Evaluation of a flow cytometric test for G6PD-deficient erythrocytes.

OBJECTIVE: Glucose-6-phosphate dehydrogenase (G6PD) deficiency, an X-linked recessive disorder, is the commonest erythrocytic enzymopathy worldwide. Reliable diagnosis and severity prediction in G6PD-deficient/heterozygous females remain challenging. A recently developed flow cytometric test for G6PD deficiency has shown promise in precisely identifying deficient females. This paper presents our experiences with this test in a subtropical setting and presents a modification in flow cytometric data acquisition strategy. METHODS: The methaemoglobin reduction + ferryl Hb generation-based flow cytometric G6PD test was compared with the screening methaemoglobin reduction test (MRT) and confirmatory G6PD enzyme activity assay (EAA) in 20 G6PD-deficient males, 22 G6PD-heterozygous/deficient females and 20 controls. Stained cells were also assessed for bright/dim G6PD activity under a fluorescent microscope. RESULTS: Flow cytometry separated and quantified %bright cells in heterozygous/deficient females, objectively classifying them into 6 normal (>85% bright cells), 14 intermediate (10-85%) and two G6PD-deficient (<10% bright cells). Concordance with MRT was 89% (55/62 cases) and with EAA was 77% (48/62 cases). Fluorometrically predicted violet laser excitation (405-nm) with signal acquisition in the 425-475 nm region was a technical advancement noted for the first time in this paper. CONCLUSION: Flow cytometry/fluorescence microscopy represent technically straightforward methods for the detection and quantification of G6PD-deficient erythrocytes. Based on our results, we recommend their application as a first-line investigation to screen females who are prescribed an oxidant drug like primaquine or dapsone.

Characterization of total adenosine deaminase activity (ADA) and its isoenzymes in saliva and serum in health and inflammatory conditions in four different species: an analytical and clinical validation pilot study.

BACKGROUND: Measurement of adenosine deaminase (ADA) can provide information about cell-mediated immunity. This report's objective was to study the enzymatic activity of total ADA (tADA) and its isoenzymes ADA1 and ADA2 in canine, equine, porcine, and bovine serum and saliva and their changes in different inflammatory situations in each species. Besides, an automated method for ADA2 measurement was developed and validated. RESULTS: tADA was present in serum and saliva of healthy animals of the four species. Erythro-9-(2-hydroxy-3-nonyl) adenine (EHNA) concentration of 0.47 mM was needed for ADA1 inhibition in canine and porcine samples (serum and saliva) and bovine saliva, whereas for equine saliva 0.94 mM was needed. ADA2 activity was not detected in bovine serum and was very low or absent in equine serum and bovine saliva. An automated procedure to measure ADA2 consisting of adding EHNA to a commercial reagent for tADA measurement provided repetitive (coefficients of variation < 8.8% in serum and < 10% in saliva) and accurate (linearity of serial sample dilutions with R2 > 0.90) results, being equivalent to a manual incubation of the sample with EHNA at a similar concentration. Salivary tADA, as well as ADA1 and ADA2, were higher in dogs with leishmaniosis, horses with acute abdominal disease and pigs with lameness than in healthy animals. tADA and isoenzymes in saliva showed a positive significant correlation with serum ferritin in dogs (r = 0.602, P < 0.01; r = 0.555, P < 0.05; and r = 0.632, P < 0.01; respectively for tADA, ADA1 and ADA2) and serum C-reactive protein in pigs (r = 0.700, P < 0.01, for both tADA and ADA1; r = 0.770, P < 0.001, for ADA2), whereas salivary ADA2 significantly correlated with serum amyloid A in horses (r = 0.649, P < 0.01). In cows, salivary tADA and ADA1 significantly increased after calving, correlating with total white blood cell count (r = 0.487, P < 0.05, for both tADA and ADA1). CONCLUSIONS: The activity of total ADA and its different isoenzymes, can be measured in serum and saliva of dogs, horses, pigs and cows by a simple and fast procedure described in this report. When measured in saliva, these analytes correlated with other biomarkers of inflammation and it could potentially be used as a biomarkers of inflammation and immune activation in the species of this study.

Leukocyte esterase strip test as a reliable intraoperative PJIs biomarker. Our experience.

ackgroud and aim of work: Prosthetic joint infection (PJI) is the most common cause of total knee replacement failure and the third most common cause of total hip replacement failure, accounting for 16.8% of all knee revisions and 14.8% of the hip revisions; nevertheless, the diagnosis of PJI is often a challenge for the orthopaedic surgeon. The aim of these study was to evaluate the reliability of the LE strip test for diagnosis of PJI. MATERIALS AND METHODS: From December 2016 to January 2019, we enrolled 50 patients with suspected PJI; 32 females and 18 males, the average age at the time of the surgery was 76 years. Twenty-four patients underwent knee revision surgery and twenty-six hip revision surgery. In all patients during the surgery, the synovial fluid was aspirated and used for leukocyte esterase strip test. The result of the tests was compared to periprosthetic tissues culture, histological examination and sonication fluid culture for PJI. RESULTS: Comparing the results obtained from the LE test with the results obtained from the other diagnostic methods, we found that the concordance between the results of the leukocyte-esterase test and those of the culture test with peri-prosthetic tissue or synovial fluid was shown to be 93%, between LE and histological examinations, the concordance was 93% and finally with the culture of the sonicated fluid the concordance was 86% of the cases. CONCLUSIONS: The results of our serie show a good intraoperative diagnostic accuracy of the LE test, especially in its ability to exclude the hypothesis of periprosthetic infection in case of a negative result.

Recent advances in histone modification and histone modifying enzyme assays.

Introduction: Histone modifying enzymes (HMEs)-catalyzed histone modifications are important epigenetic markers that play critical roles in the regulation of a variety of cellular functions, especially the regulation of gene expression. The aberrant histone modifying enzyme activity and the abnormal histone modification level are closely associated with various human diseases including cancers, making them become the promising and attractive disease biomarkers. Consequently, the development of efficient assays for accurate and sensitive detection of histone modifications and HMEs are crucial for disease diagnosis. Areas covered: In this review, we summarize the advances in histone modifications and HMEs assays in recent 5 years (2013-2018), including the development of various methods based on fluorescent, bioluminescent, colorimetric, electrochemical, surface-enhanced Raman scattering, and mass spectrometry strategies. Their principles and applications for in vitro and in vivo assays are reviewed, and the future directions are discussed as well. Expert commentary: In comparison with the conventional radioactive and Western blot assays, the newly developed histone modifications and HMEs assays exhibit distinct advantages. Especially, the introduction of novel nanomaterials and advanced analytical techniques in recent years has greatly improved the assay performances, promoting their further applications in biomedical research and clinical diagnosis.

Evaluation of the commercial kit SIRE nitratase for detecting resistant mycobacterium tuberculosis in Brazil

Abstract INTRODUCTION: This study aimed to evaluate a new commercial kit, Kit SIRE Nitratase-PlastLabor, for testing the drug susceptibility of clinical Mycobacterium tuberculosis isolates. METHODS: The accuracy of the Kit SIRE Nitratase was evaluated by examining the susceptibility (streptomycin, isoniazid, rifampicin, and ethambutol) of 40 M. tuberculosis isolates, using the proportion method with Lowenstein-Jensen medium or the BACTEC MGIT 960 system. RESULTS: The detection accuracy for streptomycin, isoniazid, rifampicin, and ethambutol was 95%, 97.5%, 100%, and 80%, respectively. CONCLUSIONS: The exceptional accuracy demonstrated by Kit SIRE Nitratase for isoniazid and rifampicin makes the kit an attractive option for screening M. tuberculosis strain resistance.

Elevation of serum lactate dehydrogenase in AL amyloidosis reflects tissue damage and is an adverse prognostic marker in patients not eligible for stem cell transplantation.

The significance of serum lactate dehydrogenase (LDH) in light chain (AL) amyloidosis has not been previously explored. We studied 1019 newly diagnosed patients and correlated the elevation of LDH above the upper limit of normal (ULN) with disease characteristics and outcome. Four hundred and nine patients had an LDH above ULN, representing 40% of the study population. Patients with an elevated LDH were older, were less likely to be male and had more extensive organ involvement compared to patients with a normal LDH. Patients with high LDH had greater cardiac and renal dysfunction. Elevated LDH was an independent prognostic marker for overall survival and for death within 6 months of diagnosis, but this was restricted to patients not eligible for stem cell transplant. Serum LDH may act as a marker for organ damage and should be explored as a potential marker for tissue healing and organ recovery.

Transient Elastography vs Aspartate Aminotransferase to Platelet Ratio Index in Hepatitis C: A Meta-Analysis

ABSTRACT Background and rationale. Many different non-invasive methods have been studied with the purpose of staging liver fibrosis. The objective of this study was verifying if transient elastography is superior to aspartate aminotransferase to platelet ratio index for staging fibrosis in patients with chronic hepatitis C. Material and methods. A systematic review with meta-analysis of studies which evaluated both non-invasive tests and used biopsy as the reference standard was performed. A random-effects model was used, anticipating heterogeneity among studies. Diagnostic odds ratio was the main effect measure, and summary receiver operating characteristic curves were created. A sensitivity analysis was planned, in which the meta-analysis would be repeated excluding each study at a time. Results. Eight studies were included in the meta-analysis. Regarding the prediction of significant fibrosis, transient elastography and aspartate aminotransferase to platelet ratio index had diagnostic odds ratios of 11.70 (95% confidence interval = 7.13-19.21) and 8.56 (95% confidence interval = 4.90-14.94) respectively. Concerning the prediction of cirrhosis, transient elastography and aspartate aminotransferase to platelet ratio index had diagnostic odds ratios of 66.49 (95% confidence interval = 23.71- 186.48) and 7.47 (95% confidence interval = 4.88-11.43) respectively. Conclusion. In conclusion, there was no evidence of significant superiority of transient elastography over aspartate aminotransferase to platelet ratio index regarding the prediction of significant fibrosis, but the former proved to be better than the latter concerning prediction of cirrhosis.

Transient Elastography vs. Aspartate Aminotransferase to Platelet Ratio Index in Hepatitis C: A Meta-Analysis.

BACKGROUND AND RATIONALE: Many different non-invasive methods have been studied with the purpose of staging liver fibrosis. The objective of this study was verifying if transient elastography is superior to aspartate aminotransferase to platelet ratio index for staging fibrosis in patients with chronic hepatitis C. MATERIAL AND METHODS: A systematic review with meta-analysis of studies which evaluated both non-invasive tests and used biopsy as the reference standard was performed. A random-effects model was used, anticipating heterogeneity among studies. Diagnostic odds ratio was the main effect measure, and summary receiver operating characteristic curves were created. A sensitivity analysis was planned, in which the meta-analysis would be repeated excluding each study at a time. RESULTS: Eight studies were included in the meta-analysis. Regarding the prediction of significant fibrosis, transient elastography and aspartate aminotransferase to platelet ratio index had diagnostic odds ratios of 11.70 (95% confidence interval = 7.13-19.21) and 8.56 (95% confidence interval = 4.90-14.94) respectively. Concerning the prediction of cirrhosis, transient elastography and aspartate aminotransferase to platelet ratio index had diagnostic odds ratios of 66.49 (95% confidence interval = 23.71-186.48) and 7.47 (95% confidence interval = 4.88-11.43) respectively. CONCLUSION: In conclusion, there was no evidence of significant superiority of transient elastography over aspartate aminotransferase to platelet ratio index regarding the prediction of significant fibrosis, but the former proved to be better than the latter concerning prediction of cirrhosis.

Association of circulating dipeptidyl-peptidase 4 levels with osteoporotic fracture in postmenopausal women.

Postmenopausal women with osteoporotic fracture (OF) had higher plasma dipeptidyl-peptidase 4 (DPP4) levels than those without. Furthermore, higher plasma DPP4 levels were significantly associated with higher bone turnover and a higher prevalence of OF. These results indicated that DPP4 may be associated with OF by mediating bone turnover rate. INTRODUCTION: Evidence indicates that dipeptidyl-peptidase 4 (DPP4) plays a distinct role in bone metabolism. However, there has been no report on the association, if any, between circulating DPP4 levels and osteoporosis-related phenotypes, including osteoporotic fracture (OF). Therefore, we performed a case-control study to investigate these associations in postmenopausal women. METHODS: This study was conducted in multiple centers in Korea. We enrolled 178 cases with OF and 178 age- and body mass index-matched controls. OF was assessed by an interviewer-assisted questionnaire and lateral thoracolumbar radiographs. Bone turnover markers (BTMs), bone mineral density (BMD), and plasma DPP4 levels were obtained in all subjects. RESULTS: After adjustment for potential confounders, subjects with OF had significantly higher DPP4 levels than those without (P = 0.021). Higher DPP4 levels were significantly positively associated with higher levels of all BTMs, but not with BMD at all measured sites. The differences in DPP4 levels according to OF status disappeared after an additional adjustment for each BTM, but not after adjustment for any BMD values. BTMs explained approximately half of the relationship between DPP4 and OF. The risk of OF was 3.80-fold (95% confidence interval = 1.53-9.42) higher in subjects in the highest DPP4 quartile than in those in the lowest quartile after adjustment for potential confounders, including femoral neck BMD. CONCLUSIONS: DPP4 may be associated with OF by at least partly mediating the bone turnover rate. Circulating DPP4 levels may be a potential biomarker that could increase the predictive power of current fracture risk assessment models.

Correlation between liver cell necrosis and circulating alanine aminotransferase after ischaemia/reperfusion injuries in the rat liver.

Circulating liver enzymes such as alanine transaminase are often used as markers of hepatocellular damage. Ischaemia/reperfusion (I/R) injury is an inevitable consequence of prolonged liver ischaemia. The aim of this study was to examine the correlation between liver enzymes and volume of liver cell necrosis after ischaemia/reperfusion injuries, using design-unbiased stereological methods. Forty-seven male Wistar rats were subjected to 1 h of partial liver ischaemia, followed by either 4 or 24 h of reperfusion. Within each group, one-third of animals were subjected to ischaemic preconditioning and one-third to ischaemic postconditioning. At the end of reperfusion, blood and liver samples were collected for analysis. The volume of necrotic liver tissue was subsequently correlated to circulating markers of I/R injury. Correlation between histological findings and circulating markers was performed using Pearson's correlation coefficient. Alanine transferase peaked after 4 h of reperfusion; however, at this time-point, only mild necrosis was observed, with a Pearson's correlation coefficient of 0.663 (P = 0.001). After 24 h of reperfusion, alanine aminotransferase was found to be highly correlated to the degree of hepatocellular necrosis R = 0.836 (P = 0.000). Furthermore, alkaline phosphatase (R = 0.806) and α-2-macroglobulin (R = 0.655) levels were also correlated with the degree of necrosis. We show for the first time that there is a close correlation between the volume of hepatocellular necrosis and alanine aminotransferase levels in a model of I/R injury. This is especially apparent after 24 h of reperfusion. Similarly, increased levels of alkaline phosphatase and α-2-macroglobulin are correlated to the volume of liver necrosis.

The relationship between liver histology and noninvasive markers in primary biliary cirrhosis.

BACKGROUND: Primary biliary cirrhosis (PBC) is a disease that affects liver with various severity and progression rates. It is important to diagnose advanced stage of the disease to lower liver-related morbidity and mortality. Since liver biopsy is an invasive method, liver biopsy tends to be replaced by noninvasive methods. In this study, we aim to show the role of aminotransferase to platelet ratio index (APRI) and fibrosis index on the basis of the four factors (FIB-4) scores, laboratory values, and their effectiveness in predicting advanced disease. MATERIALS AND METHODS: PBC patients diagnosed pathologically at Numune Education and Research Hospital were included in the study between the years 1995 and 2013. Patients were grouped according to their fibrosis level: group 1 (early stage) included 18 patients with F1 and F2 fibrosis and group 2 (advanced stage) included 22 patients with F3 and F4 fibrosis. APRI and FIB-4 scores, routine laboratory values, and their proportions were compared. The effectiveness of parameters showing advanced stage was further compared. RESULTS: There were statistically significant differences in APRI, FIB-4 scores, and aspartate aminotransferase (AST) levels between the groups with early and advanced stages of disease. Receiver operating curve analysis was used to determine APRI, FIB-4 and AST levels. The most effective parameters for diagnosing an advanced stage were APRI, AST levels, and FIB-4 scores, respectively. CONCLUSION: In conclusion, APRI and FIB-4 scores can be calculated simply and easily by routine laboratory tests at low cost and also these scores may be a predictor of advanced stage of the disease in PBC. These tests may be reproducible and may be used to monitor disease progression.

Elevated liver enzymes in inflammatory bowel disease: the role and safety of infliximab.

BACKGROUND: Abnormal liver enzymes are frequently encountered in inflammatory bowel disease (IBD) patients. Infliximab has been implicated in inducing drug-induced liver injury, autoimmune hepatitis or reactivation of hepatitis B virus. We aimed to clarify the role of infliximab in liver impairment in an IBD cohort. STUDY: A total of 305 patients with IBD, without evidence of chronic liver disease, were included in the study and retrospectively evaluated. Laboratory and clinical data were retrieved from a prospectively acquired database. In all, 176 consecutive patients treated with infliximab during the last 5 years were compared with a matched population of 129 patients who did not receive any antitumour necrosis factor treatment. RESULTS: Elevation of alanine transaminase (ALT) was frequent in the entire population (36.4%) and it was not significantly associated with the use of infliximab (P=0.284). Elevations more than 3 upper limit of normal were observed in 7.9% and these resolved spontaneously in 83%. The use of immunomodulators was the only factor that was significantly associated with liver enzyme abnormalities in multivariate analysis [odds ratio (OR) 2.666, 95% confidence interval (CI) 1.576-4.511, P<0.005]. Overall, 39% of patients on infliximab had elevated liver enzymes and this was associated with increased ALT before starting infliximab (OR 3.854, 95% CI 1.800-8.251, P=0.001) and with longer duration of infliximab treatment (OR 1.030, 95% CI 1.013-1.047, P=0.001). CONCLUSION: Elevated liver enzymes are frequently found in IBD patients and they usually resolve spontaneously. The use of immunomodulators was independently associated with increased ALT. Infliximab is relatively safe in terms of liver impairment and discontinuation of treatment is rarely required in the setting of modest elevations of ALT.

Direct measurement of catalase activity in living cells and tissue biopsies.

Spatiotemporal regulation of enzyme-substrate interactions governs the decision-making steps in biological systems. Enzymes, being functional units of every living cell, contribute to the macromolecular stability of cell survival, proliferation and hence are vital windows to unraveling the biological complexity. Experimental measurements capturing this dynamics of enzyme-substrate interactions in real time add value to this understanding. Furthermore these measurements, upon validation in realistic biological specimens such as clinical biopsies - can further improve our capability in disease diagnostics and treatment monitoring. Towards this direction, we describe here a novel, high-sensitive measurement system for measuring diffusion-limited enzyme-substrate kinetics in real time. Using catalase (enzyme) and hydrogen peroxide (substrate) as the example pair, we demonstrate that this system is capable of direct measurement of catalase activity in vitro and the measured kinetics follows the classical Michaelis-Menten reaction kinetics. We further demonstrate the system performance by measuring catalase activity in living cells and in very small amounts of liver biopsies (down to 1 µg total protein). Catalase-specific enzyme activity is demonstrated by genetic and pharmacological tools. Finally we show the clinically-relevant diagnostic capability of our system by comparing the catalase activities in liver biopsies from young and old mouse (liver and serum) samples. We discuss the potential applicability of this system in clinical diagnostics as well as in intraoperative surgical settings.

Clinical and laboratory description of a series of cases of acute viral myositis / Descrição clínico-laboratorial de uma série de casos de Miosite Aguda Viral

ABSTRACT OBJECTIVE: Describe the clinical and laboratory profile, follow-up, and outcome of a series of cases of acute viral myositis. METHOD: A retrospective analysis of suspected cases under observation in the emergency department was performed, including outpatient follow-up with the recording of respiratory infection and musculoskeletal symptoms, measurement of muscle enzymes, creatine phosphokinase (CPK), lactate dehydrogenase (LDH), transaminases (AST and ALT), blood count, C-reactive protein, and erythrocyte sedimentation rate in the acute phase and during follow-up until normalization. RESULTS: Between 2000 and 2009, 42 suspected cases were identified and 35 (27 boys) were included. The median age was 7 years and the diagnosis was reported in 89% in the first emergency visit. The observed respiratory symptoms were cough (31%), rhinorrhea (23%), and fever (63%), with a mean duration of 4.3 days. Musculoskeletal symptoms were localized pain in the calves (80%), limited ambulation (57%), gait abnormality (40%), and muscle weakness in the lower limbs (71%), with a mean duration of 3.6 days. There was significant increase in CPK enzymes (5507 ± 9180 U/L), LDH (827 ± 598 U/L), and AST (199 ± 245 U/L), with a tendency to leukopenia (4590 ± 1420) leukocytes/mm3. The complete recovery of laboratory parameters was observed in 30 days (median), and laboratory and clinical recurrence was documented in one case after 10 months. CONCLUSION: Typical symptoms with increased muscle enzymes after diagnosis of influenza and self-limited course of the disease were the clues to the diagnosis. The increase in muscle enzymes indicate transient myotropic activity related to seasonal influenza, which should be considered, regardless of the viral identification, possibly associated with influenza virus or other respiratory viruses.

RESUMO OBJETIVO: Descrever o perfil clínico-laboratorial, o acompanhamento e o desfecho de uma série de casos de miosite aguda viral. MÉTODO: Foi conduzida uma análise retrospectiva de casos suspeitos, em observação em unidade de emergência, e seguimento ambulatorial com o registro de sintomas de infecção respiratória, sintomas músculo-esqueléticos, determinação de enzimas musculares, creatina-fosfoquinase (CPK), desidrogenase lática (DHL), transaminases (AST e ALT), hemograma, proteína C reativa e velocidade de hemossedimentação, na fase aguda e no acompanhamento, até a normalização. RESULTADOS: Entre 2000 e 2009, 42 casos suspeitos foram identificados e 35 (27 meninos) foram incluídos. A mediana de idade foi de sete anos e o diagnóstico relatado em 89%, na primeira visita de emergência. Os sintomas respiratórios observados foram: tosse (31%), coriza (23%) e febre (63%), com duração média de 4,3 dias. Os sintomas músculo-esqueléticos foram: dor localizada nas panturrilhas (80%), deambulação limitada (57%), marcha anormal (40%) e fraqueza muscular nos membros inferiores (71%), com duração média de 3,6 dias. Observou-se elevação importante das enzimas CPK (5.507 ± 9.180) U/l, DHL (827 ± 598) U/l e AST (199 ± 245) U/l e tendência a leucopenia (4.590 ± 1.420) leucócitos/mm3. A recuperação completa dos parâmetros laboratoriais foi observada em 30 dias (mediana) e a recaída clínica e laboratorial em um caso após 10 meses. CONCLUSÃO: Os sintomas típicos com enzimas musculares elevadas após diagnóstico de influenza e o curso autolimitado foram os indícios para o diagnóstico. A elevação de enzimas musculares indica a atividade miotrópica transitória relacionada à influenza sazonal que deve ser considerada, a despeito da identificação viral, possivelmente associada com o vírus influenza ou outros vírus respiratórios.

AST/ALT ratio is not useful in predicting the degree of fibrosis in chronic viral hepatitis patients.

BACKGROUND AND AIM: Noninvasive tests are primarily used for staging hepatic fibrosis in patients with chronic liver disease. In clinical practice, serum aminotransferase levels, coagulation parameters, and platelet count have been used to predict whether or not a patient has cirrhosis. In addition, several studies have evaluated the accuracy of combinations (or ratios) of these measures. The present study aimed to investigate the relationship between five noninvasive models [AST/ALT ratio (AAR), aspartate aminotransferase to platelet ratio index (APRI), Bonacini cirrhosis discriminant score (CDS), age-platelet index (APind), and King's score] and the degree of hepatic fibrosis as determined by biopsy in patients with chronic hepatitis B and C. PATIENTS AND METHODS: A total of 380 patients with viral hepatitis (237 with chronic hepatitis B and 143 with chronic hepatitis C) who were seen at our clinic between January 2005 and January 2011 were retrospectively analyzed. The degree of fibrosis was determined using the Ishak score. Patients with a fibrosis score of 0-2 were considered to have low fibrosis and those with a score between 3 and 6 were considered to have high fibrosis. Five noninvasive models were compared between the groups with low and high fibrosis. RESULTS: There were statistically significant differences between the hepatitis B and C patients with high and low fibrosis with respect to APind (4.49±2.35 vs. 2.41±1.84; P<0.001 in hepatitis B and 4.83±2.25 vs. 2.92±1.88; P<0.001 in hepatitis C), APRI (1.00±1.17 vs. 0.47±0.39; P<0.001 in hepatitis B and 1.01±1.01 vs. 0.41±0.29; P<0.001 in hepatitis C), CDS (4.53±1.90 vs. 3.58±1.30; P<0.001 in hepatitis B and 4.71±2.03 vs. 3.42±1.49; P<0.05 in hepatitis C), and King's score (24.31±3.14 vs. 7.65±6.70; P<0.001 in hepatitis B and 24.82±2.55 vs. 8.33±7.29; P<0.001 in hepatitis C). There were no significant differences in the AAR between the hepatitis B and C patients with high and low fibrosis (0.78±0.31 vs. 0.74±0.34; P=0.082 in hepatitis B and 0.91±0.40 vs. 0.85±0.27; P=0.25 in hepatitis C). The area under the receiver-operating characteristic curve of the APind, APRI, CDS, and King's score in the hepatitis B group were 0.767, 0.710, 0.646, and 0.770, respectively; these values were 0.732, 0.763, 0.677, and 0.783, respectively, in the hepatitis C group. CONCLUSION: In conclusion, our data suggest that four of the five noninvasive methods evaluated in this study can be used to predict advanced fibrosis in patients with hepatitis B and C. However, there was no significant relationship between the degree of hepatic fibrosis and the AAR score, indicating that AAR is not useful in estimating the fibrosis stage in hepatitis B and C patients.

Progression of liver disease in non-alcoholic fatty liver disease: a prospective clinicopathological follow-up study.

OBJECTIVE: To perform a follow-up study on non-alcoholic fatty liver disease (NAFLD) patients in our previous study using paired liver biopsy. METHODS: Patients who were included in our previous study on NAFLD and agreed to receive a repeat liver biopsy were included in the study. Their clinical characteristics, laboratory examination results and histological analysis on the repeat liver biopsied specimens were prospectively collected and compared with those in the previous study. RESULTS: Data from 35 patients (mean age 47.5 ± 10.9 years, male 40.0%) were analyzed. The mean interval between the liver biopsies was 6.4 ± 0.8 years. NAFLD activity score (NAS) worsened in 13, remained unchanged in 9 and ameliorated in 13. Fibrosis worsened in 18 and remained unchanged in 17. Two patients who were confirmed with cirrhosis at baseline developed decompensated cirrhosis. On multivariate analysis, elevated serum aspartate aminotransferase (AST) (odds ratio [OR] 10.74, 95% confidence interval [CI] 1.00-115.86, P = 0.050) and γ-glutamyl transpeptidase (γ-GT) (OR 16.10, 95% CI 1.30-198.90, P = 0.030) at follow-up were associated with worsened NAS. Patients with borderline NASH at baseline were more likely to have worsened NAS at follow-up than those with definite NASH (OR 12.67, 95% CI 2.29-70.02, P = 0.004). However, both groups had a similar likelihood of having worsened fibrosis at follow-up. No plausible factors were found to be associated with worsened fibrosis. CONCLUSIONS: NAFLD patients with persistently elevated serum AST and γ-GT levels during follow-up should be suspected of having worsened NAS. NASH patients can have significant disease progression over a relatively short period of time and fibrosis might be irreversible without specific interventions.

Prevalência de carbapenemases em enterobactérias resistentes a carbapenêmicos em quatro hospitais terciários de Porto Alegre / Prevalence of carbapenemases in carbapenem-resistant Enterobacteriaceae in four tertiary care hospitals in Porto Alegre

INTRODUÇÃO: A disseminação de Klebsiella pneumoniae carbapenemase (KPC) no Brasil e a recente detecção de bactérias produtoras de New Delhi metalo-β-lactamase (NDM-1) em hospital terciário do sul do Brasil indicam a necessidade da avaliação da presença destas enzimas em enterobactérias resistentes a carbapenêmicos (ERC).OBJETIVO: Avaliar prevalência de carbapenemases nas ERC em quatro hospitais terciários de Porto Alegre, por meio de PCR multiplex em tempo real. MÉTODOS: Estudo descritivo, período de abril a dezembro de 2013. Isolados bacterianos de pacientes internados foram identificados pelo sistema automatizado VITEK 2, com realização do teste de suscetibilidade aos antimicrobianos. Amostras com isolados de ERC foram encaminhadas ao laboratório de referência para análise por PCR em tempo real para identificação de carbapenemases. RESULTADOS: Total de 701 isolados. As ERC predominantes foram K. pneumoniae (47% das amostras positivas) e Enterobacter cloacae (18%). As carbapenemases mais frequentes foram KPC (48%), OXA-48-like (3%) e NDM (2%). Em 47% das amostras não foi identificado o mecanismo de resistência. Isolados originados de culturas de vigilância foram associados com maior positividade para carbapenemases do que isolados de amostras clínicas (p<0,0001). Isolados de ERC pertencentes ao grupo Proteae (Proteus spp., Morganella spp., Providencia spp.) foram associados a menor positividade para carbapenemase do que isolados de outras ERC (p<0,0001). CONCLUSÃO: KPC foi a carbapenemase mais frequentemente detectada. A circulação de uma enzima OXA-48-like foi demonstrada, um achado novo e preocupante. O achado da carbapenemase NDM também é preocupante devido ao seu potencial de disseminação. Esses dados e outros estudos poderão contribuir para um entendimento maior da epidemiologia das ERC.

BACKGROUND: The spread of Klebsiella pneumoniae carbapenemase (KPC) in Brazil and the recent detection of bacteria producing New Delhi metallo-β-lactamase (NDM-1) in a tertiary care hospital in Porto Alegre indicate the need to evaluate the presence of these enzymes in carbapenem-resistant Enterobacteriaceae (CRE). AIM: To evaluate the prevalence of carbapenemases in CRE in four tertiary care hospitals in Porto Alegre using multiplex real-time PCR.METHODS: Descriptive study from April to December 2013. Bacterial isolates from hospitalized patients were identified by VITEK 2 automated system, with antimicrobial susceptibility testing. Samples with CRE isolates were sent to the reference laboratory for analysis using real-time PCR for identification of carbapenemases. RESULTS: Total of 701 isolates. The predominant CRE were K. pneumoniae (46% of positive samples) and Enterobacter cloacae (18%). The most frequent carbapenemases were KPC (48%), OXA-48-like (3%), and NDM-1 (2%). In 47% of the samples no carbapenemase was identified. Isolates originated from surveillance cultures were associated with higher positivity for carbapenemases than isolates from clinical samples (p<0.0001). CRE isolates belonging to the Proteae group (Proteus spp., Morganella spp., Providencia spp.) were associated with less positivity for carbapanemase than isolates of other CRE (p<0.0001). CONCLUSION: KPC was the most frequently detected carbapenemase. The movement of an OXA-48-like enzyme was demonstrated, a novel and worrisome finding. The finding of carbapenemase NDM is also worrisome due to its dissemination potential. These data and further studies may contribute to a better understanding of the epidemiology of CRE.

Enzymes in clinical medicine: an overview.

Enzymes are biocatalysts and because of their remarkable properties, they are extensively used in medical diagnosis. Researches in the last two decades have concentrated more on enzymes such as creatine kinase-MB, alanine transaminase, aspartate transaminase, acid phosphatase, alkaline phosphatase etc. for clinical applications. Enzymes are the preferred markers in various disease states such as myocardial infarction, jaundice, pancreatitis, cancer, neurodegenerative disorders, etc. They provide insight into the disease process by diagnosis, prognosis and assessment of response therapy. Even though the literature on the use of enzymes in various disease conditions has accumulated, a comprehensive analysis is lacking and hence this review.