USE OF MUELLER-HINTON BROTH AND AGAR IN THE GERM TUBE TEST

Candida albicans is often isolated from clinical samples, thus its presumptive differentiation from other species of the same genus can be based on its ability to form the germ tube in human serum. Nevertheless, there are two other species that share this characteristic: C. dubliniensis and C. africana. The aim of this study was to compare four different substrates to perform the germ tube (GT) test. The Candida spp. isolates were identified using a manual system (135 C. albicans, 24 C. tropicalis and one C. dubliniensis). The germ tube test was performed with fresh, previously frozen serum and Mueller-Hinton (MH) broth and agar. GT was observed in 96% (130/136) of the isolates through the fresh serum technique, 94% (128/136) through previously frozen serum, 92% (125/136) in MH agar, and 90% (122/136) in MH broth. The sensitivity of each test was higher than 90%, with 100% specificity. Both the MH agar and broth were able to identify the true positives, and false positives were not found. However, some C. albicans isolates were not identified. MH agar and broth may be used in laboratory for the rapid presumptive identification of C. albicans, as an alternative method for germ tube test.

Determination of germ tube, phospholipase, and proteinase production by bloodstream isolates of Candida albicans

Introduction Candida albicans is a commensal and opportunistic agent that causes infection in immunocompromised individuals. Several attributes contribute to the virulence and pathogenicity of this yeast, including the production of germ tubes (GTs) and extracellular hydrolytic enzymes, particularly phospholipase and proteinase. This study aimed to investigate GT production and phospholipase and proteinase activities in bloodstream isolates of C. albicans. Methods One hundred fifty-three C. albicans isolates were obtained from blood samples and analyzed for GT, phospholipase, and proteinase production. The assays were performed in duplicate in egg yolk medium containing bovine serum albumin and human serum. Results Detectable amounts of proteinase were produced by 97% of the isolates, and 78% of the isolates produced phospholipase. GTs were produced by 95% of the isolates. A majority of the isolates exhibited low levels of phospholipase production and high levels of proteinase production. Conclusions Bloodstream isolates of C. albicans produce virulence factors such as GT and hydrolytic enzymes that enable them to cause infection under favorable conditions.

An optimized one-tube, semi-nested PCR assay for Paracoccidioides brasiliensis detection

Introduction Herein, we report a one-tube, semi-nested-polymerase chain reaction (OTsn-PCR) assay for the detection of Paracoccidioides brasiliensis. Methods We developed the OTsn-PCR assay for the detection of P. brasiliensis in clinical specimens and compared it with other PCR methods. Results The OTsn-PCR assay was positive for all clinical samples, and the detection limit was better or equivalent to the other nested or semi-nested PCR methods for P. brasiliensis detection. Conclusions The OTsn-PCR assay described in this paper has a detection limit similar to other reactions for the molecular detection of P. brasiliensis, but this approach is faster and less prone to contamination than other conventional nested or semi-nested PCR assays.

Effects of Rosmarinus officinalis essential oil on germ tube formation by Candida albicans isolated from denture wearers

Introduction The aim of this study was to investigate the effects of Rosmarinus officinalis essential oil on germ tube formation by Candida albicans isolated from denture wearers. Methods Ten C. albicans isolates recovered from denture wearers were tested using 10% fetal bovine serum with or without 4% R. officinalis essential oil. Results The essential oil from R. officinalis completely inhibited germ tube formation in the investigated C. albicans isolates. Conclusions The results demonstrate that the essential oil of R. officinalis modulates C. albicans pathogenicity through its primary virulence factor (i.e., germ tube formation was suppressed).

Single-tube nested PCR assay with in-house DNA extraction for Mycobacterium tuberculosis detection in blood and urine

Abstract: INTRODUCTION : Molecular analyses are auxiliary tools for detecting Koch's bacilli in clinical specimens from patients with suspected tuberculosis (TB). However, there are still no efficient diagnostic tests that combine high sensitivity and specificity and yield rapid results in the detection of TB. This study evaluated single-tube nested polymerase chain reaction (STNPCR) as a molecular diagnostic test with low risk of cross contamination for detecting Mycobacterium tuberculosis in clinical samples. METHODS: Mycobacterium tuberculosis deoxyribonucleic acid (DNA) was detected in blood and urine samples by STNPCR followed by agarose gel electrophoresis. In this system, reaction tubes were not opened between the two stages of PCR (simple and nested). RESULTS: STNPCR demonstrated good accuracy in clinical samples with no cross contamination between microtubes. Sensitivity in blood and urine, analyzed in parallel, was 35%-62% for pulmonary and 41%-72% for extrapulmonary TB. The specificity of STNPCR was 100% in most analyses, depending on the type of clinical sample (blood or urine) and clinical form of disease (pulmonary or extrapulmonary). CONCLUSIONS: STNPCR was effective in detecting TB, especially the extrapulmonary form for which sensitivity was higher, and had the advantage of less invasive sample collection from patients for whom a spontaneous sputum sample was unavailable. With low risk of cross contamination, the STNPCR can be used as an adjunct to conventional methods for diagnosing TB.

External quality assessment in the voluntary counseling and testing centers in the Brazilian Amazon using dried tube specimens: results of an effectiveness evaluation

INTRODUCTION : In 2011, the Brazilian Ministry of Health rolled out a program for the external quality assessment of rapid human immunodeficiency virus (HIV) tests using the dried tube specimen (DTS) method (EQA-RT/DTS-HIV). Our objective was to evaluate the implementation of this program at 71 voluntary counseling and testing centers (VCTCs) in the Brazilian Legal Amazonian area one year after its introduction. METHODS : Quantitative and qualitative study that analyzed secondary data and interviews with healthcare workers (HCWs) (n=39) and VCTC coordinators (n=32) were performed. The assessment used 18 key indicators to evaluate the three dimensions of the program's logical framework: structure, process, and result. Each indicator was scored from 1-4, and the aggregate results corresponding to the dimensions were expressed as proportions. The results were compared to the perceptions of the HCWs and coordinators regarding the EQA-RT/DTS-HIV program. RESULTS: The aggregate scores for the three dimensions of structure, process, and result were 91.7%, 78.6%, and 95%, respectively. The lowest score in each dimension corresponded to a different indicator: access to Quali-TR online system 39% (structure), registration in Quali-TR online system 38.7% (process), and VCTC completed the full process in the program's first round 63.4% (result). Approximately 36% of the HCWs and 52% of the coordinators reported enhanced trust in the program for its rapid HIV testing performance. CONCLUSIONS: All three program dimensions exhibited satisfactory results (>75%). Nevertheless, the study findings highlight the need to improve certain program components. Additionally, long-term follow-ups is needed to provide a more thorough picture of the process for external quality assessment.

Myocardial Revascularization Surgery with Regional Anesthesia Without an Endotracheal Tube in Conscious Patients

OBJECTIVE: To report initial experience with myocardial revascularization surgery (MRS) performed on patients who were totally awake and without an endotracheal tube.METHODS: Between January 1994 and May 2001, 272 patients underwent MRS without extracorporeal circulation. In 24, the operations were performed without the use of an endotracheal tube and with the patients totally awake and breathing normally. The age ranged from 51-75 years with the predominant male sex. Epidural thoracic administratios of the anesthesia was performed. Surgery was performed through a habitual anterolateral thoracotomy. During the entire procedure, the left lung remained partially collapsed.RESULTS: The 24 patients progressed well through the surgery. Pneumothorax time ranged from 70-190 minutes. No electrocardiographic, echocardiographic, or enzymatic alterations occurred that characterized pre- and postoperative infarcts. Twenty-three patients were stable enough to be released after 24 hours.CONCLUSION: This technique could be performed on an large number of selected patients. However, more experience is necessary.

Comparison of the Proportion Method With Mycobacteria Growth Indicator Tube and E-test for Susceptibility Testing of Mycobacterium tuberculosis

The aim of this study was to investigate the correlation between proportion method with mycobacteria growth indicator tube (MGIT) and E-test for Mycobacterium tuberculosis. Forty clinical isolates were tested. MGIT and E-test with the first line antituberculous drugs correlated with the proportion method. Our results suggested that MGIT and E-test methods can be routinely used instead of the proportion method.

One-tube nested Polymerase Chain Reaction for detection of Chlamydia trachomatis

Here we present a one-tube nested PCR test, which allows the detection of minimal quantities of Chlamydia trachomatis in human fluids. This assay includes the use of an internal control to avoid false negative results due to the presence of inhibitors. The results obtained show that this assay is robust enough to be used for clinical diagnosis.

Serial QuantiFERON-TB Gold In-Tube assay and tuberculin skin test to diagnose latent tuberculosis in household Mexican contacts: conversion and reversion rates and associated factors using conventional and borderline zone definitions

A cohort of 123 adult contacts was followed for 18‐24 months (86 completed the follow-up) to compare conversion and reversion rates based on two serial measures of QuantiFERON (QFT) and tuberculin skin test (TST) (PPD from TUBERSOL, Aventis Pasteur, Canada) for diagnosing latent tuberculosis (TB) in household contacts of TB patients using conventional (C) and borderline zone (BZ) definitions. Questionnaires were used to obtain information regarding TB exposure, TB risk factors and socio-demographic data. QFT (IU/mL) conversion was defined as <0.35 to ≥0.35 (C) or <0.35 to >0.70 (BZ) and reversion was defined as ≥0.35 to <0.35 (C) or ≥0.35 to <0.20 (BZ); TST (mm) conversion was defined as <5 to ≥5 (C) or <5 to >10 (BZ) and reversion was defined as ≥5 to <5 (C). The QFT conversion and reversion rates were 10.5% and 7% with C and 8.1% and 4.7% with the BZ definitions, respectively. The TST rates were higher compared with QFT, especially with the C definitions (conversion 23.3%, reversion 9.3%). The QFT conversion and reversion rates were higher for TST ≥5; for TST, both rates were lower for QFT <0.35. No risk factors were associated with the probability of converting or reverting. The inconsistency and apparent randomness of serial testing is confusing and adds to the limitations of these tests and definitions to follow-up close TB contacts.

Comparison of standardized uptake values measured on 18F-NaF PET/CT scans using three different tube current intensities

Objective: To analyze standardized uptake values (SUVs) using three different tube current intensities for attenuation correction on 18FNaF PET/CT scans. Materials and Methods: A total of 254 18F-NaF PET/CT studies were analyzed using 10, 20 and 30 mAs. The SUVs were calculated in volumes of interest (VOIs) drawn on three skeletal regions, namely, right proximal humeral diaphysis (RH), right proximal femoral diaphysis (RF), and first lumbar vertebra (LV1) in a total of 712 VOIs. The analyses covered 675 regions classified as normal (236 RH, 232 RF, and 207 LV1). Results: Mean SUV for each skeletal region was 3.8, 5.4 and 14.4 for RH, RF, and LV1, respectively. As the studies were grouped according to mAs value, the mean SUV values were 3.8, 3.9 and 3.7 for 10, 20 and 30 mAs, respectively, in the RH region; 5.4, 5.5 and 5.4 for 10, 20 and 30 mAs, respectively, in the RF region; 13.8, 14.9 and 14.5 for 10, 20 and 30 mAs, respectively, in the LV1 region. Conclusion: The three tube current values yielded similar results for SUV calculation.

In vitro effect of substrate, temperature and photoperiod on Phakopsora pachyrhizi urediniospore germination and germ tube growth

In vitro experiments were conducted to assess the effects of substrate, temperature and time of exposure to temperature and photoperiod on P. pachyrhizi uredospore germination and germ tube growth. The following substrates were tested: water-agar and soybean leaf extract-agar at different leaf concentrations (0.5, 1.0, 2.0 and 4.0 g of leaves and 15g agar/L water), temperatures (10, 15, 20, 25, 30, and 35oC) and times of exposure (1, 2, 3, 4, 5, 6, 7, and 8 hours) to temperature and 12 different photoperiods. The highest germination and germ tube length was found for the soybean leaf extract agar. Maximum P. pachyrhizi uredospore germination was obtained at 21.8 and 22.3°C, and maximum germ tube growth at 21.4 and 22.1°C. The maximum uredospore germination was found at 6.4 hours exposure, while the maximum germ tube length was obtained at 7.7 h exposure. Regarding photoperiod, the maximum spore germination and the maximum uredospore germ tube length were found in the dark. Neither spore germination nor uredospore germ tube growth was completely inhibited by the exposure to continuous light.

Diversidad florística arbórea y su relación con el suelo en un bosque pluvial tropical del chocó biogeográfico

El objetivo del presente estudio fue determinar la composición y diversidad florística arbórea a escala local en un bosque del Chocó biogeográfico, una de las regiones más lluviosas del mundo, y su relación con las características del sitio. Se emplearon dos parcelas de una hectárea, en las cuales se determinó la composición y diversidad de árboles con DAP > 10 cm y se relacionaron con los nutrientes del suelo, la topografíay el espacio. No se presentaron diferencias significativas entre las parcelas para las variables estructurales y de diversidad estudiadas, con valores de abundancia de 640 y 716 individuos/ha yde riqueza de 174 y 223 especies/ha.Nuestros resultados soportan la hipótesis de hiperdominancia de unos pocos grupos biológicos en los bosques húmedos tropicales. Ladiversidad arbórea a escala local se relacionó significativamente con algunas condiciones edáficas.Se presentó asociación positiva de la diversidad con el aluminio del suelo, y negativa con la topografía.No obstante, solamente un 37% de la variación en la composición florística fue explicada por las variables espaciales y ambientales (topográficas y edáficas). Por ello, aparentemente la distribución de las especies a escala local parece estar gobernada principalmente por procesos aleatorios o biológicos (como la limitación en dispersión), conforme lo propone la teoría neutral. El papel de las adaptaciones específicas al hábitat (teoría del nicho) pareciera ser limitado.