Gram-negative pulmonary infections - advances in epidemiology and diagnosis.

PURPOSE OF REVIEW: Pulmonary infections due to Gram-negative organisms are increasing worldwide and traditional assumptions that these are limited to hospital and ventilator-acquired pneumonia are rapidly falling away. Accordingly, empiric antibiotic guidelines have to follow suit with ever broader spectrum choices in order to remain 'safe', as the Global prevalence of extensively resistant Gram-negative organisms inexorably increases. Rapid, multiplex PCR-based detection of a wide variety of potential pathogens offers the opportunity to replace empiric antibiotic choices with targeted, evidence-based therapy in clinically actionable timeframes. RECENT FINDINGS: Here, we describe the data underpinning both the increasing global prevalence of Gram-negative pulmonary infections and their increasing antibiotic resistance. We also describe the performance, characteristics and early emerging clinical impact of already available rapid molecular diagnostic platforms and how they might best be deployed. SUMMARY: It will likely be advantageous to replace the current trend for empiric prescription of increasingly broad-spectrum antibiotics with 'same day' evidence-based, targeted therapy using high performance, rapid molecular diagnostic solutions. Several challenges remain be overcome, however, to fully realize their clear potential for better, focussed deployment of antibiotics, improved patient outcomes and antibiotic stewardship.

THE PERFORMANCE OF A RAPID TEST FOR ANTI-HCV SCREENING IN ORAL FLUIDS.

UNLABELLED: Gingival crevicular fluid (GCF) and saliva samples provide advantages for screening or sero-prevalence studies on HCV using less invasive methods. The study aimed to evaluate the performance of a rapid test for HCV-antibodies (HCV-Ab) screening in oral fluids among high-risk individuals with chronic liver disease. METHODS: Chronic liver disease patients attending at the Matei Bals National Instiute for Infectious Diseases were recruited for this study. Plasma, GCF and saliva samples (pair samples) were collected from each patient included in the study. Forty-three sample pairs were tested with Laboquick (Koroglu Medical Devices) rapid test and ELISA (DIA.PRO--Diagnostic Bio-probes) for the detection of anti-HCV antibodies. RESULTS: Using rapid test, anti-HCV antibodies were detected in 36 GCFs (83.72%) and 24 saliva cases (55.8%) of infected subjects. For a better estimation of oral fluids positivity, the cut-off values were calculated following plotting the ROC curves (COV2). Comparing Laboquick and ELISA (COV2) data, matched results were noted in 95.3 % saliva samples and 93% GCF samples. CONCLUSIONS: Oral fluids could be an alternative to blood for detection of HCV-positive subjects. Anti-HCV rapid test may be useful in routine dental medicine.

Clinical relevance of the plasma load of cytomegalovirus in patients infected with HIV--a survival analysis.

To investigate whether asymptomatic cytomegalovirus (CMV) viraemia impact the course of human immunodeficiency virus (HIV) infection, this study evaluated the effect of CMV replication on progression of newly-diagnosed HIV infected individuals towards AIDS events and death. In a 3-year prospective study on co-infected patients, clinical, immunological, and virological tests were performed in a national reference hospital quarterly. CMV viraemia was quantified by RoboGene® HCMV DNA Quantification Kit (Analytik Jena, Germany), on ABI Prism® 7000 Sequence Detection System (Applied Biosystems, USA). One hundred and five patients were enrolled with a balanced sex distribution and a median age of 30.7 years. Median CD4(+) cell count at enrollment was 164/mm(3) and median HIV RNA 4.6 log10 copies/ml. Detectable CMV viraemia was found in 25.7% of the patients. Kaplan-Meier analysis showed progression of HIV infection to be significantly increased in those with active CMV replication and/or low CD4(+) cell count. Cox regression indicated the risk of developing new AIDS events was 2.6 times greater in patients with detectable CMV viraemia versus those without (CI95% 1-6.6; P = 0.04). Also in multivariate analysis, the overall risk of progression to AIDS events or death was 3-fold higher in those with detectable CMV viraemia (CI95% 1.3-6.7; P = 0.008) and 2.3-fold higher if CD4(+) cell count was below 100/mm(3) (CI95% 1-5.1; P = 0.04). In these young Romanian HIV-seropositives, active CMV replication increased morbidity, even when treated with combination antiretroviral therapy. Further studies are needed to evaluate if serial quantitative CMV-DNA levels might correlate with non-infectious inflammation-related risks in patients with HIV and active CMV infection.

Update on Kaposi sarcoma-associated herpesvirus (KSHV or HHV8) - review.

Human herpesvirus 8 (HHV8), also known as Kaposi sarcoma-associated herpesvirus (KSHV), is one of the few pathogens recognized as direct carcinogen, being involved in the pathogenesis of Kaposi sarcoma, primary effusion lymphoma and multicentric Castleman disease. KSHV is a relatively recently discovered virus, with still limited possibilities for diagnosis and treatment. Therefore, ongoing studies are trying to answer the main issues related to the management of KSHV infection and its associated diseases. This review updates the current knowledge of the KSHV infection, discussing aspects related to epidemiology, virological features, clinical manifestations, diagnosis and treatment.

Hepatitis B and C Virus Reactivation Patterns in a Romanian Cohort of Patients with Chronic Lymphoproliferative Disorders.

Hepatitis B and C virus (HBV and HCV) reactivations have become more common following the intensive use of biological therapies for the treatment of chronic lymphoproliferative disorders (CLD). We evaluate risk factors for virus reactivation and exitus in patients diagnosed with CLD and HBV or HCV infection, undergoing rituximab-chemotherapy (R-chemo). A prospective, observational study in two tertiary-care Romanian hospitals, between December 2007 and May 2010, of patients diagnosed with CLD undergoing R-chemo. HBV and HCV serological markers, viral load, fibrosis and necroinflammation were assessed at baseline and every 3-6 months. We screened 502 patients diagnosed with CLDs (77.2% non-Hodgkin lymphomas) and enrolled 57 patients with HBV and/or HCV infection with a mean age of 61.35 ± 11.1 years. The replicative virus was HBV in 23 patients (40.3%), HCV in 33 patients (57.9%). HCV reactivation rate (15.6%) was lower than for HBV (45.5%) (p = 0.02). In univariate analysis, viral reactivation was associated with aggressive CLD (p = 0.01), HBV (p = 0.01) and lymphopenia (p = 0.02). Death was associated with aggressive CLD (p = 0.01), viral reactivation (p = 0.001) and high baseline viremia (p = 0.05). In multivariate analysis, viral reactivation was associated with lymphopenia (OR 0.05, 95% CI 0.003-0.85, p = 0.03). Risk of death was 10 times higher for patients with viral reactivation (95% CI 1.54-65.5, p = 0.01). A quarter of the infected patients were diagnosed with viral reactivation. While hepatitis C was more prevalent than hepatitis B in patients with CLD, viral reactivation was found 3 times more frequently in patients with hepatitis B than C.

Leptin expression in HIV-infected patients during antiretroviral therapy.

BACKGROUND: Leptin is an adipokine with complex metabolic, neuroendocrine and immune functions. Our objective was to evaluate leptin serum levels in a cohort of Romanian HIV-infected patients undergoing antiretroviral therapy in relation to their immune-virological status, lipid and glucose metabolic abnormalities and the presence of metabolic syndrome (MS). METHODS: We enrolled consecutive non-diabetic HIV-infected patients aged 18 and over on stable cART for at least 6 months. Blood samples were tested for: leptin, CD4 T cells count, HIV viral load and lipid panel. RESULTS: A total of 90 HIV-infected patients were included in the study: 50 males (55.6%) with a mean age of 33.3 years and 40 females with a mean age of 30.4 years. Most patients (74.4%) had HIV viral load below the limit of detection and the median CD4 count for the cohort was 476 (410) cells/cmm. More than one third of the patients (41.1%) had hypoleptinemia. The prevalence of MS was 13.3%. Hypoleptinemia was significantly more frequent in men. In a subset of patients with undetectable HIV viral load, the median leptin value was 0.6 (6.07) ng/mL in patients with poor immune recovery (CD4 count ≤ 200/cmm) compared to 2 (3.07) ng/mL for those with better immune response (CD4 count > 200/cmm), without statistical significance. The median values of leptin were similar for persons with and without MS criteria. HDL-cholesterol values were positively correlated to leptin values in a linear regression model. CONCLUSION: A significant proportion of patients in our study presented low levels of leptin; this finding was not associated with immune and virological parameters or the presence of MS. Hypoleptinemia was significantly correlated with lower levels of HDL-cholesterol, a key cardiovascular risk factor.

Evaluation of bacterial translocation in patients with chronic HCV infection.

INTRODUCTION: The factors involved in the progression of liver disease towards decompensated cirrhosis are not completely elucidated. It seems that bacterial translocation (BT) from the gut to the systemic blood flow has an important role in the disease progression, but literature data are controversial. Our objectives were to evaluate the presence of BT in patients with chronic HCV infection and to assess the correlation between BT and liver fibrosis stages and inflammatory state. METHODS: We conducted a cross-sectional study on patients with chronic HCV infection in a tertiary care hospital between January and July 2013. Blood samples were collected for aspartate aminotransferase (AST), alanine aminotransferase (ALT), gamma glutamyl transpeptidase (GGT), total cholesterol, low density lipoprotein (LDL) cholesterol, triglycerides, platelets, lipopolysaccharide (LPS) and tumor necrosis-alpha (TNFα). Plasma LPS were measured by ELISA (Kamiya Biomedical Company Seattle, SUA) and TNFα by DIAsource ImmunoAssay (Louvain-la-Neuve, Belgium) kits. Liver fibrosis was evaluated by means of FibroMax (BioPredictive, Paris, France) in all patients. RESULTS: We enrolled 116 patients with CHC, with a sex ratio M/F of 0.55 and a median age of 54 (45-61) years. Most of the patients (32) had compensated cirrhosis (F4). LPS levels were higher in patients with mild fibrosis--median value of 60.34 (32-91.7) ng/mL, than in cirrhotic patients--median value 40.39 (20.2-74.4) ng/mL (p = 0.051). We found no statistical correlation between LPS levels and fibrosis (p = 0.068) or TNFα levels (p = 0.097) CONCLUSIONS: There was BT in patients with CHC but it was not correlated with liver fibrosis stages or systemic inflammation. This may suggest that LPS evaluation may not be the best technique to assess baterial translocation, but further studies are needed.

Reactivation of HBV infection in low grade lymphoma patient.

Reactivation of hepatitis B virus is a complication of chronic or HBV infection in patients with malignancies, especially hematological disorders, under cytotoxic or immunosuppressive therapy. The immunosuppression favors HBV replication with the massive infection of hepatocytes. Once immunity is restored when chemotherapy therapy is discontinued, a rapid, immune-mediated destruction of the infected hepatocytes ensues, clinically manifested as hepatitis, liver failure or even death. We report a case of HBV reactivation in a patient with B cells non-Hodgkin lymphoma, with HBsAg negative and protective titre of anti-HBs, after 5 months of combined chemotherapy. Currently, there are no data to support routine pre-emptive anti-HBV therapy in patients with negative HBsAg and undetectable viremia before the initiation of chemotherapy. The case presented in this paper is included in the group of patients that is studied in LIMFOVIR Grant (convention no 41012/2007). This research grant is funded by the National Center of Programs Management, program 4 - Partnerships in Priority Fields. The grant is coordinated by the National Institute of Infectious Diseases Prof. Dr. Matei Bals, Bucharest. The grant team include also the Emergency University Hospital Bucharest, Hematology Department, the "Carol Davila" University of Medicine and Pharmacy, Bucharest, the "Victor Babes" National Institute of Research and Development, the Institute of Electrotechnical Research, Bucharest and the Polytechnic University, Bucharest. The manager of the grant is Associated Professor dr. Victoria Arama.

Impact of highly active antiretroviral therapy on cytomegalovirus viraemia in the absence of specific anti-cytomegalovirus therapy.

OBJECTIVES: (1) to evaluate the effect of HAART on CMV viraemia in co-infected patients, in the absence of specific anti-CMV therapy; (2) to compare 2 molecular biology techniques for the detection and quantification of CMV-DNA in these patients. METHODS: We present the preliminary data of an ongoing prospective research grant on newly diagnosed HIV seropositives, in a tertiary care hospital, during June 2006- June 2008. Clinical, virological (HIV and CMV viraemia) and immunological (CD4) screening was performed every 3 months. The CMV viraemia was performed by RoboGene Human Cytomegalovirus Quantification kit (aj Roboscreen). We retested all undetectable CMV viremia found in patients with CD4 <50/mmc, by CMV PCR kit (Qiagen Diagnostics). Both PCR reactions were performed on ABI Prism 7000 (Applied Biosystems). RESULTS: Up to date, our study has included 105 HIV-infected subjects, who were seropositive for anti-CMV IgG antibodies. Average follow-up was 18 months. CMV viraemia was found detectable in 21 cases at first visit and in other 5 at the second visit. 22 cases had CD4 <50/mmc, among which 14 had undetectable CMV viraemia. The results of both molecular biology techniques were widely the same. HAART was prescribed to 86% of the patients; all the patients having detectable CMV viraemia received HAART, but not any specific anti-CMV therapy. Under HAART, all the detectable CMV loads which were retested in time became undetectable at next visits, after a median of 16.5 weeks from the introduction of therapy. CONCLUSIONS: CMV viraemia detection was useful in early diagnosis of asymptomatic CMV infection. As opposed to transplant cases, molecular biology techniques for the detection and quantification of CMV-DNA in HIV-patients have not been standardized yet. In our study, the two kits RoboGene Human Cytomegalovirus (HCMV) Quantification kit (aj Roboscreen) and CMV PCR kit (Qiagen Diagnostics) were comparable. HAART made the reduction of CMV viral load, without any specific anti-CMV therapy. As in the case of other opportunistic infections, undetectable natural history of CMV infection seemed to have been improved by controlling HIV infection.

Predictors of tuberculosis in acute aseptic meningitis syndrome.

Tuberculous meningitis (TBM) is a medical emergency. Early and accurate diagnosis is crucial for preventing morbidity and mortality. Our aim was to define the predictors of tuberculous rather than other causes of acute aseptic meningitis syndrome (AAMS). A retrospective study of 225 patients with AAMS admitted in a tertiary care infectious diseases center between 2001 and 2004 was performed. In order to assess the value of clinical and biological variables for the diagnosis of TBM, we compared the variables between two groups of patients, one with microbiologically documented TBM and one with certain non-TBM acute lymphocytic meningitis. The assessed variables had good specificities, but relatively low sensitivities, ruling in the diagnosis of TBM rather than ruling it out. The most reliable predictors in multivariate analysis were: duration of symptoms before admission > 7 days, altered clinical stage, CSF/blood glucose ratio < 0.5 and CSF protein concentration > 200mg/ml. Based on the multivariate model which retained four variables, the probability of TBM in a certain patient could have been increased from 17% to 99.9% when all variables were present, or decreased to 0% when all variables were absent. This study suggests that simple clinical and laboratory features are useful in diagnosing TBM in a high-prevalence region.