Strain Dairen I (DI) and strain IHD-T of neurovaccinia virus were readily inactivatedin HCl-citrate buffer solution, pH 2.2 or 2.6, at room temperature, 23 C, reaching about10-3 or less survival in 2 min and 10-4 or less in 5 min. Treatment of L cells with thebuffer solution, pH 2 to 2.5, at room temperature for 5 min after virus adsorption at37 C effectively reduced the titer of residual active virus, and provided a simple, effectivemethod to unmask the low-grade replication of strain DI with little deleteriouseffect on the cells. The method seems useful for the analytical study on the viralreplication, particularly the early virus-cell interactions. The clonal analysis, preliminaryin nature, of the viral yield of DI infected L cells provided data suggestive ofselection for viruses better adapted to L cells

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Numerical classification was carried out using a "ghypothetical mean organism"h and11 species of the genus Mycobacterium and 6 species of the genus Nocardia. Statisticalsurvey of the results indicated the presence of three taxonomic groups, the slowlygrowing mycobacteria, the rapidly growing mycobacteria and nocardiae. The presentauthor had previously divided the genus Mycobacterirum into two subgenera, Mycobacteriumand Rlycomycobacterium, which corresponded approximately to slowlygrowing mycobacteria and rapidly growing mycobacteria, respectively. In the numericalclassification, these two subgenera behaved as if they were two genera. It was suggestedthus that these two subgenera of genus Mycobacterium should be transferred to twogenera. Beside morphological characters, three biochemical characters were shown to beuseful for differentiating between rapidly growing mycobacteria and nocardiae: (I) Atwo-week arylsulfatase activity; (2) Acid formation from mannose; (3) Utilization forgrowth of trimethylene diamine as a simultaneous nitrogen and carbon source.

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In order to provide further information on the chemical nature of the antigenicdeterminants of the mannan of Saccharomyces cerevisiae, the mannan was digested by Arthrobacter α-mannosidase, and 9, 21, 35, 59 and 62%-partially degraded mannanswere prepared in the present study. Acetolysis of each degraded mannan showed thatonly a small amount of the tetrasaccharide was detectable in the 35%-digested mannan, whereas the predominant product of the 59 and 62%-digested mannan was mannose.The result of a quantitative precipitation reaction with the degraded mannans showedthat the precipitation activities were partially or completely destroyed by the action ofthe enzyme. The lack of the tetrasaccharide moieties of the mannan were noticeableby a decrease in the precipitating ability. It was observed that the decreasing ratio ofeither the maximum amount of the antibody N precipitable by the mannan or percent degradation of the mannan were essentially equal and yielded nearly a straightrelationship between 0 and 2.0 hr digestion. However, the 59 and 62%-digested mannans, containing trace amounts of di- and trisaccharides in the branching parts, showedno significant antigenic activities. Furthermore, the molar ratio of the tetrasacchariderelative to the trisaccharide also gradually decreased. These observations confirm thatthe tetrasaccharide moiety, Manα1→3Manα1→2Manα1→2Man, plays an importantrole as the antigenic determinant. The core mannan moiety completely lost both theprecipitating ability and inhibitory activity in ranges employed up to 1500μg. Thesefindings offer a direct proof that the core mannan moiety of mannan is not responsiblefor antigenic activity, and functions merely as the "gcarrier" of the antigenic determinantswhich dominate the immunological specificity.

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The relationship between the number of circulating leukocytes and amount ofvirus-inhibiting factor (IF) or interferon appearing in blood stream in response toinjection of E. coli endotoxin or Newcastle disease virus (NDV) was studied in miceafter whole-body X-irradiation. Two days after irradiation with 500R, the leukocytecount fell to 7% of the normal value. Production of IF, however, was not depressedwhen induced with endotoxin 2 days after irradiation, while NDV induced a slightlylower titer of IF, 1/1.6 of that in non-irradiated mice. When induced 5 days afterirradiation, though leukocyte count remained at 8 to 9% of the normal value, IF titersattained were only 1/3 and 1/4 of the control value with endotoxin and NDV, respectively.In mice given 1000 R before 2 days, leukocyte number was less than 1% of thenormal value, while IF titers reached 1/2.7 and 1/5 of that in the control mice withendotoxin and NDV, respectively. The lack of parallel between the leukocyte count andamount of IF produced suggest that circulating leukocytes do not produce IF.

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It was demonstrated that spiramycin (SP)-resistance could be related to the decrease in binding of ribosomes to SP and that the SP-binding to ribosomes was related with inhibition of polypeptide synthesis by SP in a cell-free system in staphylococcal strains. These facts were also observed in Mac (macrolide)-inducible strains, in which resistance to Mac antibiotics is enhanced by prior treatment with subinhibitory concentrations of erythromycin. From these results, it was concluded that the mechanism of resistance to Mac antibiotics is accounted for by alteration of ribosomes in staphylococcal strains and that this alteration of ribosomes is caused not only by mutation but also by induction.

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Extracts of tobacco leaves, 3 to 22 hr after infection with ³²P-labeled tobacco mosaic virus (TMV), were analysed by sucrose density-gradient centrifugation. As reported previously, a shoulder appeared on the lighter side of the main peak representing intact parent viruses. The shoulder was shown to contain partially uncoated virus particles with sedimentation constants of about 140-150 S, which withstood dialysis against a phosphate buffer. RNA from the uncoated particles was isolated on the sucrose densitygradient, and its sedimentation constant was estimated at 16-18 S. Intact TMV-RNA and RNA having 16-18 S were also obtained directly from homogenates of tobacco leaves infected with ³²P-TMV. ³²P-TMV was partially stripped by SDS and was added to tobacco leaf sap to test the RNase activity in tobacco leaves. A part of RNA which was exposed by SDS was digested and the ³²P-radioactivity in the top zone of the centrifugation tube increased. This finding supported the idea that the partially uncoated virus particles in infected leaves lost a portion of their RNA during extraction by the action of the RNase contained in the homogenate.

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In a survey of 228 tuberculous patients who had received kanamycin (KM) treatment, KM-resistant Escherichia coli or Escherichia freundii were isolated from 29. From the patients who had not received KM-therapy, KM-resistant strains were not isolated. Of the 29 KM-resistant strains isolated, 5 E. coti strains were also resistant to other aminoglycoside antibiotics, i.e., paromomycin, fradiomycin and streptomycin. They were also resistant to chloramphenicol and sulfanilamide and all of these resistances could be transferred to E. coli strains by conjugation. The resistance was considered to be conferred by an R factor. The remaining 24 KM-resistant strains were identified as E. freundii. In these level of KM-resistance was rather low and conjugal transfer was not observed. All were resistant to paromomycin but sensitive to fradiomycin. When 381 KM-sensitive strains were plated on an agar plate containing 25μg of KM/ml, 23 KM-resistant strains were isolated. The mutants thus obtained were similar to the second type of 24 KM-resistant strains isolated from patients. Thus it was strongly suggested that the 24 strains carrying low levels of KM- and paromomycin-resistance, that is, conjugally nontransmissible, appeared by spontaneous mutation in the intestines of patients who bad received KM-treatment.

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A limited multiplication of Mycobacterium lepraemurium was observed in cultures of mouse foot-pad cells up to the third sub-culture. After 96 hr of incubation at 33 C for phagocytosis, infected cells were washed to remove unphagocytized bacteria and transferred to new culture vessels. There was a 4.9-fold increase in the number of bacteria during 70 days of successive cultures up to the third culture in flasks, giving an overall generation time of 30.4 days. Acid-fast stained cover-slips taken from the culture in Leighton tubes revealed intracellular multiplication of M. lepraemurium, frequently in bundles of hundreds of bacteria, and gave an average generation time of 16.4 days in the primary, 15.7 days in the secondary, and 8.1 days in the tertiary culture. Elongation of M. lepraemurium was observed as early as 6 days after infection. All attempts to grow the acid-fast bacteria from the original bacillary suspension and the cell suspension at the end of the experiment on artificial culture media have failed. However, subcutaneous inoculations of these materials to mice produced the typical lesions caused by M. lepraemurium.

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