Measles infection of human mononuclear cells. I. Acute infection of peripheral blood lymphocytes and monocytes.

A study of the susceptibility of human peripheral blood mononuclear cells to measles virus infection and replication is reported. Resting lymphocytes obtained from adults showed very low levels of infection and virus replication while lymphocytes activated by plant mitogens or allogenic lymphocytes supported mononuclear cells obtained from the umbilical cord of healthy neonates were more susceptible to measles virus infection than those of adults; however, activated cord lymphocytes supported viral replication in the range observed with adult activated lymphocytes. Monocytes obtained from adults were relatively resistant to measles virus infection and replication while neonatal cord blood monocytes supported viral replication to the degree observed with activated lymphocytes. It is hypothesized that infection of acitivated lymphocytes may explain the depression of cell-mediated immunity seen during acute measles virus infection. The significance of the finding that neonatal monocytes are more susceptible to viral infection and replication than adult monocytes is discussed.

HIV-1 inhibition by azidothymidine in a concurrently randomized placebo-controlled trail.

Two independent measures of human immunodeficiency virus type 1 (HIV-1) infection, virus isolation, and serum levels of p24 antigen were evaluated in a double-blind randomized clinical trial of the safety and efficacy of a nucleoside analogue, 3'-azido-3'-deoxythymidine (AZT) versus placebo in a single center. Pretreatment studies from 38 AIDS and AIDS-related complex (ARC) patients were comparably positive for virus isolation from their lymphocytes; all patients were qualitatively virus positive. Before AZT treatment, there was significantly decreased virus recovery in patients with higher numbers of CD4-positive lymphocytes. Within 1 month of AZT therapy, the time in culture required to register virus positivity was increased markedly in the AZT-treated group, and over the following several months progressive diminution in virus recovery was noted. Similar changes were not seen in patients concurrently receiving placebo treatment. Before treatment, 16 of 20 and 12 of 16 patients in the AZT and placebo groups, respectively, were p24 antigen positive. Marked reduction in serum p24 levels were noted in 11 of 16 (69%) of the p24 antigen-positive AZT-treated patients compared to 3 of 12 (25%) of the p24 antigen-positive placebo-treated patients (p = 0.02). There was a marked virologic response in 14 of 20 (70%) of the AZT-treated patients compared to 4 of 18 (22%) placebo-treated patients (p = 0.004). A higher frequency of positive clinical and immunological effects also were noted in the AZT-treated patients relative to placebo-treated patients (p = 0.02 and p = 0.06, respectively).

Spectrum of antiviral activity and mechanism of action of zidovudine. An overview.

Zidovudine is a potent in vitro inhibitor of human immunodeficiency virus (HIV) with varying efficacy against other retroviruses. With the exception of Epstein-Barr virus, all non-retroviruses tested so far have been insensitive to inhibition by zidovudine. In vivo, efficacy of zidovudine was demonstrated against Rauscher murine leukemia virus and feline leukemia virus. In both experimental models, infections completely resolved in animals when the drug was administered soon after infection. These results suggest that prompt initiation of zidovudine therapy, following a known exposure to HIV, should be considered. Mechanism studies show that zidovudine is phosphorylated to the monophosphate and diphosphate derivatives by the host cell cytosolic thymidine kinase and thymidylate kinase, respectively. The identity of the enzyme that phosphorylates zidovudine diphosphate is not known, but is believed to be the cellular nucleoside diphosphate kinase. The triphosphate of zidovudine appears to be the active form of the drug. Zidovudine triphosphate competes well with thymidine 5'-triphosphate for binding to the HIV reverse transcriptase and also functions as an alternative substrate. Incorporation of zidovudine monophosphate results in chain termination. However, it is not clear which mechanism, chain termination or competition with thymidine 5'-triphosphate, or a combination of both, is responsible for the inhibition of HIV replication.

Disease and latency characteristics of clinical herpes virus isolated after acyclovir therapy.

Herpes simplex virus (HSV) type 1 from a bone marrow transplant recipient and HSV type 2 from a patient with genital herpes infection were examined for sensitivity to acyclovir after both patients received therapy with the drug. A 38- and 83-fold shift in sensitivity was detected in association with a marked decrease in viral thymidine kinase activity in isolates from both patients. The resistant HSV-1 isolate was approximately 900 times less neurovirulent to Balb/C mice but had similar cutaneous virulence in hairless mice compared with the patient's sensitive strain. In contrast, there was no difference in pathogenicity between the sensitive and resistant HSV-2 isolates. Latency was detected in the trigeminal ganglia of mice after snout inoculation with both the sensitive and resistant HSV-1 isolates. The ganglion isolate from the resistant HSV-inoculated mouse was found to be sensitive to acyclovir, implying a selection for or reversion of the sensitive phenotype. No trigeminal ganglion latency was detected after inoculation with either HSV-2 isolate. Resistance to acyclovir can arise during therapy as a result of diminished viral thymidine kinase activity but does not appear to be associated with increased virulence.

Spectrum of sensitivity of acyclovir of herpes simplex virus clinical isolates.

HSV-1 and HSV-2 clinical isolates were tested for their in vitro sensitivity to acyclovir. The median ID50 for 32 genital HSV-2 isolates was 0.215 micrograms/ml and this was not significantly higher than a median value of 0.125 micrograms/ml for 28 oral HSV-1 isolates (p = 0.08). A wider range of ID50s was observed with the HSV-2 isolates compared with the HSV-1 isolates. The concentration of drug required to inhibit virus replication by 90 and 99 percent was approximately 10- and 100-fold greater than that producing 50 percent inhibition.

Oral acyclovir therapy of genital herpes simplex virus type 2 infections in guinea pigs.

Oral acyclovir was evaluated for its effectiveness in treating guinea pigs with primary herpes simplex virus type 2 infections. Guinea pigs inoculated intravaginally with acyclovir-susceptible strains (for which 50% inhibitory concentrations of acyclovir in cell culture were found to be in the range of 0.15 to 1.2 micrograms/ml) and treated with 5.0 mg of acyclovir per ml in the drinking water beginning 48 h postinfection showed significant reductions in lesion severity. This dosage produced serum acyclovir levels of 1.3 micrograms/ml. Lower concentrations of oral acyclovir (less than or equal to 2.5 mg/ml in the drinking water), which produced serum acyclovir levels of less than 1.0 microgram/ml, were less consistently effective against these same virus strains. When an acyclovir-resistant isolate (for which the 50% inhibitory concentration of acyclovir in cell culture was found to be 8.5 micrograms/ml) was used to initiate infection, treatment with 5 or 10 mg/ml (yielding serum levels of 1.3 and 3.5 micrograms/ml) in the drinking water had only minimal clinical benefit. However, the degree of response was difficult to determine because of the attenuated disease produced by the acyclovir-resistant virus. In vitro virus sensitivity may be predictive of the serum drug levels that need to be obtained to produce a successful response to therapy.

Clinical and laboratory experience with acyclovir-resistant herpes viruses.

The emerging field of antiviral sensitivity testing must depend upon some of the lessons learned from prior experience with anti-bacterial sensitivity testing, but also take into account additional factors including intracellular drug distribution and metabolism as well as the unique features of viral replication. There is an urgent need to standardise in-vitro sensitivity testing, and an international collaborative study is essential so that different laboratories may compare and define their sensitivity results in the context of the outcome of therapy in infected patients. It is only by such a study that the influence of patient and therapeutic factors such as immune function, anatomical location of the infection and dosage regimens used can be understood.

Resistance to arbovirus challenge in mice immediately after vaccination.

Mice vaccinated with a single injection of formalin-inactivated suspensions of mouse brain tissue infected with arboviruses were markedly protected against a challenge injection administered hours later. The protection observed during the first 2 days after vaccination seemed to be nonspecific, in that it appeared not only with the homologous system but also between arboviruses of different antigenic groups; this phase may be associated with an interferon-like activity of the serum. Overlapping the nonspecific phase was one of specific protection, which seemed to be well-established in its own right by day 3 or 4 after vaccination. Serum neutralizing antibodies against the homologous viruses were detected as early as 24 h after vaccination and in almost all instances by day 3.

Quantitation of influenza vaccine hemagglutinin by immunoelectrophoresis.

A technique for the quantitation of hemagglutinin (HA) in influenza vaccines by immunoelectrophoresis (IEP) is described. This method gives accurate, and reproducible results and requires approximately 9 h to complete. The HA content of four manufacturers' vaccines correlated with the chick cell agglutination (CCA) values for each manufacturers' vaccine. However, when comparisons were made between vaccines produced by different manufacturers, up to a four-fold difference in HA content was observed. Comparison with single radial immunodiffusion (SRID) gave comparable results for vaccines produced by three of four manufacturers; the fourth manufacturers' vaccine gave 1.6-fold higher results by SRID. Human seroconversion rates in seronegative individuals correlated better with IEP determined HA content than the current assay of vaccine potency using CCA.

Inactivated influenza vaccine efficacy: diminished antigenicity of split-product vaccines in mice.

Groups of 60 to 120 mice were given a single intraperitoneal inoculation of varying dilutions of commercially prepared and licensed bivalent (A/England and B/Mass) and monovalent (A/Aichi or B/Hong Kong) inactivated influenza vaccines, and their antibody responses at 14 days were quantitated by hemagglutination inhibition tests. Split-product vaccines prepared by the treatment of A/England, B/Mass, and B/Hong Kong whole virus with Tween-80 and either tributylphosphate or ether produced significantly lower mean antibody titers than did equivalent whole-virus preparations. The rates of seroconversion (<1:8 to >/=1:8) at the various dilutions tested were also significantly reduced when these split-product vaccines were given. When the antigen content of all vaccines was quantitated by the chick cell agglutination test, between 10 and 100 times more split-product antigen than whole-virus antigen was required to produce seroconversion in 50% of the mice tested. Differences between split-product and whole-virus A/Aichi vaccines were less marked. These data point out the need to consider factors other than hemagglutinin content alone in determining the immunogenicity of inactivated influenza vaccines.

Antibody production and protection against influenza virus in immunodeficient mice.

The roles of T and B cells in the immune response to influenza virus were studied by using mice deficient in either T cells (athymic nude) or immunoglobulin production (CBA/N). The serological responses of these mice to either whole or disrupted A/Aichi/2/68 influenza virus vaccines were examined, and the protective effect of these inoculations was tested by challenge infection with mouse-adapted A/Aichi/2/68 influenza virus. In contrast to normal mice, neither strain of immunodeficient mouse produced detectable serum antibody after inoculation with either type of vaccine. CBA/N mice immunized with intact virus vaccine were protected, however, against subsequent lethal challenge. CBA/N mice inoculated with disrupted virus vaccine and nude mice inoculated with either disrupted or whole virus vaccine were not protected against viral challenge. Evidence of immunological memory was observed in CBA/N and nude mice that had survived live virus challenge after immunization with inactivated vaccine.

In vitro sensitivity to acyclovir in genital herpes simplex viruses from acyclovir-treated patients.

Genital isolates of herpes simplex virus (HSV) from patients given acyclovir or placebo were tested in vitro for sensitivity to acyclovir. Isolates obtained before therapy were sensitive to acyclovir concentrations of 0.01-19 micrograms/ml, with 86 of 97 isolates inhibited by less than 1 microgram/ml. Before therapy, six patients had isolates of HSV type 2 with ID50 values (concentrations of drug reducing viral cytopathic effect by 50%) of greater than 3 micrograms/ml. Plaque purification revealed mixed populations of virus; some clones were associated with high and some with low rates of acyclovir phosphorylation. Sensitivity to acyclovir decreased in isolates obtained after therapy from four of 25 patients given acyclovir and three of 30 patients given placebo. The occurrence of this change with similar frequency in the two groups suggests that factors other than the use of acyclovir influence the in vitro sensitivity of clinical HSV isolates to this agent. In one patient in whom an acyclovir-resistant, thymidine kinase-negative strain of HSV type 2 emerged during therapy, infection subsequently recurred. The isolate responsible for the recurrence was sensitive to acyclovir and had a high level of acyclovir-phosphorylating activity.

Isolation of foamy virus from rhesus, African green and cynomolgus monkey Leukocytes.

Foamy virus (FV) was recovered regularly from the leukocyte of rhesus and cynomolgus monkeys and somewhat less often from African green monkey leukocytes. Virus was found in virtually all organs of experimentally infected rhesus monkeys. No illness or pathologic abnormalities were noted in these animals or in any of the naturally infected animals in spite of the prolonged period of viral persistence in various organs and tissues.

Recurrent genital herpes and suppressive oral acyclovir therapy. Relation between clinical outcome and in-vitro drug sensitivity.

To evaluate the association between in-vitro resistance of herpes simplex virus type 2 to acyclovir and breakthrough recurrences of herpes despite chronic suppressive therapy, we determined the in-vitro sensitivity of herpes simplex virus isolated before, during, and after therapy. One hundred eighty-three virus isolates from 107 patients were tested. Before therapy, the median amount of drug required to inhibit 50% of the virus in tissue culture (ID50) was 0.91 microgram/mL. The median ID50 after therapy was 0.99. Six isolates from patients with culture-positive breakthrough recurrences were evaluated. The median ID50 was 0.90 microgram/mL (range, 0.39 to 1.55). The development of breakthrough recurrences could not be correlated with infection with strains of herpes simplex virus type 2 that were resistant to acyclovir in vitro. Acyclovir-resistant strains are not commonly recovered from patients during acyclovir therapy, nor does there seem to be a high frequency of resistance after 4 months of chronic suppressive therapy.

Rapid screening of antiretroviral combinations.

Increasing evidence supports the view that therapy with combinations of antiretroviral drugs provides greater and more sustained benefits in the treatment of HIV infection than either monotherapy or sequential therapy. As the number of licensed and developmental antiretroviral agents grows, in vitro analysis is increasingly being used to aid in the selection of effective combinations. An assay has been designed to ascertain the inhibitory action of drug combinations on HIV-infected MT4 cells, allowing rapid evaluation of those that may be of use in the clinic. Manipulation of this system also provides data on the efficacy of drugs under conditions of high viral load and against resistant strains, providing valuable information for the treatment of antiretroviral-experienced patients with advanced disease.

In vitro comparison of selected triple-drug combinations for suppression of HIV-1 replication: the Inter-Company Collaboration Protocol.

Ten different three-drug combinations have been analyzed for their ability to prevent HIV-induced cytopathic effects (CPEs) in a continuous human T-lymphoblastoid cell line. Agents acting at the same as well as at different sites in the HIV-1 replicative cycle were used. Each compound was analyzed at peak and trough plasma levels achieved in monotherapy and in the presence of HIV-1 strains 3B and MN at a viral inoculum varying from 1 x TCID50 (50% tissue culture inhibitory dose) to 1,000 x TCID50. Using a viral inoculum of 10 x TCID50 HIV-1 3B, it was determined that triple-drug combinations had greater antiviral activities than the corresponding double-drug combinations, which had greater antiviral activities than zidovudine (AZT) monotherapy. The most consistent triple-drug combination, demonstrating superior activity at all concentrations of virus, was AZT + dideoxyinosine + lamivudine which reduced the AZT IC95 (95% inhibitory concentration) by 208-, 57-, 133-, and 25-fold at of 1,000, 100, 10, and 1 x TCID50 HIV-1 3B, respectively, as compared with the IC95 for AZT monotherapy. For all antiviral regimens tested, higher viral inoculum resulted in less inhibition of viral replication and a higher IC95 for AZT. This observation argues for therapeutic intervention at an earlier stage in HIV infection, when viral burden is lower.