Limited hepatitis B virus replication space in the chronically hepatitis C virus-infected liver.

We compared the kinetics and magnitude of hepatitis B virus (HBV) infection in hepatitis C virus (HCV)-naive and chronically HCV-infected chimpanzees in whose livers type I interferon-stimulated gene (ISG) expression is strongly induced. HBV infection was delayed and attenuated in the HCV-infected animals, and the number of HBV-infected hepatocytes was drastically reduced. These results suggest that establishment of HBV infection and its replication space is limited by the antiviral effects of type I interferon in the chronically HCV-infected liver.

Adaptation of a genotype 3 hepatitis E virus to efficient growth in cell culture depends on an inserted human gene segment acquired by recombination.

An infectious cDNA clone of a genotype 3 strain of hepatitis E virus adapted to growth in HepG2/C3A human hepatoma cells was constructed. This virus was unusual in that the hypervariable region of the adapted virus contained a 171-nucleotide insertion that encoded 58 amino acids of human S17 ribosomal protein. Analyses of virus from six serial passages indicated that genomes with this insert, although initially rare, were selected during the first passage, suggesting it conferred a significant growth advantage. RNA transcripts from this cDNA and the viruses encoded by them were infectious for cells of both human and swine origin, the major host species for this zoonotic virus. Mutagenesis studies demonstrated that the S17 insert was a major factor in cell culture adaptation. Introduction of 54 synonymous mutations into the insert had no detectable effect, thus implicating protein, rather than RNA, as the important component. Truncation of the insert by 50% decreased the levels of successful transfection by ~3-fold. Substitution of the S17 sequence by a different ribosomal protein sequence or by GTPase-activating protein sequence resulted in a partial enhancement of transfection levels, whereas substitution with 58 amino acids of green fluorescent protein had no effect. Therefore, both the sequence length and the amino acid composition of the insert were important. The S17 sequence did not affect transfection of human hepatoma cells when inserted into the hypervariable region of a genotype 1 strain, but this chimeric genome acquired a dramatic ability to replicate in hamster cells.

A naturally occurring human/hepatitis E recombinant virus predominates in serum but not in faeces of a chronic hepatitis E patient and has a growth advantage in cell culture.

Hepatitis E virus is the aetiological agent of acute hepatitis E, a self-limiting disease prevalent in developing countries. Molecular analysis of viral genomic RNA from a chronically infected patient confirmed the recent discovery that chronic infection correlated with extensive diversification of the virus quasispecies: the hypervariable region of some virus genomes in this USA patient contained large continuous deletions and a minor proportion of genomes in faeces and serum had acquired a mammalian sequence that encoded 39 aa of S19 ribosomal protein fused to the virus non-structural protein. Genomes with this insert were selected during virus passage in cultured cells to become the predominant species, suggesting that the inserted sequence promoted virus growth. The results demonstrated that hepatitis E virus can mutate dramatically during a prolonged infection and suggests it may be important to prevent or cure chronic infections before new variants with unpredictable properties arise.

Lack of protective immunity against reinfection with hepatitis C virus.

Some individuals infected with hepatitis C virus (HCV) experience multiple episodes of acute hepatitis. It is unclear whether these episodes are due to reinfection with HCV or to reactivation of the original virus infection. Markers of viral replication and host immunity were studied in five chimpanzees sequentially inoculated over a period of 3 years with different HCV strains of proven infectivity. Each rechallenge of a convalescent chimpanzee with the same or a different HCV strain resulted in the reappearance of viremia, which was due to infection with the subsequent challenge virus. The evidence indicates that HCV infection does not elicit protective immunity against reinfection with homologous or heterologous strains, which raises concerns for the development of effective vaccines against HCV.

Characterization of hepatitis E-specific cell-mediated immune response using IFN-gamma ELISPOT assay.

In developing countries, hepatitis E (HEV) and hepatitis A (HAV) are the major causes of acute viral hepatitis with similar feco-oral modes of transmission. In contrast to the high seroprevalence of hepatitis A infection, a low seroprevalence of HEV among children in endemic areas has been reported. These data suggest the possibility that silent HEV infection is undiagnosed by the current available methods. Many of the serological tests used for HEV diagnosis have poor specificity and are unable to differentiate among different genotypes of HEV. Moreover, the RT-PCR used for HEV isolation is only valid for a brief period during the acute stage of infection. Cell-mediated immune (CMI) responses are highly sensitive, and long lasting after sub-clinical infections as shown in HCV and HIV. Our objective was to develop a quantitative assay for cell-mediated immune (CMI) responses in HEV infection as a surrogate marker for HEV exposure in silent infection. Quantitative assessment of the CMI responses in HEV will also help us to evaluate the role of CMI in HEV morbidity. In this study, an HEV-specific interferon-gamma (IFN-gamma) ELISPOT assay was optimized to analyze HEV-specific CMI responses. We used peripheral blood mononuclear cells (PBMC) and sera from experimentally infected chimpanzees and from seroconverted and control human subjects to validate the assay. The HEV-specific IFN-gamma ELISPOT responses correlated strongly and significantly with anti-HEV ELISA positive/negative results (rho=0.73, p=0.02). Moreover, fine specificities of HEV-specific T cell responses could be identified using overlapping HEV ORF2 peptides.

Detection of SIV antigens by HIV-1 antigen capture immunoassays.

We tested three commercially available HIV-1 antigen capture systems for their ability to detect simian immunodeficiency viruses (SIV). All three kits detected antigens from six distinct SIV isolates, but with varying degrees of sensitivity. For the overall detection of SIV in cell culture, our assay for reverse transcriptase was more sensitive than HIV-1 antigen capture. HIV-1 antigen capture systems were useful for the detection of SIV antigenemia in experimentally infected macaques. The limited sensitivity of HIV-1-specific antigen capture suggests that SIV-specific antigen capture reagents should be developed.

Radioimmunoassay and characterization of woodchuck hepatitis virus core antigen and antibody.

Solid-phase radioimmunoassays for woodchuck hepatitis virus core antigen (WHcAg) and antibody (anti-WHc) were developed. WHcAg in woodchuck liver homogenates was characterized by ultracentrifugation in CsCl gradients; both heavy (1.35 g/cm3) and light (1.31 g/cm3) cores were obtained from the liver of an animal during acute WHV infection, which is consistent with observations in hepatitis B virus infection in man. Endpoint titers of anti-WHc were higher in chronic WHV carriers than in animals recovered from acute infections. Both IgM and IgG anti-WHc antibodies were produced by infected woodchucks. A survey of colony woodchucks demonstrated that 88/89 animals having one or more markers of past or ongoing WHV infection were positive for anti-WHc. Thus, serum anti-WHc appears to be a sensitive marker of WHV infection.

Working memory and retrieval: a resource-dependent inhibition model.

Four experiments examined individual differences in working memory (WM) capacity and how those differences affect performance on retrieval from both primary and secondary memory. The results showed that WM differences appear only in retrieval from primary memory and then only under conditions that lead to interference or response competition within the task. This suggests that WM capacity is important to retrieval that is based on controlled effortful search but not search that is based on automatic activation. A view is presented suggesting that individual differences in attentional resources lead to differences in the ability to inhibit or suppress irrelevant information. The paradigm also allowed more general comparisons between the processes involved in retrieval from primary and secondary memory. As expected, it was found that retrieval from primary memory was a function of set size. However, for sets larger than 2 items, retrieval from secondary memory was independent of set size.

The role of working memory capacity in retrieval.

The verbal fluency task requires generation of category exemplars and appears to be an example of what M. Moscovitch (1995) calls a strategic test of memory retrieval. Four experiments explored the role of individual differences in working memory (WM) capacity on verbal fluency under various secondary load conditions. High WM participants consistently recalled more exemplars. However, load conditions caused a decline in recall only for high WM participants. Low WM participants showed no effect of secondary workload on exemplar generation. WM group differences and load effects were observed even in the 1st min of retrieval, which suggests that differences were not due to differences in knowledge. A model of retrieval is supported that relies on cue-based-automatic activation, monitoring of output for errors, controlled suppression of previously recalled items, and controlled strategic search.

A controlled-attention view of working-memory capacity.

In 2 experiments the authors examined whether individual differences in working-memory (WM) capacity are related to attentional control. Experiment 1 tested high- and low-WM-span (high-span and low-span) participants in a prosaccade task, in which a visual cue appeared in the same location as a subsequent to-be-identified target letter, and in an antisaccade task, in which a target appeared opposite the cued location. Span groups identified targets equally well in the prosaccade task, reflecting equivalence in automatic orienting. However, low-span participants were slower and less accurate than high-span participants in the antisaccade task, reflecting differences in attentional control. Experiment 2 measured eye movements across a long antisaccade session. Low-span participants made slower and more erroneous saccades than did high-span participants. In both experiments, low-span participants performed poorly when task switching from antisaccade to prosaccade blocks. The findings support a controlled-attention view of WM capacity.

Dissociable brain mechanisms for inhibitory control: effects of interference content and working memory capacity.

In this study, event-related fMRI was used to examine whether the resolution of interference arising from two different information contents activates the same or different neuronal circuitries. In addition, we examined the extent to which these inhibitory control mechanisms are modulated by individual differences in working memory capacity. Two groups of participants with high and low working memory capacity [high span (HS) and low span (LS) participants, respectively] performed two versions of an item recognition task with familiar letters and abstract objects as stimulus materials. Interference costs were examined by means of the recent negative probe technique with otherwise identical testing conditions across both tasks. While the behavioral interference costs were of similar magnitude in both tasks, the underlying brain activation pattern differed between tasks: The object task interference-effects (higher activation in interference trials than in control trials) were restricted to the anterior intraparietal sulcus (IPS). Interference effects for familiar letters were obtained in the anterior IPS, the left postero-ventral and the right dorsolateral prefrontal cortex (PFC) as well as the precuneus. As the letters were more discernible than the objects, the results suggest that the critical feature for PFC and precuneus involvement in interference resolution is the saliency of stimulus-response mappings. The interference effects in the letter task were modulated by working memory capacity: LS participants showed enhanced activation for interference trials only, whereas for HS participants, who showed better performance and also lower interference costs in the letter task, the above-mentioned neuronal circuitry was activated for interference and control trials, thereby attenuating the interference effects. The latter results support the view that HS individuals allocate more attentional resources for the maintenance of task goals in the face of interfering information from preceding trials with familiar stimulus materials.

Epidemiology of acute hepatitis in the Stann Creek District of Belize, Central America.

Hepatitis is common in the Stann Creek District of southern Belize. To determine the etiologies, incidence, and potential risk factors for acute jaundice, we conducted active surveillance for cases. Cases of jaundice diagnosed by a physician within the previous 6 weeks were enrolled. Evaluation included a questionnaire and laboratory tests for hepatitis A, B, C, D, and E, a blood film for malaria, and a serologic test for syphilis. Etiologies of jaundice among 62 evaluable patients included acute hepatitis A, 6 (9.7%), acute hepatitis B, 49 (79.0%), hepatitis non-A-E, 2 (3.2%), and malaria, 5 (8.1%). There were no cases of acute hepatitis E. One patient each with antibody to hepatitis C and D were detected. The annualized incidence of hepatitis A was 0.26 per 1,000. All cases of hepatitis A were in children 4-16 years of age. The annualized incidence of hepatitis B, 2.17 per 1,000, was highest in adults aged 15-44 years (4.4 per 1,000) and was higher in men (36 cases; 3.09 per 1,000) than women (13 cases; 1.19 per 1,000). Four (31%) of the women with hepatitis B were pregnant. The annualized incidence was significantly higher in Mestizo (6.18 per 1000) and Maya (6.79 per 1,000) than Garifuna (0.38 per 1,000) or Creole (0.36 per 1,000). Persons with hepatitis B were significantly more likely to be born outside of Belize (82%), had been in Belize < 5 years (73%), and lived and worked in rural areas (96%) than was the general population. Of those > or = 14 years of age with hepatitis B, only 36% were married. Few persons admitted to transfusions, tattoos, IV drug use, multiple sexual partners, visiting prostitutes, or sexually transmitted diseases. Only 1 of 49 had a reactive test for syphilis. Six patients were hospitalized (including 3 with acute hepatitis B and one with hepatitis A), and none to our knowledge died. Acute hepatitis B is the most common cause of viral hepatitis in the Stann Creek District, but the modes of transmission remain obscure. Infants, women attending prenatal clinics, and new workers are potential targets for immunization with hepatitis B vaccine.

An outbreak of fulminant hepatitis delta in the Waorani, an indigenous people of the Amazon basin of Ecuador.

An outbreak of delta hepatitis occurred during 1998 among the Waorani of the Amazon basin of Ecuador. Among 58 people identified with jaundice, 79% lived in four of 22 Waorani communities. Serum hepatitis B surface antigen (HBsAg) was found in the sera of 54% of the jaundiced persons, and 14% of asymptomatic persons. Ninety-five percent of 105 asymptomatic Waorani had hepatitis B core (HBc) IgG antibody, versus 98% of 51 with jaundice. These data confirm that hepatitis B virus (HBV) infection is highly endemic among the Waorani. Sixteen of 23 (70%) HBsAg carriers identified at the onset of the epidemic had serologic markers for hepatitis D virus (HDV) infection. All 16 were jaundiced, where as only two of seven (29%) with negative HDV serology were jaundiced (P = .0006). The delta cases clustered in families, 69% were children and most involved superinfection of people chronically infected with HBV. The data suggest that HDV spread rapidly by a horizontal mode of transmission other than by the sexual route.

Echoic memory processes in good and poor readers.

Four experiments were conducted to determine whether echoic memory plays a role in differences between good and poor readers. The first two experiments used a suffix procedure in which the subject is read a list of digits with either a tone control or the word go appended to the list. For lists that exceeded the length of the subjects memory span by one digit (i.e., that avoided ceiling effects), the poor readers showed a larger decrement in the suffix condition than did the good readers. The third experiment was directed at the question of whether the duration of echoic memory is different for good and poor readers. Children shadowed words presented to one ear at a rate determined to give 75-85% shadowing accuracy. The items presented to the nonattended ear were words and an occasional digit. At various intervals after the presentation of the digit, a light signaled that the subject was to cease shadowing and attempt to recall any digit that had occurred in the nonattended ear recently. Whereas good and poor readers recalled the digit equally if tested immediately after presentation, the poor readers showed a faster decline in recall of the digit as retention interval increased. A fourth experiment was conducted to determine whether the differences in echoic memory were specific to speech stimuli or occurred at a more basic level of aural persistence. Bursts of white noise were separated by 9-400 ms of silence and the subject was to say whether there were one or two sounds presented. There were no differences in detectability functions for good and poor readers.

Working-memory capacity, proactive interference, and divided attention: limits on long-term memory retrieval.

Two experiments examined how individual differences in working-memory capacity (WM) relate to proactive interference (PI) susceptibility. We tested high and low WM-span participants in a PI-buildup task under single-task or dual-task ("load") conditions. In Experiment 1, a finger-tapping task was imposed during encoding and retrieval of each list; in Experiment 2, tapping was required during encoding or retrieval. In both experiments, low spans showed greater PI than did high spans under no load, but groups showed equivalent PI under divided attention. Load increased PI only for high spans, suggesting they use attention at encoding and retrieval to combat PI. In Experiment 2, only low spans showed a dual-task cost on List 1 memory, before PI built up. Results indicate a role for attentional processing, perhaps inhibitory in nature, at encoding and retrieval, and are discussed with respect to theories of WM and prefrontal cortex function.

Working-memory capacity as long-term memory activation: an individual-differences approach.

One explanation of the correlation often observed between working-memory span scores and reading comprehension is that individuals differ in level of activation available for long-term memory units. Two experiments used the fan manipulation to test this idea. In Experiment 1, high- and low-working-memory Ss learned a set of unrelated sentences varying in the number of shared concepts (fan) and then performed speeded recognition for those sentences. Low-working-memory Ss showed a larger increase in recognition time as fan increased. When the slope of the fan effect was partialed out of the relationship between working-memory span and verbal abilities, the relationship was reduced to nonsignificance. In Experiment 2, Ss learned thematically related sentences that varied in fan. Low-span Ss showed the positive fan effect typically found with thematically unrelated sentences, whereas high-span Ss showed a negative fan effect. The results are discussed in terms of a general capacity theory.

Suffix interference in the recall of linguistically coherent speech.

Four experiments are presented that address the stimulus suffix effect for linguistically coherent spoken materials. In Experiment 1, definitions of low-frequency words were presented for online written recall. Each definition was followed by a nonword speech suffix presented in the same voice as the definition, the same nonword presented in a different voice, or a tone. The results yielded a significant reduction in the recall of the terminal words of the definitions in the speech suffix conditions compared with the tone control. This general pattern was replicated in Experiment 2, in which subjects did not begin their recall until the suffix item or tone was presented, although the magnitude of the suffix effect was reduced in this experiment. In Experiment 3, sentences that were part of a cohesive story were presented for on-line recall. Here, the suffix effect was considerably reduced compared with the suffix effect found with the definitions presented in Experiments 1 and 2. This pattern was replicated in Experiment 4, in which subjects did not begin their recall of the story sentences until the speech suffix or tone was presented. Overall, the results suggest that auditory memory interference can take place for linguistically coherent speech, although the magnitude of the interference decreases as one increases the level of linguistic structure in the to-be-recalled materials. Implications of the present results for current models of natural language processing are discussed.

Individual differences in working memory and comprehension: a test of four hypotheses.

A relationship has consistently been found between measures of working memory and reading comprehension. Four hypotheses for this relationship were tested in 3 experiments. In the first 2 experiments, a moving window procedure was used to present the operation-word and reading span tasks. High- and low-span subjects did not differentially trade off time on the elements of the tasks and the to-be-remembered word. Furthermore, the correlation between span and comprehension was undiminished when the viewing times were partialed out. Experiment 3 compared a traditional experimenter-paced simple word-span and a subject-paced span in their relationship with comprehension. The experimenter-paced word-span correlated with comprehension but the subject-paced span did not. The results of all 3 experiments support a general capacity explanation for the relationship between working memory and comprehension.

Absence of mutagenicity of coralyne and related antileukemic agents: structural comparison with the potent carcinogen 7,12-dimethylbenz[a]anthracene.

The structural similarity between antileukemic alkaloid coralyne and the carcinogenic and antineoplastic hydrocarbon 7, 12-dimethylbenz[a]anthracene, as well as the similarity between the antileukemic alkaloid nitidine and the carcinogenic hydrocarbon 5-methylchrysene, prompted a mutagenicity evaluation of coralyne sulfoacetate, nitidine chloride, the 8-ethyl homolog of coralyne, nitidine methosulfate, and the tetramethoxy analog of nitidine by the Ames method against the histidine-auxotroph strains of Salmonella typhimurium TA-1537, TA-1538, TA-98, and TA-100; 7,12-dimethylbenz[a]anthracene was used as a reference standard. The mutagenicity of these antileukemic compounds was either completely eliminated or drastically reduced, but the mutagenic response was generally high for 7,12-dimethylbenz[a]anthracene. The results suggest that the presence of a quaternary nitrogen atom and alkoxy groups could be important in alleviating the mutagenicity of the parent mutagenic and carcinogenic hydrocarbons.

Serologic markers of hepatitis B virus (HBV) and hepatitis D virus infection in carriers of hepatitis B surface antigen who are frequently exposed to HBV.

Serum hepatitis B e antigen (HBeAg) and HBV DNA are indicators of active replication of HBV, whereas IgM antibody to hepatitis B core antigen (IgM anti-HBc) may indicate an active immune response to chronic HBV infection. Fifty-eight carriers of hepatitis B surface antigen (HBsAg) who had frequent parenteral exposures were studied for the presence of HBeAg, HBV DNA, IgM anti-HBc and hepatitis delta virus (HDV) serologic markers. Active replication of HBV was detected in 36.2% (25% of drug addicts, 16.7% of thalassemia patients, and 46.9% of hemodialysis patients) and seropositivity for IgM anti-HBc in 55.2% of the HBsAg carriers. Among the 39 HBsAg carriers who were negative for HBeAg, IgM anti-HBc was detected significantly more frequently than HBV DNA (46.1% vs. 5.1%, p less than 0.001). Serologic evidence of HDV infection was detected in 35% of drug addicts, 50% of thalassemia patients and in 9.4% of hemodialysis patients. These data revealed that continued replication of HBV was more frequent in hemodialysis patients than in drug addicts and thalassemia patients who are HBsAg carriers and the opposite was true for the prevalence of HDV infection.