Viral RNA Titers Research Articles

Outpatient biopsies of the palatine tonsil: access to lymphoid tissue for assessment of human immunodeficiency virus RNA titers.

Our objective was to assess the feasibility of using tonsillar lymphoid biopsy specimens obtained on an outpatient basis to quantitate a patient's lymphoid human immunodeficiency virus (HIV) RNA titers. A pilot cohort study was performed. We evaluated ten HIV-seropositive patients who ranged in age from 26 to 48 years and had CD4+ cell counts ranging from 110 to 833 at enrollment. The main outcome measures were tolerance and safety of outpatient tonsil biopsies and quantitation of HIV RNA titers in tonsillar lymphoid biopsy specimens, plasma, and peripheral blood mononuclear cells determined by a new method of HIV RNA signal amplification with branched DNA probes. Outpatient tonsil biopsies were well tolerated and were performed without complications. Nine of 10 tonsil biopsies from the HIV-seropositive patients examined were positive for significant concentrations of HIV RNA, ranging from 106 to 101 HIV RNA equivalents per gram of tissue. All of the HIV RNA-positive tonsillar lymphoid specimens had HIV RNA titers that were 101 to 104 times greater than those recovered from plasma (per milliliter) of the same patient obtained at the time of biopsy. Sufficient tonsillar tissue can be obtained in an outpatient clinic setting to quantitate lymphoid HIV titers by the new branched-DNA signal amplification method with relative ease and without complication. The biopsy method described here affords ready access to the lymphoreticular system, which may help to advance our understanding of the pathogenesis of myriad immune diseases without the need for excisional node biopsies.

A monoclonal natural autoantibody that promotes remyelination suppresses central nervous system inflammation and increases virus expression after Theiler's virus-induced demyelination

We have used an established experimental model of multiple sclerosis to investigate the potential beneficial relationship between natural autoimmunity and remyelination after central nervous system (CNS) demyelination. Intracerebral infection of SJL/J mice with Theiler's murine encephalomyelitis virus (TMEV) produces chronic, progressive, inflammatory CNS demyelination. Chronically infected SJL/J mice show minimal spontaneous remyelination, which is in part due to a T cell-mediated immune response inhibiting myelin repair. We previously identified a monoclonal natural autoantibody, designated SCH94.03, that promotes remyelination when passively transferred to chronically infected SJL/J mice. The mechanism whereby SCH94.03 promotes remyelination is unknown, although previous reports suggest that natural autoantibodies can modulate immune system function. In this report we demonstrate that treatment with SCH94.03 reduced by 2- to 3-fold the number of CD4(+) and CD8(+) cells infiltrating the CNS of SJL/J mice chronically infected with TMEV, in the absence of global lymphocyte depletion. Associated with the decreased inflammation was a 2- to 3-fold increase in virus antigen expression without a significant increase in viral RNA or virus titers. Treatment with SCH94.03 also suppressed the humoral immune response to a T cell-dependent antigen in chronically infected mice. Immunohistochemical staining showed that SCH94.03 labeled MHC class II-positive dendritic cells in peripheral lymphoid organs. These results are consistent with the proposed immunomodulatory function of natural autoantibodies and suggest that one mechanism whereby SCH94.03 promotes CNS remyelination in chronically infected SJL/J mice is through inhibition of a pathogenic immune response.

Does hepatitis C viral (HCV) genotype or initial HCV RNA titer predict induction of response to interferon alfa 2a in chronic hepatitis C: WM Lee with the Hepatitis Study Group, Roche Clinical Research and Roche Molecular Systems, Nutley and Branchburg, NJ, and the University of Texas Southwestern Med School, Dallas, TX

Does hepatitis C viral (HCV) genotype or initial HCV RNA titer predict induction of response to interferon alfa 2a in chronic hepatitis C: WM Lee with the Hepatitis Study Group, Roche Clinical Research and Roche Molecular Systems, Nutley and Branchburg, NJ, and the University of Texas Southwestern Med School, Dallas, TX

Superinfection of chimpanzees carrying hepatitis C virus of genotype II/1b with that of genotype III/2a or I/1a

Three chimpanzees persistently infected with hepatitis C virus of genotype II/1b were challenged with hepatitis C virus of genotype III/2a and 6 wk later with hepatitis C virus of genotype I/1a. They were tested for titers of total and genotype-specific hepatitis C virus RNA, as well as for serum transaminase levels, until 52 wk after the first challenge. One chimpanzee (CH489) with intermittent low hepatitis C virus RNA titers of genotype II/1b in serum was superinfected with hepatitis C virus of genotype III/2a between wk 1 and 7 after the challenge; superinfection was accompanied by fluctuating high transaminase levels. Later, the animal was superinfected with hepatitis C virus of genotype I/1a. Superinfection was accompanied by persistently high transaminase levels immediately after the challenge. Hepatitis C virus of genotype I/1a persisted, whereas hepatitis C virus of genotype II/1b was undetectable 22 wk after the challenge and thereafter. In another chimpanzee (CH353) with intermittent low hepatitis C virus RNA titers of genotype II/1b, hepatitis C virus of genotype III/2a induced fluctuating high levels of serum transaminases without revealing itself in serum. Then, HCV of genotype I/1a superinfected her, induced persistently high transaminase levels and took over HCV of genotype II/1b at 22 wk after the challenge and thereafter. The third chimpanzee (CH451) with persistently high HCV RNA titers of genotype II/1b did not reveal HCV RNA of genotype III/2a in serum after the challenge, although transaminases sharply increased. Low-titered HCV RNA of genotype I/1a was detected at 18 wk after the challenge. He continued to have elevated transaminase levels and high-titered HCV RNA of genotype II/1b throughout the observation period. These results indicate superinfection with HCV of different genotypes in chimpanzees, newly introduced HCV replacing or being expelled by the predecessor HCV with each infection inducing an episode of liver injury. (Hepatology 1994;20:1131–1136).

Superinfection of chimpanzees carrying hepatitis C virus of genotype II/1b with that of genotype III/2a or I/1a

Three chimpanzees persistently infected with hepatitis C virus of genotype II/1b were challenged with hepatitis C virus of genotype III/2a and 6 wk later with hepatitis C virus of genotype I/1a. They were tested for titers of total and genotype-specific hepatitis C virus RNA, as well as for serum transaminase levels, until 52 wk after the first challenge. One chimpanzee (CH489) with intermittent low hepatitis C virus RNA titers of genotype II/1b in serum was superinfected with hepatitis C virus of genotype III/2a between wk 1 and 7 after the challenge; superinfection was accompanied by fluctuating high transaminase levels. Later, the animal was superinfected with hepatitis C virus of genotype I/1a. Superinfection was accompanied by persistently high transaminase levels immediately after the challenge. Hepatitis C virus of genotype I/1a persisted, whereas hepatitis C virus of genotype II/1b was undetectable 22 wk after the challenge and thereafter. In another chimpanzee (CH353) with intermittent low hepatitis C virus RNA titers of genotype II/1b, hepatitis C virus of genotype III/2a induced fluctuating high levels of serum transaminases without revealing itself in serum. Then, HCV of genotype I/1a superinfected her, induced persistently high transaminase levels and took over HCV of genotype II/1b at 22 wk after the challenge and thereafter. The third chimpanzee (CH451) with persistently high HCV RNA titers of genotype II/1b did not reveal HCV RNA of genotype III/2a in serum after the challenge, although transaminases sharply increased. Low-titered HCV RNA of genotype I/1a was detected at 18 wk after the challenge.(ABSTRACT TRUNCATED AT 250 WORDS)

Hepatitis B virus subtypes and hepatitis C virus genotypes in patients with chronic liver disease in Nepal

A total of 145 patients with chronic liver disease, including 20 with chronic hepatitis, 63 with cirrhosis and 62 with primary hepatocellular carcinoma from Nepal were tested for markers of hepatitis B virus or hepatitis C virus infection. HBsAg was detected in 57 (39%) and hepatitis C virus RNA in 12 (8%); the cause of liver disease was not known in the remaining 76 (52%). HBsAg was found in 5 (1.3%) of 379 normal controls, whereas hepatitis C virus-associated antibodies were detected in 13 (3.4%), none of whom was positive for serum hepatitis C virus RNA. Subtypes of 102 HBsAg samples, from patients and asymptomatic carriers, were adw in 35 (34%), adr in 4 (4%) and ayw in 48 (47%); the remaining 15 (15%) were of atypical subtypes such as ad, ay and a. Of 12 hepatitis C virus RNA samples, genotype I was detected in 1, genotype II in 5 and genotype V in 1; the remaining five samples were not to be classified by polymerase chain reaction with primers specific for genotypes I to V deduced from hepatitis C virus core sequences, despite high hepatitis C virus RNA titers in all of them. Sequences of 192 amino acids in the entire E1 region of unclassifiable hepatitis C virus isolates from five patients differed from each other in 17% to 23%, and varied from reported isolates of defined genotypes in 13% to 44%. These results indicate that atypical subtypes of hepatitis B virus and novel genotypes of hepatitis C virus would prevail in Nepal.

Buoyant density of hepatitis C virus recovered from infected hosts: Two different features in sucrose equilibrium density-gradient centrifugation related to degree of liver inflammation

Hepatitis C virus is reported to have a low buoyant density in sucrose. To determine the density of hepatitis C virus in the circulation of infected hosts and its association with the degree of liver inflammation, we examined serum samples from 10 patients who were positive for both hepatitis C virus antibody (C100 antigen) antibody and serum hepatitis C virus RNA. After the serum was ultracentrifuged in sucrose density gradient (10% to 60%), the hepatitis C virus RNA titer in each collected fraction was quantified by means of competitive reverse transcription-polymerase chain reaction. In samples from five blood donors, the hepatitis C virus RNA titer had a single peak at fractions with densities of 1.08 to 1.11 gm/ml. In samples from five patients with ALT abnormalities, the titer had two peaks at fractions with 1.09 to 1.10 gm/ml and 1.22 to 1.25 gm/ml. After the selected samples were treated with detergents and ultracentrifuged, the titer in the 1.08 to 1.11 gm/ml fractions decreased and that in the 1.22 to 1.25 gm fractions increased. This result implied that the hepatitis C virus density changed with removal of the viral envelope by lipid solvents. Thus the buoyant density of hepatitis C virus in sucrose was 1.08 to 1.11 gm/ml for an intact virion and 1.22 to 1.25 gm/ml for what was presumed to be a nucleocapsid. These results demonstrated that HCV virion is a dominant form in the circulation of blood donors without ALT abnormalities. In patients with liver inflammation HCV particles with higher densities of 1.22 to 1.25 gm/ml coexist with virion in the circulation, which might be presumed nucleocapsids.

Quantitation and typing of serum hepatitis C virus RNA in patients with chronic hepatitis C treated with interferon-β

We quantified serum hepatitis C virus RNA titers and determined hepatitis C virus subtypes in chronic hepatitis C patients treated with interferon-beta to investigate relationships among serum ALT response, serum hepatitis C virus titer and hepatitis C virus subtype. Of 146 chronic hepatitis C patients who received interferon-beta therapy, 24 patients with sustained serum ALT normalization (complete responders) and 26 patients without serum ALT normalization (nonresponders) were randomly selected. Detection, typing and quantitation of hepatitis C virus were performed by means of the "single-tube" polymerase chain reaction method. Of the 24 complete responders, 21 (87.5%) became negative for hepatitis C virus RNA, whereas 21 (80.8%) of the 26 nonresponders remained positive. Hepatitis C virus infections with types I, II, III, IV, II + III and III + IV occurred in 0 (0%), 22 (51.2%), 10 (23.3%), 1 (2.3%), 7 (16.5%) and 3 (7.9%) patients, respectively. The mean pretreatment hepatitis C virus RNA titer of complete responders (0.4 +/- 2.0 x 10(4) CID50/ml) was significantly lower than that of nonresponders (3.8 +/- 4.5 x 10(4) CID50/ml) (p < 0.01). Regardless of HCV subtype, patients with more than 10(4) CID50/ml of HCV did not show serum ALT normalization, whereas complete serum ALT response was seen in most cases with less than 10(2) CID50/ml HCV. These results show that mixed infections with different hepatitis C virus subtypes appear to be more common than previously reported and that the pretreatment serum level of hepatitis C virus RNA is a more important predictor of outcome of interferon therapy than is virus genotype.

Quantitation and typing of serum hepatitis C virus RNA in patients with chronic hepatitis C treated with interferon-β

We quantified serum hepatitis C virus RNA titers and determined hepatitis C virus subtypes in chronic hepatitis C patients treated with interferon-β to investigate relationships among serum ALT response, serum hepatitis C virus titer and hepatitis C virus subtype. Of 146 chronic hepatitis C patients who received interferon-β therapy, 24 patients with sustained serum ALT normalization (complete responders) and 26 patients without serum ALT normalization (nonresponders) were randomly selected. Detection, typing and quantitation of hepatitis C virus were performed by means of the “single-tube” polymerase chain reaction method. Of the 24 complete responders, 21 (87.5%) became negative for hepatitis C virus RNA, whereas 21 (80.8%) of the 26 nonresponders remained positive. Hepatitis C virus infections with types I, II, III, IV, II + III and III + IV occurred in 0 (0%), 22 (51.2%), 10 (23.3%), 1 (2.3%), 7 (16.5%) and 3 (7.9%) patients, respectively. The mean pretreatment hepatitis C virus RNA titer of complete responders (0.4 ± 2.0 × 104 CID50/ml) was significantly lower than that of nonresponders (3.8 ± 4.5 × 104 CID50/ml) (p < 0.01). Regardless of HCV subtype, patients with more than 104 CID50/ml of HCV did not show serum ALT normalization, whereas complete serum ALT response was seen in most cases with less than 102 CID50/ml HCV. These results show that mixed infections with different hepatitis C virus subtypes appear to be more common than previously reported and that the pretreatment serum level of hepatitis C virus RNA is a more important predictor of outcome of interferon therapy than is virus genotype. (HEPATOLOGY 1993;18:1319–1325.)

Hepatitis C virus infection in kidney transplant recipients.

To determine the prevalence and significance of hepatitis C virus infection in kidney transplant recipients, paired serum samples collected from 100 renal allograft recipients on admission for kidney transplantation and 1 yr after transplantation were tested for antibody to hepatitis C virus with second-generation enzyme immunoassay and recombinant immunoblot assay and for hepatitis C virus RNA with reverse transcription-polymerase chain reaction. Before kidney transplantation, hepatitis C virus antibody was detected with second-generation enzyme immunoassay in 18 patients (12 second-generation recombinant immunoblot assay-positive, 6 second-generation recombinant immunoblot assay-indeterminate). Nine of 12 second-generation recombinant immunoblot assay-positive and 2 of 6 second-generation recombinant immunoblot assay-indeterminate samples were hepatitis C virus RNA positive. In addition, 7 of 82 patients who had no detectable antibody on second-generation enzyme immunoassay or second-generation recombinant immunoblot assay were hepatitis C virus RNA positive. After kidney transplantation, hepatitis C virus antibody was detected in 19 patients (12 second-generation recombinant immunoblot assay-positive, 7 second-generation recombinant immunoblot assay-indeterminate, 14 seropositive for hepatitis C virus antibody). Eleven of 12 patients with second-generation recombinant immunoblot assay-positive results and 4 of 7 with second-generation recombinant immunoblot assay-indeterminate results were positive for hepatitis C virus RNA. Hepatitis C virus RNA was present in 28 patients 1 yr after kidney transplantation. Six patients appeared to have acquired active hepatitis C virus infection 1 yr after kidney transplantation (seroconverted to hepatitis C virus RNA positivity).(ABSTRACT TRUNCATED AT 250 WORDS)

Hepatitis C virus infection in kidney transplant recipients

To determine the prevalence and significance of hepatitis C virus infection in kidney transplant recipients, paired serum samples collected from 100 renal allograft recipients on admission for kidney transplantation and 1 yr after transplantation were tested for antibody to hepatitis C virus with second-generation enzyme immunoassay and recombinant immunoblot assay and for hepatitis C virus RNA with reverse transcription-polymerase chain reaction. Before kidney transplantation, hepatitis C virus antibody was detected with second-generation enzyme immunoassay in 18 patients (12 second-generation recombinant immunoblot assay-positive, 6 second-generation recombinant immunoblot assay-indeterminate). Nine of 12 second-generation recombinant immunoblot assay-positive and 2 of 6 second-generation recombinant immunoblot assay-indeterminate samples were hepatitis C virus RNA positive. In addition, 7 of 82 patients who had no detectable antibody on second-generation enzyme immunoassay or second-generation recombinant immunoblot assay were hepatitis C virus RNA positive. After kidney transplantation, hepatitis C virus antibody was detected in 19 patients (12 second-generation recombinant immunoblot assay-positive, 7 second-generation recombinant immunoblot assay-indeterminate, 14 seropositive for hepatitis C virus antibody). Eleven of 12 patients with second-generation recombinant immunoblot assay-positive results and 4 of 7 with second-generation recombinant immunoblot assay-indeterminate results were positive for hepatitis C virus RNA. Hepatitis C virus RNA was present in 28 patients 1 yr after kidney transplantation. Six patients appeared to have acquired active hepatitis C virus infection 1 yr after kidney transplantation (seroconverted to hepatitis C virus RNA positivity). We found no correlation between the presence of hepatitis C virus markers in the pretransplant and posttransplant sera and clinical/biochemical parameters or clinical outcome. Hepatitis C virus infection was clinically silent in most patients. Semiquantitation of serum hepatitis C virus RNA in 25 patients with hepatitis C virus infection showed that the hepatitis C virus RNA titer remained the same (n = 12) or increased at least 10-fold (n = 12), and only 1 patient had a decrease in serum hepatitis C virus RNA level. These data suggest that (a) the prevalence of hepatitis C virus infection is high in kidney transplant recipients, (b) active hepatitis C virus replication may be present in the absence of hepatitis C virus antibody, (c) many of these patients who are second-generation enzyme immunoassay-positive but second-generation recombinant immunoblot assay-indeterminate are hepatitis C virus RNA positive, (d) hepatitis C virus infection is usually clinically silent during the initial follow-up period and (e) antirejection immunosuppression may enhance hepatitis C virus replication. (HEPATOLOGY 1993;18:1027-1031).

Changes in serum hepatitis C virus RNA titer and response to interferon therapy

Changes in serum hepatitis C virus RNA titer and response to interferon therapy