibilidad antifungica en especies de fusarium / Inoculum standadization by densitometry for the antifungal susceptibility testing of fusarium species

Las especies del género Fusarium han emergido como patógenos oportunistas en las últimas décadas. La aparición de cepas resistentes y la introducción de nuevos antifúngicos, hace necesaria la realización de las pruebas de susceptibilidad en los laboratorios de microbiología clínica. El objetivo de este estudio fue estandarizar la preparación del inóculo por densitometría para las pruebas de susceptibilidad a los antifúngicos en especies de Fusarium. Se utilizaron 15 aislamientos clínicos de Fusarium spp. y se prepararon los inóculos por espectrofotometría y por contaje de unidades formadoras de conidias en cámara de Neubauer, siguiendo los protocolos establecidos por el CLSI y EUCAST respectivamente, determinando en paralelo sus lecturas por densitometría para ambos procedimientos. Las lecturas densitométricas a través del uso del Densimat®, permitieron establecer un intervalo de 0,5 ­ 0,7 unidades Mc Farland para la preparación del inóculo por la metodología descrita por el CLSI y un rango de 0,2 ­ 0,8 unidades Mc Farland para la metodología según el EUCAST. Con este estudio, pionero en Venezuela, se logró estandarizar la preparación del inóculo óptimo en las pruebas de susceptibilidad para Fusarium spp. utilizando la densitometría como método alternativo, comparable y sustitutivo de los procedimientos descritos internacionalmente, con considerables ventajas (útil, disponible y reproducible) para ser implementado en los laboratorios de microbiología clínica. La variabilidad en cuanto a la capacidad de esporulación y tamaño de las conidias, sobre todo en las especies poco frecuentes de Fusarium, sugiere la necesidad de estandarizar el inóculo por especie.

Fusarium species have emerged like opportunistic pathogens in the last decades. The appearance of new resistant strains and the introduction of new antifungal agents, make necessary susceptibility tests in clinical microbiology laboratories. The objective of this study was to standardize inoculum preparation by densitometry for antifungal susceptibility testing in Fusarium species. Fifteen clinical isolates of Fusarium spp. were used, and the inocula were prepared by spectrophotometry and by conidia forming count in Neubauer chamber, following the establishment protocols of CLSI and EUCAST, respectively, determining in parallel their densitometry readings for both procedures. Densitometry readings by Densimat® allowed to establish an interval of 0.5 - 0.7 Mc Farland units for the inoculum preparation by the CLSI methodology, and a rank of 0.2 - 0.8 Mc Farland units for the methodology according to the EUCAST. With this study, pioneer in Venezuela, it was achieved the standardization of the optimal inoculums preparation for the susceptibility tests in Fusarium spp., using densitometry like an alternative, comparable and substitute method of the described internationally standard procedures, with considerable advantages (useful, available and reproducible) to be applied in clinical microbiology laboratories. The variability as far as both sporulation capability and conidia's size, mainly in less frequent species of Fusarium, suggests the needs to standardize the inoculums by species.