D. A. J. Tyrrell

INHIBITION OF RESPIRATORY VIRUS INFECTION BY LOCALLY APPLIED INTERFERON

Abstract In-vitro studies showed several strains of influenza and rhinoviruses to be as sensitive to the antiviral action of human interferon as a known interferon-sensitive virus, provided that a yield-reduction assay was used. In volunteers, a single day of prophylaxis with intranasal interferon only slightly delayed the onset of an influenza-B infection, and did not prevent the illness or reduce its severity. However, by using a greater daily dosage of interferon and combining a day of prophylaxis with 3 days of treatment, a statistically significant prevention of clinical symptoms and virus-shedding was achieved after challenge of volunteers with rhinovirus 4.

Isolation of rhinoviruses and coronaviruses from 38 colds in adults

Nasal washings were collected from 27 normal adults during 38 naturally acquired colds. The washings were exhaustively tested using tissue cultures, organ cultures and electron microscopy. Washings yielding no identifiable agent were inoculated into human volunteers, and further specimens obtained from the latter were examined by the same techniques in vitro. Viruses were identified in association with 25 of the original 38 colds (65.7%). Fifteen were rhinoviruses (39.5%), seven coronaviruses (18.4%), two were parainfluenza viruses, and one was influenza virus. Use of organ cultures and of volunteers significantly increased the isolation rate. No agent was cultivated from the remaining 13 specimens, although tests in volunteers showed that cold‐producing agents were present in five of them (13%). Three specimens gave doubtful results in volunteers, and five others, all collected within a period of six weeks in December and January, apparently contained no infectious agent.

A COMPARISON OF LIVE AND KILLED INFLUENZA-VIRUS VACCINES: Report to the Medical Research Council's Committee on Influenza and other Respiratory Virus Vaccines

Abstract 98 volunteers were vaccinated with formalin-inactivated influenza-B virus intramuscularly or with living influenza-B virus intranasally. They, and a group of control volunteers, were subsequently challenged with live virus administered intranasally. 71% of those given killed vaccine developed an antibody response but 15% were infected on challenge. 51% of those who had received live virus had an antibody response and 2% were infected on challenge. Vaccination also reduced the frequency of clinical reactions. The living vaccine used in this trial produced too many reactions to be used on a large scale.

HUMAN DIPLOID CELL STRAIN RABIES VACCINE: Rapid Prophylactic Immunisation of Volunteers with Small Doses

The clinical and antibody responses of volunteers to three intradermal schedules of human diploid cell strain rabies vaccine (0.4 ml on day 0; 0-1 ml on days 0, 1, 2, 3; and 0-1 ml on days 0, 3, 7, and 14) are described. Vaccine was administered to 114 contacts of two rabid patients in order to evoke a rapid antibody response and the antibody titres of 30 of those who were vaccinated and bled were measured. High antibody titres were obtained in all subjects irrespective of their immunisation schedule; there were only minimal local reactions. All volunteers had titres greater than 1/78 (1.7 I.U./ml) by day 14, and 7 of 10 receiving 0.1 ml into each limb on day 0 had detectable antibody by day 7.

Drug resistant rhinoviruses from the nose of experimentally treated volunteers

SummaryViruses were isolated from nasal washings of volunteers receiving experimental therapy for rhinovirus type 9 infection with intranasal sprays of a new synthetic antiviral R61837. On a screening test nine subjects yielded drug sensitive virus and four resistant virus. In four others the virus was sensitive at first but became resistant later, while in one the reverse occurred. Evidence is given that at least some of the resistant viruses were present in the respiratory tract and were not selected during virus isolation. Of six viruses studied in detail, five had a low degree of resistance and one was highly resistant. The degree of resistance of the five was similar for an antiviral chalcone, dichloroflavan and disoxaril. The sixth was different in that the resistance to disoxaril was relatively less than to the other drugs. The significance of these results is discussed—these are the first experiments in man to show the selection of drug resistant rhinovirus.

Experiments on the spread of colds. II. Studies in volunteers with coxsackievirus A21.

The amount of virus in nasal and other secretions after infection with coxsackievirus A 21 has been measured daily in ten volunteers. Most virus was found in nasal secretion, less in throat secretion and small amounts were found intermittently in the saliva and faeces. Virus administered as small drops or on a swab was more infectious for man if put on to the nasal mucosa than on to the throat or outside the nose. It was also infectious by the conjunctival route. Virus was sprayed in droplets of about the same size range as those found in a natural sneeze. Virus survived better in larger (> 4 μ) than in smaller droplets. About one tissue culture infectious dose of virus in such droplets also infected volunteers. The symptoms produced by these experimental infections have been analysed. The disease produced was largely independent of the dosage and route of infection. Those with pre-existing antibody resisted infection better than those with no antibody. Antibody rises were detected in about two-thirds of infected volunteers. Volunteers with colds shed virus in large drops on sneezing, or into the handkerchief on blowing the nose, but virus was recovered from the air only after simulated sneezes by volunteers with high concentrations of virus in their nasal secretions. Virus died off rapidly on fabric at room temperature and humidity, and was only resuspended as airborne droplets when large doses such as 0·02 ml. of virus of high titre (10 7·5 TCD 50/ml.) were used. Infection was transmitted from an infected volunteer to an uninfected partner living in the same flat in three out of twenty tests. Infection was not transmitted in experiments when volunteers mixed for a few hours with subjects with colds, or inhaled air into which a subject with a cold had just sneezed.

Use of synthetic oligonucleotide probes to detect rhinovirus RNA

SummaryCurrent methods of detecting a human rhinovirus (HRV) infection are either based on isolation of virus in appropriate susceptible cell lines, which is time-consuming and requires considerable expertise, or are dependent on knowing the serotype. The extistence of over 100 immunologically distinct serotypes makes serotype specific assays, such as ELISA, unsuitable for general diagnostic assays. In this study a general rhinovirus assay is described which utilises synthetic oligonucleotides as probes in a filter hybridization assay. The probes are designed to bind to short but highly conserved regions of the rhinovirus genome. Indeed, the probes successfully detected all 57 rhinovirus serotypes tested. Furthermore, the test was used to demonstrate rhinovirus infection in clinical samples from 57 volunteers, inoculated with HRV, collected on six consecutive days. Clinical samples were taken prior to inoculation and on days 2–7 after inoculation. The filter hybridization assay gave results comparable to virus culture on days 2 and 3 post-inoculation, but was more sensitive on subsequent days.

Detection of Human Rhinoviruses and Their Molecular Relationship Using cDNA Probes

We describe here a cDNA: RNA hybridization system for the study of human rhinoviruses. We have constructed an M13 probe from the 5′ end of the genome of rhinovirus 14 (HRV‐14) and used this to detect directly viral RNA. Of the 56 human rhinoviruses so far investigated 54 or 96.4% gave clearly positive hybridization signals. However, the strength of this signal depended very much on the molecular relationship of these viruses. Thus, HRV‐3, 4, 17, 72, and, to a slightly lesser extent, HRV‐2, 6, 9, 13, 19, 31, 42, 49, 64, and 69 appear to be closely related to HRV‐14 whereas HRV‐5, 7, 8, 16, 32, 40, 45, 55, 56, 63, 80, 82, and 85 appear to be relatively divergent. Further, evidence is provided in this study that indicates that it would be feasible to use cDNA probes to detect human rhinoviruses in nasal washings. However, the sensitivity of detection was clearly affected by both the inclusion of inhibitors of endogenous RNase activity in the RNA extraction mixture and also in the method of extracting the viral RNA. From reconstruction experiments in nasal washings and under optimal conditions, we can detect virus at 102.8 TCID50/ml.

Studies of experimental rhinovirus type 2 infections in polar isolation and in England.

After five months of total isolation a wintering party of seventeen British Antarctic Survey (BAS) personnel was inoculated under double blind concitions with placebo, or rhinovirus type 2 which had been propagated in tissue culture. The clinical and virological responses of these subjects were compared with those of volunteers in England who received a similar dose of the same strain. The virus used was apparently partly attenuated for man; at the dosage used its effects in England were similar to a smaller dose of an unattenuated strain, but in the Antarctic it caused relatively severe infections. Both the symptoms and the laboratory evidence of virus infection appeared to be more pronounced in the BAS subjects than in the volunteers in England who received the same challenge. In the former group the infection readily spread to those who were originally given placebo. In the BAS subjects serum antibody titres were well maintained during the isolation period but a significant fall in nasal immunoglobulin concentration was recorded during the 5 months of isolation after the virus challenge. Possible mechanisms for the increased sensitivity to rhinovirus of subjects who have been totally isolated in a small closed community are discussed.

Studies on a rhinovirus (EC11) derived from a calf: I. Isolation in calf tracheal organ cultures and characterization of the virus

Abstract A virus having the characteristics of a rhinovirus was isolated from a calf with a respiratory illness. Isolation of the virus in calf tracheal organ cultures was achieved after initial failure of attempts to transmit the agent in monolayer cultures. After eight serial passages in organ culture the virus was adapted to growth in calf kidney monolayers. Except for its initially very poor growth in calf kidney cells the virus appears similar to other bovine rhinoviruses. Pools of bovine sera were found to have neutralising activity against this virus, but it was not neutralised by an antiserum against the Sd-1 rhinovirus of Bogel and Bohm.