Nitroblue tetrazolium (NBT)-dye tests were performed on blood specimens obtained at the time of admission or during the course of hospitalization from 498 patients and 150 healthy persons. In the healthy group, persons over 3 years of age showed mean percentages of 19.16±5.03 of NBT-positive neutrophils. On the other hand, infants from 3 months to 3 years of age showed decreased values of NBT-dye reduction. Patients suffering from acute bacterial or fungal infections, or cryoglobulinemia showed high percentages of NBT-positive neutrophils. Most patients with viral or tuberculous infections demonstrated normal or slightly attenuated values of NBT-dye reduction. Patients who had been receiving corticosteroids for long periods, those in severe stages of diabetes mellitus, and those receiving radiotherapy or anticancer drugs had remarkably impaired NBT-dye reduction. We examined the myeloperoxidase reactions, since we knew that patients with myeloperoxidase deficiencies have been reported to have disseminated candidiasis and to be defective in the ability of neutrophils to kill Candida albicans. We confirmed the fact that most persons had equal levels of myeloperoxidase and were unable to find anyone with a deficient myeloperoxidase syndrome among 450 individuals.

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Three hundred and thirty-eight strains of Escherichia coli from clinical sources were examined for their drug resistances and R factors in 1971. With reference to four drugs, i.e., tetracycline(TC), chloramphenicol(CM), streptomycin(SM) and sulfanilamide(SA), the SA-resistant strains were isolated most frequently followed by SM-, TC- and CM-resistant ones in decreasing order. Among those resistant to the aforementioned four drugs, 20.3, 19.0, 7.7 and 18.3% were single-, double-, triple- and quadruple-resistant, respectively. The R factors were demonstrated at a frequency of 29.2% among these resistant strains. From the strains possessing quadruple-resistance, the conjugationally transferable R factors were demonstrated most frequently (60.5%) followed by triple- (59.1%), double(21.7%) and single-resistant strains (4.5%). The isolation frequencies of strains resistant to kanamycin (KM) and ampicillin (APC) were 1.5 and 4.7%, respectively; being fewer in numbers compared with those resistant to the four drugs. Strains resistant to cephaloridine, nalidixic acid, colistin. gentamicin C complex and furatridine were not isolated. Strains resistant to KM and APC were demonstrated mostly from those possessing multiple resistance with reference to the aforementioned four drugs. Among the strains carrying either APC- or KM-resistance, 76.9 and 100% respectively, were found to carry R factors, indicating that most of the APC- and KM-resistances were conferred by the R factors. Twenty percent of the R+ strains with reference to resistance to the four drugs carried two types of R factors in a cell, being in a hetero-R state, and 66.7% of the strains carrying R factors with APC- or KM-resistance were in a hetero-R state.

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Infection of a subline of L cells adapted to grow in suspension (Ls) with Fushimi strain of HVJ (HVJ-F) resulted in a virus carrier state. Ls cells, when cultured in monolayer, showed morphological changes following infection of HVJ-F and were detached from the glass wall. However, when the detached cells were transferred to a new environment of suspension culture within 5 days after infection, the carrier state was again established. HVJ-F caused only lethal infection in L cells maintained exclusively in monolayer (Lm). On the other hand, both Ls and Lm, irrespective of their culture conditions, were lethally infected by Nagoya 1-60 strain of HVJ. The overall results showed that culture condition as well as the kind of host cells or virus strains is an important factor regulating the establishment and maintenance of the virus carrier state.

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The antibody response of mice to bovine γ-globulin(BGG) was suppressed either specifically by an intravenous injection of deaggregated soluble BGG (sBGG) or nonspecifically by X-irradiation. Immunization with the subcutaneous injection of BGG in Freund's incomplete adjuvant was given to mice either various days after sBGG injection or immediately after X-irradiation. Antigen-elimination (AE) test and passive hemagglutination(PHA) test were employed for estimating the immune status. The AE test indicated that the induction of tolerance was accomplished in the first 2 days after sBGG injection and that the tolerant state was stable at least for about 30 days thereafter. The degree of suppression by 1000 μg of sBGG corresponded to that obtained by X-irradiation at the dose of 400 R or more, and 100 μg of sBGG was equivalent to 300 R X-irradiation. The PHA test indicated, however, that such a correspondence as mentioned above between the dose of tolerogen and that of X-irradiation was not so stable as was seen by the AE test. Thus, the PHA titers of tolerized animals tended to recover up to the level of untolerized animals during the period of time from 10 days to 20 days after the tolerogen injection. Such discrepancies between the features in the AE test and those in the PHA test seemed attributable to a low avidity antibody formation in the tolerized animals, as judged by the hemagglutination-dissociation test. Hemagglutination by means of the sera from tolerized animals was seen to be reversed by the addition of free antigen more easily than the hemagglutination achieved by the sera of control animals or X-irradiated animals. The relationship between PHA titers and AE capacities of antibodies was investigated by the passive immunization of normal mice previously given the antigen. The result showed that the PHA titer did not always correlate with the AE capacity.

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Mice were made tolerant to bovine γ-globulin (BGG) by an intravenous injection of deaggregated soluble BGG (sBGG). Two weeks later, they were injected with immunogenic forms of BGG either into foot pads or intravenously, Time course of the number of antibody-forming cells and the relative avidity of plaque-forming antibodies in tolerized mice were compared with those of sublethally X-irradiated mice and of normal immunized mice. The number of plaque-forming cells (PFC) per popliteal lymph node of tolerant or X-irradiated mice was shown to be smaller than in the controls. The growth rate of PFC in the nodes in the early logarithmic phase was almost the same between tolerized mice and normal mice, but X-irradiation resulted in a reduction of the growth rate. Although X-irradiation did not influence the average avidity of plaque-forming antibodies, the induction of tolerance by sBGG slightly reduced the avidity in popliteal lymph nodes and conspicuously in spleens about three weeks after the challenge immunization, the relative avidity in the latter being as low as one tenth of the normal immunized mice. In popliteal lymph nodes of tolerized mice at day 9 after immunization, a reduction in the relative avidity of plaque-forming antibodies was not observed, though the number of PFC was as small as one fifth of the controls. However, X-irradiation of the hind limbs of tolerized mice prior to immunization caused a reduction in the avidity of plaque-forming antibodies in the nodes. These results indicated that the precursors of high avidity antibody producers were liable to tolerance induction, and that cells in the recirculating pool were inactivated by the tolerogen more easily than those in such remote sites as the popliteal lymph nodes.

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In 1970, a great outbreak of hand-foot-and-mouth disease (HFMD) due to Coxsackie A16 virus (Cox A16) occurred throughout Japan. When serologic relationship between the viruses isolated during the epidemic and the prototype strain of the serotype was examined by the tube neutralization test, the crude new isolates were found to be poorly neutralized by both an antiserum against the prototype strain and those against the isolates, although they were neutralized significantly in the plaque reduction test. However, about 2% of virus in crude suspensions of the isolates remained unneutralized even in the plaque reduction test and this fraction could be eliminated by filtering the virus materials through a 100 mμ Millipore filter. Therefore, the difficulty in neutralization of the isolates by the tube method could be accounted for by the presence of aggregated viruses. Even when filtered viruses were used, the reciprocal neutralization kinetic studies revealed a significant serological difference between the isolates and the prototype strain. Such serological properties of the isolates were not influenced by the cell types used for virus isolation or passages. All the results suggest that the Cox A16 isolates from the epidemic of HFMD in Japan, 1970, are serologically different from the prototype strain.

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Complement-fixing (CF) and complement fixation-inhibiting (CFI) antibodies were investigated sequentially in three horses infected with equine infectious anemia (EIA) virus. The CF activity was first demonstrated 6 or 8 days after onset of the first pyrexia. The CFI activity developed a short period later, and caused a decrease of apparent CF titer of the whole serum. However, both antibodies tended to increase with advance of the disease course in two persistently infected horses, whereas they became completely undetectable during the late-stage in the remaining horse which showed no evidence for recurrence of pyrexia or persistence of viremia. The CF activities determined with varying dilutions of serum were distinctly different in pattern between the early-stage serum having the CF activity alone and the late-stage serum having both the CF and the CFI activities. The CF antibody was precipitated by 27-30% saturation with ammonium sulfate (SAS) while the CFI activity distributed in a wider range of precipitates formed by 26-32% SAS. The CFI activity was demonstrated to a higher titer when a relatively small amount of antigen was sensitized with CFI antibody prior to addition of reference CF antibody than when the three reagents were mixed at one time. The late-stage serum with a strong CFI activity against EIA antigen had no effect on the CF reaction of other viral antigenantibody systems such as equine influenza and equine rhinopneumonitis.

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Millipore-filtered herpes virus and a hyperimmune rabbit serum were reacted to analyze the unneutralizable persistent fraction (PF). When the PF was serially diluted with antibody-containing diluent, the plaque-formers reduced in number. When the serial dilutions were made with antibodyfree diluent, plaque numbers were disproportionately smaller in lower dilutions. The PF filtered through a 0.22μ Millipore membrane showed only a slight loss of infectivity, but a further incubation of the filtrate at 37 C resulted in a marked reduction of titer. This effect was less pronounced when the membrane porosity was larger. Additional virus given to the PF was quickly neutralized by excess antibody. On the other hand, dilution of the virus-serum mixture followed by incubation at 37 C or sonication did not further reactivate the virus. When neutral complexes were sedimented by ultracentrifugation and resuspended in antibody-free diluent, a partial reactivation slightly exceeding the usual PF level occurred with a concomitant release of antibody. It is proposed that the PF may be free virus resulting from reversible virus-antibody reaction.

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