Aberrant hippocampal transmission and behavior in mice with a stargazin mutation linked to intellectual disability.

Mutations linked to neurodevelopmental disorders, such as intellectual disability (ID), are frequently found in genes that encode for proteins of the excitatory synapse. Transmembrane AMPA receptor regulatory proteins (TARPs) are AMPA receptor auxiliary proteins that regulate crucial aspects of receptor function. Here, we investigate a mutant form of the TARP family member stargazin, described in an ID patient. Molecular dynamics analyses predicted that the ID-associated stargazin variant, V143L, weakens the overall interface of the AMPAR:stargazin complex and impairs the stability of the complex. Knock-in mice harboring the V143L stargazin mutation manifest cognitive and social deficits and hippocampal synaptic transmission defects, resembling phenotypes displayed by ID patients. In the hippocampus of stargazin V143L mice, CA1 neurons show impaired spine maturation, abnormal synaptic transmission and long-term potentiation specifically in basal dendrites, and synaptic ultrastructural alterations. These data suggest a causal role for mutated stargazin in the pathogenesis of ID and unveil a new role for stargazin in the development and function of hippocampal synapses.

Chronic warming stimulates growth of marsh grasses more than mangroves in a coastal wetland ecotone.

Increasing temperatures and a reduction in the frequency and severity of freezing events have been linked to species distribution shifts. Across the globe, mangrove ranges are expanding toward higher latitudes, likely due to diminishing frequency of freezing events associated with climate change. Continued warming will alter coastal wetland plant dynamics both above- and belowground, potentially altering plant capacity to keep up with sea level rise. We conducted an in situ warming experiment, in northeast Florida, to determine how increased temperature (+2°C) influences co-occurring mangrove and salt marsh plants. Warming was achieved using passive warming with three treatment levels (ambient, shade control, warmed). Avicennia germinans, the black mangrove, exhibited no differences in growth or height due to experimental warming, but displayed a warming-induced increase in leaf production (48%). Surprisingly, Distichlis spicata, the dominant salt marsh grass, increased in biomass (53% in 2013 and 70% in 2014), density (41%) and height (18%) with warming during summer months. Warming decreased plant root mass at depth and changed abundances of anaerobic bacterial taxa. Even while the poleward shift of mangroves is clearly controlled by the occurrences of severe freezes, chronic warming between these freeze events may slow the progression of mangrove dominance within ecotones.

CFTR biomarkers: time for promotion to surrogate end-point.

In patients with cystic fibrosis, cystic fibrosis transmembrane conductance regulator (CFTR) biomarkers, such as sweat chloride concentration and/or nasal potential difference, are used as end-points of efficacy in phase-III clinical trials with the disease modifying drugs ivacaftor (VX-770), VX809 and ataluren. The aim of this project was to review the literature on reliability, validity and responsiveness of nasal potential difference, sweat chloride and intestinal current measurement in patients with cystic fibrosis. Data on clinimetric properties were collected for each biomarker and reviewed by an international team of experts. Data on reliability, validity and responsiveness were tabulated. In addition, narrative answers to four key questions were discussed and agreed by the team of experts. The data collected demonstrated the reliability, validity and responsiveness of nasal potential difference. Fewer data were found on reliability of sweat chloride concentration; however, validity and responsiveness were demonstrated. Validity was demonstrated for intestinal current measurement, but further information is required on reliability and responsiveness. For all three end-points, normal values were collected and further research requirements were proposed. This body of work adds useful information to support the promotion of CFTR biomarkers to surrogate end-points and to guide further research in the area.

Diversity of Archaea in Brazilian savanna soils.

Although the richness of Bacteria and Fungi in Cerrado' soils has been reported, here we report, for the first time, the archaeal community in Cerrado's soils. DNA extracted from soil of two distinct vegetation types, a dense subtype of sensu strict (cerrado denso) and riverbank forest (mata de galeria), was used to amplify Archaea-specific 16S rRNA gene. All of the fragments sequenced were classified as Archaea into the phylum Thaumarchaeota, predominantly affiliated to groups I.1b and I.1c. Sequences affiliated to the group I.1a were found only in the soil from riverbank forest. Soils from 'cerrado denso' had greater Archaea richness than those from 'mata de galeria' based on the richness indexes and on the rarefaction curve. ß-Diversity analysis showed significant differences between the sequences from the two soil areas studied because of their different thaumarchaeal group composition. These results provide information about the third domain of life from Cerrado soils.

Opioid-free mastectomy under thoracic epidural anesthesia-analgesia in a patient with critical tracheal stenosis: a case report.

Mastectomy is traditionally performed under general anaesthesia and invasive ventilation, and is often complemented with regional techniques. In this setting, tracheal stenosis can pose a challenge to airway management. The aim of this report is to describe the successful management of a 68-year-old woman with severe subglottic tracheal stenosis undergoing mastectomy due to breast cancer. Surgery was performed without airway instrumentation under an opioid-free regimen consisting of thoracic epidural, propofol and dexmedetomidine perfusion, and non-opioid analgesics. Spontaneous ventilation and adequate perioperative analgesia were achieved. Opioid-free anaesthesia without airway instrumentation, consisting of thoracic epidural anaesthesia and sedation, is a good alternative in patients undergoing mastectomy in whom airway manipulation is best avoided.

Treatment of anaemia with erythropoiesis-stimulating agents in patients with chronic kidney disease does not lower mortality and may increase cardiovascular risk: a meta-analysis.

BACKGROUND/AIMS: Interpretation of the results of earlier meta-analyses in chronic kidney disease (CKD) patients on the impact of anaemia treatment with erythropoiesis-stimulating agents (ESAs) on clinical outcomes has been hampered by the inclusion of small trials and trials of short duration. We re-evaluated the benefits and harms of treating anaemia, including only relevant clinical trials. METHODS: We conducted a systematic review and meta-analysis of randomised controlled trials performed in adults with CKD which allocated patients to different doses of ESAs, and we compared the effect of these interventions on vascular access thrombosis, stroke, risk of end-stage renal disease (ESRD) and all-cause mortality. Additional inclusion criteria were studies with a duration of at least 1 year and enrolling more than 500 participants. RESULTS: Five trials (7,902 participants) met the inclusion criteria and were included in the meta-analysis. The number of patients enrolled in each trial ranged from 596 to 4,038. The mean/median duration of follow-up ranged from 14 to 36 months. A higher haemoglobin target was associated with increased risk of vascular access thrombosis (RR 1.343; 95% CI 1.162-1.554; p = 0.0005) and stroke (RR 1.735; 95% CI 1.323-2.275; p = 0.0005), and no effect on risk of ESRD (RR 1.089; 95% CI 0.986-1.203; p = 0.094) or all-cause mortality (RR 1.148; 95% CI 0.977-1.350; p = 0.093). CONCLUSION: In CKD patients, treatment of anaemia with ESAs targeting a higher haemoglobin value does not lower mortality or reduce the risk of ESRD, and may increase cardiovascular risk.

Use of a mobile application for self-management of pancreatic enzyme replacement therapy is associated with improved gastro-intestinal related quality of life in children with Cystic Fibrosis.

BACKGROUND: Most patients with cystic fibrosis (CF) suffer from pancreatic insufficiency (PI), leading to fat malabsorption, malnutrition, abdominal discomfort and impaired growth. Pancreatic enzyme replacement therapy (PERT) is effective, but evidence based guidelines for dose adjustment are lacking. A mobile app for self-management of PERT was developed in the context of the HORIZON 2020 project MyCyFAPP. It contains an algorithm to calculate individual PERT-doses for optimal fat digestion, based on in vitro and in vivo studies carried out in the same project. In addition, the app includes a symptoms diary, educational material, and it is linked to a web tool allowing health care professionals to evaluate patient's data and provide feedback. METHODS: A 6-month open label prospective multicenter interventional clinical trial was performed to assess effects of using the app on gastro-intestinal related quality of life (GI QOL), measured by the CF-PedsQL-GI (shortened, CF specific version of the Pediatric Quality of Life Inventory, Gastrointestinal Symptoms Module). RESULTS: One hundred and seventy-one patients with CF and PI between 2 and 18 years were recruited at 6 European CF centers. Self-reported CF-PedsQL-GI improved significantly from month 0 (M0) (84.3, 76.4-90.3) to month 6 (M6) (89.4, 80.35-93.5) (p< 0.0001). Similar improvements were reported by parents. Lower baseline CF-PedsQL-GI was associated with a greater improvement at M6 (p < 0.001). CONCLUSIONS: The results suggest that the MyCyFAPP may improve GI QOL for children with CF. This tool may help patients to improve self-management of PERT, especially those with considerable GI symptoms.

99mTc-sestamibi SPECT/CT and 18F-FDG-PET/CT have similar performance but different imaging patterns in newly diagnosed multiple myeloma.

PURPOSE: F-fluorodeoxiglucose (F-FDG)-PET/CT has been widely used to evaluate multiple myeloma. Tc-sestamibi (MIBI) scintigraphy has also been proposed for assessing multiple myeloma, but its use with state-of-the-art single-photon emission computed tomography/computed tomography (SPECT/CT) technology has not been fully evaluated.This study aimed to compare these two imaging modalities in multiple myeloma staging. MATERIALS AND METHODS: Sixty-two patients with recently diagnosed multiple myeloma were submitted to whole-body F-FDG-PET/CT and whole-body MIBI scans plus SPECT/CT of the chest and abdomen/pelvis. Number of focal lesions, contiguous soft tissue involvement (CSTI), extramedullary lesions (EMLs) and diffuse bone marrow (BM) involvement were recorded. RESULTS: PET/CT was positive in 59 patients (95%) and MIBI SPECT/CT in 58 (93%) (P = 0.69). MIBI detected more diffuse bone marrow involvement than PET/CT (respectively 78 vs. 58% of the patients), while PET/CT demonstrated more focal lesions than MIBI SPECT/CT (81 vs. 54% of the patients) (P = 0.002). PET/CT detected EMLs in four subjects and MIBI in one subject. CSTI was found in 28 (45%) and 23 (37%) patients on PET/CT and MIBI images, respectively (P = 0.36). Three patients with lytic lesions and no FDG uptake were MIBI positive, and two subjects with lytic lesions without MIBI uptake were FDG positive. CONCLUSION: MIBI SPECT/CT performs similarly to F-FDG-PET/CT in identifying sites of active disease in multiple myeloma staging. MIBI is more efficient than FDG for detecting the diffuse involvement of bone marrow but less efficient for detecting focal lesions. Some patients presented a 'mismatch' pattern of FDG/MIBI uptake.

Evidence of the growth plate and the growth of long bones in juvenile dinosaurs.

Histological and ultrastructural evaluation of the ends of long bones of juvenile dinosaurs from the Upper Cretaceous Two Medicine Formation of Montana revealed the preservation of growth plates. Growth plates are discs of cartilage present near the ends of growing long bones that generate bone elongation. Comparison of the fossils with modern taxa demonstrated homology of the growth plate in birds and dinosaurs. The presence of an avian-type growth plate in dinosaurs adds a shared derived anatomical character corroborating inclusion of birds within the Dinosauria. Additionally, possession of a growth plate, which in birds is capable of producing rapid determinate long bone growth, implies that an avian developmental pattern may have been present in these dinosaurs.

Galectin and aldolase-like molecules are responsible for the specific IgE response in humans exposed to Dirofilaria immitis.

Dirofilaria immitis is the agent of the heartworm disease in canids and felids, and of pulmonary dirofilariosis in man. Like other filariae, D. immitis harbours endosymbion Wolbachia bacteriae. In this work we analyse the response of specific IgE antibodies against both D. immitis antigens and the Wolbachia surface protein (WSP) in two groups of persons living in an area of canine endemia, one presenting high levels of total IgE (group 1) and other with normal levels (group 2). Infections with D. immitis were demonstrated by the presence of specific IgG in 228 individuals(48.8%) of the group 1 and only in one of the group 2. Specific IgE antibody response against D. immitis antigens was detected only in individuals of the group 1. IgE response against WSP was not detected in any group. The IgE response was directed mainly against two molecules of 33 and 42 kDa of the antigenic extract of D. immitis. These molecules were identified by mass spectrometry as a galectin and an aldolase, respectively. Their possible role in the survival mechanisms of the parasite and their contribution to development of allergic reactions in individuals resident in areas with heartworm disease are discussed.

Short and medium-term effects of two-phase olive mill waste application on olive grove production and soil properties under semiarid mediterranean conditions.

A five-year field study was conducted to evaluate the potential use of oiled and de-oiled two-phase olive mill waste (TPOMW and DTPOMW, respectively) as soil amendment on a representative olive grove soil: a Cutanic Luvisol. Treatments included a non-amended control, TPOMW1, TPOMW2, DTPOMW1 and DTPOMW2 (30, 60, 27 and 54Mgha(-1) dry weight equivalent). Significant increases (P<0.05) in organic carbon, total N, available P and K, and aggregate stability were observed in the amended soils. Leaf analysis showed significant increases in N, P, and K concentrations in treated plots after the two first years of TPOMW or DTPOMW amendments. Also, a general increase in olive production was observed in the treated plots, this increase being higher in the TPOMW1 and DTPOMW1 treated soils. After five years of repeated TPOMW and DTPOMW application, the increase in yield was 29%, 9.8%, 30%, and 19% for TPOMW1, TPOMW2, DTPOMW1, and DTPOMW2, respectively. Raw TPOMW and DTPOMW have the potential to be valuable soil amendments and source of organic matter, with a positive effect on olive yield, and closing the cycle of residues-resources.

Microbiota modification by probiotic supplementation reduces colitis associated colon cancer in mice.

AIM: To investigate the effect of probiotic supplementation during the development of an experimental model of colitis associated colon cancer (CAC). METHODS: C57BL/6 mice received an intraperitoneal injection of azoxymethane (10 mg/kg), followed by three cycles of sodium dextran sulphate diluted in water (5% w/v). Probiotic group received daily a mixture of Lactobacillus acidophilus, Lactobacillus rhamnosus and Bifidobacterium bifidum. Microbiota composition was assessed by 16S rRNA Illumina HiSeq sequencing. Colon samples were collected for histological analysis. Tumor cytokines was assessed by Real Time-PCR (Polymerase Chain Reaction); and serum cytokines by Multiplex assay. All tests were two-sided. The level of significance was set at P < 0.05. Graphs were generated and statistical analysis performed using the software GraphPad Prism 5.0. The project was approved by the institutional review board committee. RESULTS: At day 60 after azoxymethane injection, the mean number of tumours in the probiotic group was 40% lower than that in the control group, and the probiotic group exhibited tumours of smaller size (< 2 mm) (P < 0.05). There was no difference in richness and diversity between groups. However, there was a significant difference in beta diversity in the multidimensional scaling analysis. The abundance of the genera Lactobacillus, Bifidobacterium, Allobaculum, Clostridium XI and Clostridium XVIII increased in the probiotic group (P < 0.05). The microbial change was accompanied by reduced colitis, demonstrated by a 46% reduction in the colon inflammatory index; reduced expression of the serum chemokines RANTES and Eotaxin; decreased p-IKK and TNF-α and increased IL-10 expression in the colon. CONCLUSION: Our results suggest a potential chemopreventive effect of probiotic on CAC. Probiotic supplementation changes microbiota structure and regulates the inflammatory response, reducing colitis and preventing CAC.

Characteristics of seminal plasma proteins and their correlation with canine semen analysis.

The objectives were to separate canine seminal plasma proteins (with SDS-PAGE) and to determine the correlation between specific proteins and semen characteristics. Three ejaculates from 20 mixed-breed dogs, of unknown fertility, were collected by digital manipulation. Ejaculate volume and color, sperm motility, sperm vigor, percentage of morphologically normal spermatozoa, and membrane integrity (hypoosmotic swelling test and fluorescent staining) were assessed. For each dog, seminal plasma was pooled from all three ejaculates and proteins were separated with SDS-PAGE, using polyacrylamide concentrations of 13% and 22% in the separation gels. After staining, gel images were digitized to estimate molecular weights (MW) and integrated optical density (IOD) of each lane and of individual bands. Total seminal plasma protein concentration was 2.19+/-1.56 g/dL (mean+/-SD; range 1.12-5.19 g/dL). A total of 37 protein bands were identified (although no dog had all 37 bands). In the 13% gel, molecular weights ranged from 100.6 to 17.1 kDa, with four bands (49.7, 33.2, 26.4, and 19.5 kDa) present in samples from all dogs. In the 22% gel, molecular weights ranged from 15.6 to 3.6 kDa, with nine bands (15.6, 13.5, 12.7, 11.7, 10.5, 8.7, 7.8, 5.6, and 4.9 kDa) present in samples from all dogs. Combined for both gels, the majority of bands (85%) had molecular weights <17 kDa, with B20 (15.6 kDa) in high concentrations in samples from all dogs. There were positive correlations (P < or = 0.01) between two bands, B4 (67 kDa) and B5 (58.6 kDa), and sperm motility (r=0.66 and r=0.46), sperm vigor (r=0.56 and r=0.66), percentage of morphologically normal spermatozoa (r=0.55 and r=0.59), the hypoosmotic swelling test (r=0.76 and r=0.68), and fluorescent staining (r=0.56 and r=0.59), respectively. In conclusion, 37 proteins were identified in seminal plasma; two were significantly correlated with semen characteristics.

[Insulin and angiotensin II signaling pathways cross-talk: implications with the association between diabetes mellitus, arterial hypertension and cardiovascular disease]. / Cross-talk das vias de sinalização de insulina e angiotensina II: implicações com a associação entre diabetes mellitus e hipertensão arterial e doença cardiovascular.

Insulin (Ins) and angiotensin II (AII) play pivotal roles in the control of two vital and closely related systems: the metabolic and the circulatory, respectively. A failure in the proper action of each of these hormones results, to a variable degree, in the development of two highly prevalent and commonly overlapping diseases--diabetes mellitus (DM) and hypertension (AH). In recent years, a series of studies has revealed a tight connection between the signal transduction pathways that mediate Ins and AII actions in target tissues. This molecular cross-talk occurs at multiple levels and plays an important role in phenomena that range from the action of anti-hypertensive drugs to cardiac hypertrophy and energy acquisition by the heart. At the extracellular level, the angiotensin-converting enzyme controls AII synthesis but also interferes with Ins signaling through the proper regulation of AII and the accumulation of bradykinin. At an early intracellular level, AII, acting through JAK-2/IRS-1/PI3-kinase, JNK and ERK, may induce the serine phosphorylation and inhibition of key elements of the Ins-signaling pathway. Finally, by inducing the expression of the regulatory protein SOCS-3, AII may impose a late control on the Ins signal. This review will focus on the main advances obtained in this field and will discuss the implications of this molecular cross-talk in the common clinical association between DM and AH.

Isolation and partial characterization of starch from banana cultivars grown in Colombia.

Banana starch is resistant to hydrolysis by digestive enzymes due to its structure and dietary fibre content. Starch was isolated from the following three cultivars of Colombian Musaceae: Gros Michel (dessert), Dominico Harton and FHIA 20 (cooking); also, the amylose and amylopectin contents, morphology of the granules, thermal properties, pasting, molecular characteristics and digestibility were determined. The total starch content, amylose content and digestibility (gelatinized starch) were higher in cooking varieties; the purity and gelatinization temperature were similar for the three varieties, but the enthalpy was higher in the dessert variety. The three varieties showed higher viscosities in the pasting profile compared to commercial maize starch in both acid and neutral conditions. Starch granules presented with heterogeneous sizes and shapes (elongated and ovals) that had birefringence. The Dominico Hartón variety showed the lowest rapidly digestible starch (RDS) value in the gelatinized sample that is in agreement with the greater proportion of long chains.

Respiratory sleep disturbance in children and adolescents with cystic fibrosis.

UNLABELLED: Sleep disturbance has been described in cystic fibrosis (CF) patients as relevant to clinical and lung function predictive factors helping to improve the diagnosis and early intervention. Related paediatric studies are scarce. OBJECTIVE: To describe respiratory sleep disturbance (RSD) and its association with spirometric indices in a population of CF children. A second aim was to determine if spirometric indices and wake-time SpO2 are predictors of sleep disturbance. METHODS: A cross-sectional study involving 33CF paediatric patients. All participants underwent in-lab polysomnography (PSG), pulse oximetry and spirometry. A standardized sleep questionnaire was completed for each patient. Two subgroups were considered: I - Normal (FEV1>-1.64 z-score); II - Obstructed (FEV1≤-1.64 z-score). RESULTS: Participant's median age was 12 (6-18) years, 16 (48.5%) were male. Twenty-nine patients (87.9%) presented sleep complaints. Sleep efficiency was reduced; sleep latency and waking after sleep onset (WASO) increased. N1 increased, N2, N3, REM and awakenings were normal. The apnoea-hypopnoea index was 0.6/h (sd 0.9); respiratory disturbance index (RDI) was 6.6/h (sd 5.2). Mean awaking (97% (sd 1.1)) and sleep SpO2 (95% (sd 2.7)) were normal; mean nocturnal oximetry desaturation index was 2.36/h; minimal nocturnal SpO2 was 89% (sd 4.1). We found associations between mean nocturnal SPO2 and mean values of FEV1 (r=0.528; p=0.002) and FEF25-75 (r=0.426; p=0.013). There were significant differences in nocturnal SpO2 between normal and obstructed patients (p<0.000). PSG data correlated with the questionnaire answers for night awakenings and WASO (p=0.985) and difficult breathing during sleep and RDI (p=0.722). This study points to most CF children having sleep complaints, and highlights the correlation between subjective assessment of sleep and PSG and spirometric results. Awake-time SpO2 and spirometric values are possible risk predictors for nocturnal desaturation.

BCR-ABL binds to IRS-1 and IRS-1 phosphorylation is inhibited by imatinib in K562 cells.

In the present study we used K562 cells to demonstrate that insulin receptor substrate 1 (IRS-1) is expressed and constitutively phosphorylated in BCR-ABL(+) cells. We observed association between BCR-ABL/IRS-1, IRS-1/phosphoinositide 3'-kinase (PI3-kinase), and IRS-1/Grb2 in the K562 cell line. Our findings demonstrate that imatinib treatment resulted in marked attenuation of BCR-ABL/IRS-1 association and of IRS-1-stimulated PI3-kinase activity in K562 cells. We concluded that the IRS-1 protein is involved in the signalling pathway of the BCR-ABL tyrosine kinase.

High frequency gene transfer by microprojectile bombardment of intact conidia from the entomopathogenic fungus Paecilomyces fumosoroseus.

Two different methods, (i) PEG and (ii) biolistic, were employed to transform protoplasts and conidia of Paecilomyces fumosoroseus using hygromycin resistance as selectable marker. Transformation frequencies varied from 1.9 to 2.5 transformants microgram-1 of DNA by the PEG method, and from 33 to 153 transformants microgram-1 of DNA by the biolistic procedure.

Differential growth by growth plates as a function of multiple parameters of chondrocytic kinetics.

Differential elongation of growth plates is the process by which growth-plate chondrocytes translate the same sequence of gene regulation into the appropriate timing pattern for a given rate of elongation. While some of the parameters associated with differential growth are known, the purpose of this study was to test the hypothesis that eight independent variables are involved. We tested this hypothesis by considering four different growth plates in 28-day-old Long-Evans rats. Temporal parameters were provided by means of oxytetracycline and bromodeoxyuridine labeling techniques. Stereological parameters were measured with standard techniques. For all four growth plates, the calculated number of new chondrocytes produced per day approximated the number of chondrocytes lost per day at the chondro-osseous junction. This suggests that the proposed equations and associated variables represent a comprehensive set of variables defining differential growth. In absolute numbers, the proximal tibial growth plate produced about four times as many chondrocytes per day as the proximal radial growth plate (16,400 compared with 3,700). In the proximal tibia, 9% of growth is contributed by cellular division; 32%, by matrix synthesis throughout the growth plate; and 59%, by chondrocytic enlargement during hypertrophy. In the more slowly elongating growth plates, the relative contribution to elongation from cellular enlargement decreases from 59 to 44%, with a relative increase in contribution from matrix synthesis ranging from 32% in the proximal tibia 49% in the proximal radius. This study suggests that differential growth is best depicted as a complex interplay among cellular division, matrix synthesis, and cellular enlargement during hypertrophy. Differential growth is best explained by considering a set of eight independent variables, seven of which vary from growth plate to growth plate. Thus, this study confirms the importance of cellular hypertrophy during elongation and adds to our understanding of the importance of locally mediated regulatory systems controlling growth-plate activity.

Spontaneous hepatitis B surface antigen clearance in a long-term follow-up study of patients with chronic type B hepatitis. Lack of correlation with hepatitis C and D virus superinfection.

We investigated the frequency of HBsAg clearance and the possible role of viral superinfection in a long-term follow-up of 184 patients with chronic hepatitis B (CHB). Our subjects were 184 patients with chronic hepatitis B and the follow-up was 12-216 months (mean 66.2 +/- 53.7 months). The investigative methods used were: immunoenzymatic assays for HBV, HCV, HDV, and HIV markers; polymerase chain reaction (PCR) for HBV DNA; and liver biopsy and immunoperoxidase. During the follow-up, 20 of the 184 patients cleared serum HBsAg. A comparison of patients with persistent HBsAg(group I) and of those who cleared this marker (group II) showed a significant difference in mortality (P = 0.002) between the two groups and a tendency to a more severe exacerbation (flare) in group II (P = 0.07). Antibodies to hepatitis C and D virus as well as antibodies to HIV were equally distributed in both groups. Thirteen patients (7.9%) from group I, but none from group II, subsequently developed hepatocellular carcinoma. These results suggest that the frequency of spontaneous clearance of HBsAg during chronic HBV infection is low. No determinant factor for the clearance was found, including the presence of liver cirrhosis. Serum HBV DNA was undetectable by PCR after clearance in 16 out of 17 patients.