Attenuation of wild-type yellow fever virus by passage in HeLa cells.

During the 1960s three different research groups reported that passage of wild-type yellow fever (YF) virus [strain Asibi (YF-Asibi)] in HeLa cells resulted in attenuation of the virus for monkeys so that the virus no longer caused viscerotropic disease. We have repeated and extended this observation to analyse the process of attenuation of YF virus during cell culture passage. A large plaque (LP) variant of YF-Asibi virus became attenuated for both monkeys and mice following six serial subcultures in HeLa cells (YF-Asibi-LP HeLa p6). Thus, attenuation was probably due to a genetic change in the virus population rather than to selective enrichment of a pre-existing variant of YF-Asibi-LP virus. No evidence was obtained to implicate defective interfering particles in the attenuation process. Comparison of the YF-Asibi-LP viruses before and after passage in HeLa cells, using a panel of envelope protein-reactive monoclonal antibodies (MAbs), showed that MAbs which specifically neutralize YF-Asibi-LP virus, and not YF 17D-204 vaccine virus, also neutralized YF-Asibi-LP HeLa p6. This indicated that the epitopes involved in the biological process of neutralization were not altered during attenuation. However, two MAbs that recognize envelope protein epitopes did distinguish between HeLa- and non-HeLa-passaged YF-Asibi-LP virus. One of these (MAb 117) which is YF wild-type-specific, recognized YF-Asibi-LP virus but not YF-Asibi-LP HeLa p6 virus, whereas the other (MAb411), which is YF vaccine-specific, recognized YF-Asibi-LP HeLa p6 virus but not YF-Asibi-LP virus. These results suggest that antigenic changes in the viral envelope protein may determine the relative virulence or attenuation of YF virus.

Analysis of methylated and oxidized purines in urine by capillary gas chromatography-mass spectrometry.

Methylated purine bases from urine can be quantitated by capillary gas chromatography-mass spectrometry with selected ion monitoring. The method is based on a separation of the purines in urine using XAD-2 column chromatography and bonded-phase extraction methods. The extracted samples are derivatized with N-(tert-butyldimethylsilyl)-N-methyltrifluoroacetamide and analyzed by capillary gas chromatography-mass spectrometry. The tert-butyldimethylsilyl derivatives of the purines showed excellent GC and GC/MS properties suitable for low-level detection in extracts of human urine as well as in urine extracts from animals treated with the methylating carcinogen N-nitrosodimethylamine. The oxidized purine bases 7-methyl-8-hydroxyguanine, 8-hydroxyadenine, and 8-hydroxyguanine were also detected in urine. To our knowledge, the latter two modified bases have not been previously reported in normal urine.

The influence of ethionine-supplemented soy protein diet on cell-mediated and humoral immunity.

These studies were designed to investigate the influence of ethionine, a suspected carcinogen, on cell-mediated (CMI) and humoral immunity. It is believed that ethionine, an analog of methionine which is produced by intestinal bacteria, could have significant relevance to health. To study the effect of ethionine on immune responsiveness, three groups of mice were allowed to feed ad libitum for 5 weeks on one of the following regimens: diet 1, a basal diet of 16% soy protein; diet 2, soy protein supplemented with 0.6% dl-methionine; and diet 3, soy protein supplemented with 0.1% dl-ethionine. The immunological parameters measured were responsiveness to mitogens, [phytohemagglutinin (PHA), concanavalin A (Con A), pokeweed mitogen (PWM), and lipopolysaccharide (LPS)], delayed-type hypersensitivity (DTH) to dinitrofluorobenzene (DNFB), and antibody formation to sheep red blood cells (SRBC). There were no significant differences in mitogen and antigen responses in mice maintained on diets 1 and 2 as measured by thymidine uptake in proliferating lymphocytes. However, there was a significant suppression in mitogen responsiveness in mice that received diet 3. DTH was also suppressed in mice on diet 3. Antibody levels were similar in all groups. Thus, there was clear evidence of suppression of CMI by ethionine in these studies.

Comparison of operant- and independence-training procedures for mentally retarded adults.

Seventy-five moderatley to profoundly retarded adults in an exit unit of a large state institution were randomly assigned to one of three groups: standard treatment, independence training, and no treatment control. Pre- and posttreatment measures, with progress being defined as an increase in the percentage of operationally defined steps completed for one behavior, were taken for all three groups on one of two types of targeted self-help behavior. A significant increase in correctly completed steps of the target behavior from pre- to posttreatment, as well as from pretreatment to follow-up assessment, was noted for the independence-training group. Independence training proved to be significantly more effective, and staff members preferred it over both standard treatment and no-treatment control at the postassessment.