Analysis of linkage and segregation of unselected markers was made using a recombination system of Mycobacterium smegmatis strains, Jucho and Lacticola. It was demonstrated that recombinants primarily possessed the genetic characters derived from Jucho. This suggested that strains Lacticola and Jucho did not play equivalent roles in zygote formation, and that zygotes were made up essentially of the chromosome from Jucho and contained only a segment of the chromosome from Lacticola. The ordering of the genes in strain Lacticola might be as follows: φrb, gly-2⁺, leu-7⁺, and str-9. Results of backcrosses of recombinants with parental strains showed that there were four different mating types in the recombinants. This indicated that at least two different factors controlled mating in mycobacteria. D4 phage and phage Rabinowitschi did not have mating type specificities.

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The action of cephalexin against Staphylococcus aureus No. 50774 and Escherichia coli NIH had been studied by phase contrast microscopy and electron microscopy. S. aureus grown on the cephalexin-containing agar exhibited abnormal cell division when observed nuder the phase contrast microscope. Exposure of E. coli to cephalexin resulted in the formation of long fiaments and spheroplasts The degree of morphological changes was found to be dependent on the concentration of cephalexin added. Electron micrescopic studies revealed that S. aureus upon exposure to cephalexin produced a swelling of the cross wall, which resulted in lysis of the cell or the production of a protoplast. E. coli exposed to cephalexin produced long filamentous forms. A number of mesosomes were seen in the firamentous forms. However, the presence of a cross wall or plasma membrane partitioning the elongated cell thus formed was never observed. The above-mentioned lindings offer morphological evidence that the antibacterial action of cephalexin is ascribed to the inhibition of cell division and possibly cell wall svnthesis.

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Virus labeled with ³H-uridine or ³²P-orthophosphate was purified by CsCl equilibrium centrifugation of concentrated virus materials from infectious BHK21-WI2 cell culture fluids. A single clearly visible band formed in the gradient coinciding with a sharp peak of radioactivity having a buoyant density of 1.19g/ml. Infectisity exhibited a broader distribution with a peak coinciding kith the risible band, in which numerous virions of the sirus were observed with the aid of an electron microscope using the phosphotungstic negativc staining technique. Ribonucleic acid (RNA) extracted from the purified virus by the phenol method exhibited a rather broad distribution of radioactivity with a major peak at about 12S when analyzed by the sucrose density gradient centrifugation technique. Viral RNA centrifuged in the gradient after ribonuclcase (RNase) treatment showed a single sharp peak at about 12S These findings seemed to indicate that the virion of this virus contained double-stranded RNA. Double strandeclness of the viral RNA was further corroborated by an examination of the base composition, reduced resistance to RNase in low salt concentration and a sharp thermal transition with a relatively high melting temperature of 85C. The virus could not be classified either in the rhabdovirus group, although the shape of the vision resembled that of the rhabdoviruses, or in the reosirus group since the virus was ether-sensitive. It seemed necessars to create a new genus for this virus.

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The Mivadera strain of Newcastle discase virus (NDV) consisted predominantly of virus particles forming small plaques on monolayers of chick embryo fibroblasts (CEF) and contained small amounts of virus particles forming large plaques These large and small-plaque-forming clones of this virus (NDV-L and NDV-S) were isolated. The small size of the NDV-S plaques did not appear to be due to an agar inhibitor. NDV-L produced a much higher vield of intectice virus particles in CEF and they were released more completely from the infected cells than were those produced by NDV-S. The yield of infective vitus of NDV-L per cell from cultures of CEF was comparable to the yield from the allantoic cells. The infectivitv hemagglutinin ratio for NDV-L from CEF was as high as the ratio for virus from the allantoic cells, but the ratio for NDV-S from CEF was lower NDV-S demonstrated an autointerference phenomenon in CEF when infected at high multiplicities, but NDV-L did not. Contrary to virus multiplication, NDV-S exhibited a more rapid and marked cytopathic effect on monolayers of CEF than NDV-L In the allantoic cavity of eggs NDV-S produced slightly higher virus yields than NDV-L. No correlation existed between plaque size of the two viruses and the capacity to induce interferon synthesis or the susceptibility to the action of interferon. The properties of both distinctive plaque isolates were stable on egg passage.

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In a previous paper we reported that an inbred strain of SL mice and an outbred strain of CFI mice belonged to the high-responder strains in antibody production after primary immunization with hamster erythrocytes (H-RBC), while inbred strains of C57BL/6, AKR and C3H/He mice belonged to low-responder strains. In the present study we obtained the following results. 1) Pre-sensitization with hamster lymphoma cnhanced antibody production after an intravenous injection of H-RBC. There was no strain difference in the pattern of antibody production against H-RBC among pre-sensitized mice. 2) The pattern of enhanced antibody production after an intravenous injection of H-RBC into pre-sensitized mice assumed the primary type in terms of time of appearance of hemolysin plaque-forming cells (PEC) in the spleens and the conversion from 2-mercaptoethanol sensitive to 2-mercaptoethanol resistant antibody production, when the intervals between both treatments were within 7 days. 3) Pre-sensitization with lymphoma induced not only an increase in numbers of PFC after an intracenous injection of H-RBC, but also an increase in the size of the hemolysin plaques. These results suggested that sensitization with hamster lymphoma stimulated some kinds of immuno-competent cells, which could contribute to antibody production against H-RBC after a booster injection of H-RBC.

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A portion of the seed virus remained active even in the presence of excess antibody. With an early serum or relatively low concentrations of a hyperimmune serum, the presence of such unneutralizable virus was uncovered by the addition of complement. This unneutralizable persistent fraction (PF) was distinguished from sensitized virus surviving in insufficient antibody, because the latter was inactivated by high concentration of antibody as quickly as the control virus. The level of the PF was constant regardless of the serum species, serum lot, dilution of serum, antibody class and complement requirement of antibody, being approximately) 0.1% of the seed virus. Millipore filtration of the seed virus lowered this PF level to varying degrees depending upon the porosity However, when filtered virus-serum mixtures were incubated at 37C for a Sufficient time and then filtered again, a portion of the unneutralized virus did pass a 0.22μ membrane. Furthermore, virus sensitized with insufficient antibody showed no increase in resistance to complement after 10hr incubation at 4C, These results proved that viral aggregation was not the essential cause of the PF. It was confirmed, on the other hand, that the neutralization of virus by hyperimmune IgG followed a one-hit curve. Analysis of these data appeared to support Rappaport's postulation that the PF represented antibody-coated virus whose critical antigenic sites were all left unbound.

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An endotoxin was extracted from Salmonella typhosa, strain O-901, with trichloroacetic acid and phenol-water (TCA-PW ET). TCA-PW ET was treated with 0.5N lithium hydroxide at 37C for 30min. The hydrolysate was chromatographed in a Sephadex G-150 column. The absorbancy at 485mμ showed five successive peaks (Frs. 1 through 5). Analytical centrifugation indicated that these fractions were composed of diverse sizes. The sedimentation coefficient of Fr. 5 was the smallest being approximately 6.5S. The composition of each fraction was shown to be different by chemical analysis. Fraction 5 consisted mostly of hexosamine and fatty acids and some neutral sugars. With gas chromatography Fr. 5 showed a distinct difference when compared with either TCA-PW ET or Frs. 1 to 3 in its reduced lauric acid as well as increased β-hydroxymyristic acid contents. Rabbit pyrogenicity of Fr. 5 as compared with TCA-PW ET was approximately one thousandth, while chick embryo lethality was almost the same. Biological activities of the other fractions were intermediate in relative potencies. Fr. 5 was further chromatographed in Sephadex G-75 and G-50 columns and six fractions were obtained. All of these fractions were highly lethal with slight differences. Phosphorus, heptose, 2-keto-3-deoxyoctonate and glucosamine contents of the endotoxin products were relatively high, while the fatty acid contents were partly decreased. Relationship between lethality and chemical constituents of the endotoxin products is discussed.

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