Individualized significance of the -251 A/T single nucleotide polymorphism of interleukin-8 in severe infections.

Based on the concept of the individualized nature of sepsis, we investigated the significance of the -251 A/T (rs4073) single nucleotide polymorphism (SNP) of interleukin (IL)-8 in relation to the underlying infection. Genotyping was performed in 479 patients with severe acute pyelonephritis (UTI, n = 146), community-acquired pneumonia (CAP, n = 109), intra-abdominal infections (IAI, n = 119), and primary bacteremia (BSI, n = 105) by restriction fragment length polymorphism of the polymerase chain reaction (PCR) product and compared with 104 healthy volunteers. Circulating IL-8 was measured within the first 24 h of diagnosis by an immunosorbent assay. Carriage of the AA genotype was protective from the development of UTI (odds ratio 0.38, p: 0.007) and CAP (odds ratio 0.30, p: 0.004), but not from IAI and BSI. Protection from the development of severe sepsis/septic shock was provided for carriers of the AA genotype among patients with UTI (odds ratio 0.15, p: 0.015). This was accompanied by greater concentrations of circulating IL-8 among patients with the AA genotype. It is concluded that carriage of rs4073 modifies susceptibility for severe infection in an individualized way. This is associated with a modulation of circulating IL-8.

Combination of vial culture and broad-range PCR for the diagnosis of spontaneous bacterial peritonitis: experience in a Greek tertiary care hospital.

Spontaneous bacterial peritonitis (SBP) is often difficult to diagnose because bacteria in ascites cannot be detected accurately by conventional culture. In this study, we evaluated the use of broad-range 16S rRNA PCR, applied either directly to a total of 32 ascitic fluids (AFs) or to the AF vial cultures, after a long incubation of 14 days; the results were compared with those of AF vial cultures. Escherichia coli was isolated in four of 32 AF vial cultures (12.5%). The application of 16S rRNA PCR directly to AF detected only one of the four positive samples (sensitivity 25%, specificity 100%, positive predictive value (PPV) 100%, negative predictive value (NPV) 90.32%). However, the application of 16S rRNA PCR to AF vial cultures after 14 days of incubation correctly identified all the positive samples, including one more that was positive for Brucella mellitensis (sensitivity 100%, specificity 80%, PPV 80%, NPV 100%). The elongation of the incubation period of the AF vial cultures, combined with the use of 16S rRNA in negative vials, increases the possibility of identifying the causative agents of SBP and could be applied in the clinical laboratory.

Clinical significance of organ- and non-organ-specific autoantibodies on the response to anti-viral treatment of patients with chronic hepatitis C.

BACKGROUND: Development of organ- and non-organ-specific autoantibodies has been reported in hepatitis C virus patients treated with interferon-alpha plus/minus ribavirin. AIMS: To address whether prevalence and the titre of gastric parietal autoantibodies and non-organ-specific autoantibody in hepatitis C virus-treated patients were affected by therapy, and if the development of these antibodies carries any clinical significance on the response to treatment, as few studies in adults have been strictly designed to address the above hypothesis. METHODS: Samples at three time-points (baseline, end of treatment, end of follow-up) from 102 hepatitis C virus patients (39 sustained responders, 26 relapsers, 33 non-responders; four lost in follow-up) were studied for gastric parietal autoantibodies and/or non-organ-specific autoantibody by indirect immunofluorescence, commercial and in-house enzyme-linked immunosorbent assays. RESULTS: Sustained virological and biochemical response was associated with antinuclear antibody absence (end of treatment or end of follow-up), decrease of smooth-muscle antibody titres during therapy and gastric parietal autoantibodies negativity at baseline. However, after multivariate analysis only antinuclear antibody positivity at the end of treatment and increase of smooth-muscle antibody titres were associated with worst treatment response, independently of known factors of worst treatment outcome. CONCLUSIONS: We were able to demonstrate a negative correlation between the efficacy of anti-viral treatment for hepatitis C virus and the presence of antinuclear antibody and smooth-muscle antibody before treatment, or their increase during therapy.

A real-world study focused on the long-term efficacy of mycophenolate mofetil as first-line treatment of autoimmune hepatitis.

BACKGROUND: Front-line therapy with mycophenolate mofetil (MMF) in autoimmune hepatitis (AIH) has shown high on-treatment remission rates. AIM: To study prospectively in a real-world fashion the long-term outcome of a large group of consecutive treatment-naïve AIH patients. METHODS: Between 2000 and 2014, 158 patients were recruited but only 131 were eligible for treatment (109 MMF/prednisolone; 22 prednisolone ± azathioprine). Long-term data on outcome after drug withdrawal were evaluated. Patients stopped treatment after having achieved complete response (normal transaminases and IgG) for at least the last 2 years. RESULTS: At diagnosis, 31.6% of patients had cirrhosis and 72.8% insidious presentation. A total of 102 of 109 (93.6%) responded initially to MMF within 2 (1-18) months. A total of 78 of 109 (71.6%) had complete response on treatment and 61 of 78 (78.2%) maintained remission off prednisolone. MMF-treated patients had increased probability of complete response compared to those receiving azathioprine (P = 0.03). Independent predictors of complete response were lower ALT at 6 months (P = 0.001) and acute presentation (P = 0.03). So far, treatment withdrawal was feasible in 40/109 patients and 30 (75%) are still in remission after 24 (2-129) months. Remission maintenance was associated with longer MMF treatment (P = 0.005), higher baseline ALT (P < 0.02), lower IgG on 6 months (P = 0.004) and histological improvement. CONCLUSIONS: Mycophenolate mofetil proved to be an efficient first-line treatment for AIH, achieving so far the highest rates of remission maintenance off treatment (75%) ever published for at least a median of 2 years, although the remission criteria used were strict. However, the risk of potential bias and overestimation of intervention benefits from MMF cannot be completely excluded as this is a real world and not a randomised controlled trial.

Markers of hepatitis viruses and human T-lymphotropic virus types I/II in patients who have undergone open-heart surgery: evidence of increased risk for exposure to HBV and HEV.

BACKGROUND: Open-heart procedure is characterized by a high-risk for contracting blood-borne infections. We evaluated the prevalence of several markers of hepatitis viruses (B-E) and human T-cell lymphotropic virus types I/II (HTLV-I/II) in a consecutive series of patients who had undergone open-heart surgery. METHODS: 204 patients and 158 selected age- and sex-matched healthy volunteers were investigated. Samples were collected at least 6-12 months postoperatively. Commercial enzyme immunoassays and confirmatory immunoblot assays for HCV, HEV and HTLV-I/II were used. RESULTS: None of the subjects tested positive for antibodies to HTLV-I/II. Prevalence of markers of past HBV infection and antibodies to HEV (anti-HEV) were higher in patients than in healthy controls (anti-HBc: 45.1% vs. 31%, p=0.009; anti-HBs: 31.9% vs. 22.2%, p=0.02; anti-HBe: 32.4% vs. 10.1%, p=0.000; anti-HEV: 5.4% vs. 0%, p=0.008). HBsAg and antibodies to HCV did not differ between the groups. CONCLUSIONS: HTLV, HBsAg and HCV infection markers did not differ between patients and healthy controls. However, patients had significantly increased prevalence of markers of previous HBV infection suggesting that an intensive vaccination schedule against HBV preoperatively might be helpful in minimizing the risk. The increased prevalence of anti-HEV in cardiac patients requires further investigation. Prospective studies are needed in order to definitely address whether the high prevalence of exposure to HBV and HEV infections in patients who had undergone open-heart surgery is procedure-related or not and whether it has any impact on morbidity of these patients.

Review article: autoimmune hepatitis -- current management and challenges.

BACKGROUND: Autoimmune hepatitis (AIH) is a disease of unknown aetiology characterised by interface hepatitis, hypergammaglobulinaemia, circulating autoantibodies and a favourable response to immunosuppression. AIM: To review recent advancements in understanding aetiopathogenesis, clinical, serological and histological features, diagnostic criteria and treatment strategies of AIH. METHODS: Published studies on AIH extracted mainly from PubMed during the last 15 years. RESULTS: Autoimmune hepatitis has a global distribution affecting any age, both sexes and all ethnic groups. Clinical manifestations are variable ranging from no symptoms to severe acute hepatitis and only seldom to fulminant hepatic failure. Autoimmune attack is perpetuated, possibly via molecular mimicry mechanisms, and favoured by the impaired control of regulatory T-cells. A typical laboratory finding is hypergammaglobulinaemia with selective elevation of IgG, although in 15-25% of patients - particularly children, elderly and acute cases - IgG levels are normal. Liver histology and autoantibodies, although not pathognomonic, still remain the hallmark for diagnosis. Immunosuppressive treatment is mandatory and life-saving; however, to meet strict response criteria, the conventional therapy with prednisolone with or without azathioprine is far from ideal. CONCLUSIONS: Autoimmune hepatitis remains a major diagnostic and therapeutic challenge. The clinician, the hepato-pathologist and the laboratory personnel need to become more familiar with different expressions of the disease, interpretation of liver histology and autoimmune serology. According to the strict definition of treatment response issued by the 2010 AASLD guidelines, many patients are nonresponders to conventional treatment. Newer immunosuppressive agents targeting pathogenetic mechanisms can improve patient management, which needs to be tailored on a case-by-case basis.

Anti-α actinin antibodies as new predictors of response to treatment in autoimmune hepatitis type 1.

BACKGROUND: We reported that combined presence of autoantibodies (Abs) against filamentous-actin (AFA) and α-actinin are specific for autoimmune hepatitis type 1 (AIH-1) diagnosis. AIM: To explore our data and assess whether anti-α-actinin and AFA Abs could be used as indicators of response to treatment and predictors of AIH-1 flares in a large cohort of AIH-1 patients. METHODS: Seven hundred and sixty-four serial serum samples of 86 consecutive AIH-1 patients, 509 pathological and 110 normal controls were tested for the presence of anti-α-actinin and AFA Abs by an in-house IgG-specific ELISA and a standardised commercially available ELISA respectively. Patients sera were divided into baseline group (active disease before treatment initiation, n = 86) and then according to treatment response into group A-responders (n = 40 patients), group B-relapsers/incomplete responders (n = 37 patients) and group C-not-treated (n = 9 patients). RESULTS: Anti-α-actinin and AFA levels were significantly higher at baseline. Double reactivity against α-actinin and AFA was associated with disease activity (OR 4.9; 95% CI: 2.7-9). Anti-α-actinin optical densities (ODs) before treatment decreased significantly at first remission (P < 0.05). Treatment response was associated with anti-α-actinin Abs negativity before treatment (OR 3.4; 95% CI: 1.3-8.9) and absence of double positivity for anti-α-actinin and AFA Abs before treatment (OR 3.8; 95% CI: 1.4-10.4). Responders had lower baseline levels of anti-α-actinin than relapsers and/or incomplete responders (P = 0.002). Binary logistic regression revealed lower levels of anti-α-actinin as the only independent predictors of response (P = 0.05). CONCLUSIONS: Anti-α-actinin Abs at baseline appear to predict treatment response and therefore they might be used for monitoring treatment outcome in AIH-1.

Real-time quantification of human telomerase reverse transcriptase mRNA in liver tissues from patients with hepatocellular cancer and chronic viral hepatitis.

We determined, for the first time, the human telomerase reverse transcriptase (hTERT) mRNA expression, using real-time quantitative PCR, in liver tissues from patients with hepatocellular cancer (HCC; n = 13), chronic hepatitis B (n = 19) and C (n = 13). Liver tissues from the 45 patients and 17 patients without liver disease in whom liver biopsy was performed during cholecystectomy (control group), were investigated for telomerase activity (TA) and hTERT mRNA expression using the LightCycler technology. TA was detected in all HCC tissues compared with 15.6% of chronic hepatitis (P < 0.001) and none of controls (P < 0.001). TA levels and hTERT mRNA were higher in HCC compared with chronic hepatitis (P < 0.001) and normal livers (P < 0.001). hTERT mRNA expression was correlated with TA (P < 0.05). Chronic hepatitis patients who tested negative for TA and hTERT mRNA had significantly lower disease duration (58 +/- 85 months) compared with those tested positive (144 +/- 50 months; P < 0.05). Detection of TA and quantification of hTERT mRNA expression in liver tissues could be useful and additional markers for HCC diagnosis and may serve as prognostic markers for HCC development in chronic viral hepatitis patients. However, we were not able to draw general conclusions at this moment, as the number of chronic hepatitis patients positive for hTERT mRNA was relatively small. Real-time quantification of hTERT mRNA expression as a diagnostic/prognostic marker in patients with chronic hepatitis B and C and its relationship with hepatocarcinogenesis needs further evaluation.

Direct detection of Cardiobacterium hominis in serum from a patient with infective endocarditis by broad-range bacterial PCR.

Bacterial DNA was detected directly in the serum of a patient with endocarditis by broad-range 16S rRNA PCR followed by sequencing and analysis of the results by the BLAST search. Using these methods, Cardiobacterium hominis was identified in 2 days from the date of serum collection. The microorganism was also isolated and identified using conventional methods (bacterial culture and biochemical tests) 17 days from the date of sample collection. This is the first report showing the direct detection of C. hominis in a patient's serum using molecular-based methods, emphasizing their potential usefulness as additional and rapid diagnostic tools for the detection and identification of fastidious bacteria.

Autoantibodies and defined target autoantigens in autoimmune hepatitis: an overview.

Autoimmune hepatitis (AIH) is a disease of unknown aetiology characterised by hypergammaglobulinaemia, non-organ and liver-related autoantibodies, association with HLA-DR3 or DR4 and a favourable response to immunosuppression. The current classification of AIH and the several autoantibodies/target autoantigens found in this disease are reported. The importance of these markers in the differential diagnosis and the study of pathogenesis of AIH is also given. AIH is subdivided into two major types: AIH type 1 (AIH-1) and AIH type 2 (AIH-2). AIH-1 is characterised by the detection of smooth muscle autoantibodies (SMA) and/or antinuclear antibodies (ANA). Antineutrophil cytoplasmic autoantibodies (ANCA), in most cases of perinuclear pattern (p-ANCA), by the indirect immunofluorescence assay, antibodies against the asialoglycoprotein receptor (anti-ASGP-R) and antibodies to soluble liver antigens or liver-pancreas (anti-SLA/LP) may be useful for the identification of individuals who are seronegative for ANA/SMA. AIH-2 is characterised by the presence of specific autoantibodies against liver and kidney microsomal antigens (anti-LKM type 1 or infrequently anti-LKM type 3) and/or autoantibodies against liver cytosol 1 antigen (anti-LC1). Anti-LKM-1 and anti-LKM-3 autoantibodies are also detected in some patients with chronic hepatitis C (HCV) and chronic hepatitis D (HDV). For these reasons, the distinction between AIH and chronic viral hepatitis is of particular importance. Cytochrome P450 2D6 (CYP2D6) is the major target autoantigen of anti-LKM-1 autoantibodies in both conditions (AIH-2 and HCV infection). Recent data have demonstrated the expression of CYP2D6 on the surface of hepatocytes, suggesting a pathogenetic role of anti-LKM-1 autoantibodies in liver injury. Family 1 of UDP-glycuronosyltransferases has been identified as the target autoantigen of anti-LKM-3. The molecular target of anti-SLA/LP autoantibodies has been identified recently as a 50 kDa protein with unknown structure and function. A liver-specific enzyme, the formiminotransferase cyclodeaminase, was identified as the target autoantigen of anti-LC1 autoantibodies. Anti-ASGP-R and anti-LC1 autoantibodies appear to correlate better with the severity of AIH and the response to treatment. The latter may suggest a pathogenic role of these autoantibodies in the hepatocellular damage in AIH. In general, however, autoantibodies should not be used to monitor treatment or to predict AIH activity or outcome. Finally, current knowledge concerning a specific form of AIH that may develop in some patients with a rare genetic syndrome, the autoimmune polyglandular syndrome type-1 (APS-1), is also discussed. Autoantibodies against liver microsomes (anti-LM) are the specific autoantibodies found in AIH as a disease component of APS-1. However, anti-LM autoantibodies have also been described in cases of dihydralazine-induced hepatitis. Cytochrome P450 1A2 has been identified as the target autoantigen of anti-LM autoantibodies in both disease entities.

Anti-cardiolipin antibodies in patients with chronic viral hepatitis are independent of beta2-glycoprotein I cofactor or features of antiphospholipid syndrome.

BACKGROUND: Although controversial, some authorities have implicated hepatitis C virus (HCV) as a cause of anti-phospholipid syndrome (APLS). Anti-cardiolipin antibodies (anti-CLAbs) in APLS are cofactor-dependent ('pathogenic' antibodies). We conducted a study in order to determine the prevalence of anti-CLAbs in HCV patients, and furthermore to address whether these autoantibodies are cofactor-dependent or not and whether they are associated with features of APLS. Patients with hepatitis B virus (HBV) were also evaluated in order to assess whether there are differences in the prevalence and the clinical significance of anti-CLAbs between these two major types of chronic viral hepatitis. MATERIALS AND METHODS: One hundred and seventy-four consecutive HCV patients, 50 HBV patients and 267 healthy were investigated for the presence of anti-CLAbs and antibodies against beta2-glycoprotein I (beta2-GPI), which is the most important cofactor of the 'pathogenic' anti-CLAbs in APLS. IgG anti-CLAbs were determined by an in-house quantitative ELISA and anti-beta2-GPIAbs using a commercial ELISA kit. RESULTS: 21.3% of the HCV and 14% of the HBV patients tested positive for IgG anti-CLAbs (P < 0.0001 compared with healthy controls). Neither age, sex, certain epidemiologic and laboratory parameters nor the clinical status and the histologic findings were associated with anti-CLAbs detection in both diseases. 2.3% of the HCV (P < 0.05 compared with healthy controls) and 2% of the HBV patients tested positive for anti-beta2-GPIAbs. Presence of anti-CLAbs was not associated with features of APLS. CONCLUSIONS: A significant proportion of the HCV and HBV patients had detectable IgG anti-CLAbs. However, the anti-CLAbs titres were relatively low, and in most cases seem to be cofactor-independent ('nonpathogenic'). The latter is further supported by the lack of their association with clinical features of APLS. Furthermore, anti-CLAbs appear to be detected irrespective of the demographic, laboratory, clinical and histologic status in both HCV and HBV. However, prospective studies of longer duration may be required in order to address whether anti-CLAbs in patients with chronic viral hepatitis are or are not of clinical importance.