bluuebunny/crossref_metadata_2025 huggingface dataset:
Evidence That the Multi‐Drug‐Resistance of Streptococcus faecalis BIO‐4R Is Not Carried by a Plasmid
Authors: Yamashita Yutaka , Ozawa Kyosuke , Takeda Yoshifumi , Miwatani Toshio
Published: November 2013

The multi‐drug‐resistant strain Streptococcus faecalis BIO‐4R was studied to see if it carried a plasmid responsible for antibiotic resistance. From results indicating that the antibiotic resistance of S. faecalis BIO‐4R was not transferred to recipient bacteria, that the organism did not produce enzymes which inactivated antibiotics, and that the presence of covalently closed circular DNA was not demonstrated by dye‐cesium chloride buoyant density gradient centrifugation, it was concluded that the organism did not carry such a plasmid. Studies on polyphenylalanine synthesis by cell‐free extracts of the organism showed that its ribosomes were resistant to chloramphenicol, tetracycline, streptomycin and kanamycin. These results, although rather indirectly, support the above notion that the multi‐drug‐resistance of S. faecalis BIO‐4R is not mediated by a plasmid.
Relation between Delayed Skin Reactivity and Macrophage Migration Inhibition or Lymphocyte Transformation in Tuberculin‐Type Hypersensitivity and Jones‐Mote Hypersensitivity
Authors: Himeno Kunisuke , Nomoto Kikuo , Kuroiwa Ataru , Miyazaki Sumio , Takeya Kenji
Published: November 2013

Precise time‐course studies on delayed skin reaction, lymphocyte transformation and macrophage migration inhibition were carried out from day 3 to 270 and from day 3 to 120, respectively, in guinea pigs immunized with bovine gamma‐globulin (BGG) in complete Freund's adjuvant (CFA) and those immunized with BGG in incomplete Freund's adjuvant (IFA). a) Delayed skin reactions could be elicited for a long period of time after immunization with BGG in CFA in the presence of prominent antibody production and were accompanied by induration. b) Delayed reactions could be elicited transiently after immunization with BGG in IFA and were not accompanied by induration. c) At the peak of hypersensitivity, infiltrating cells at the reaction sites were composed largely of mononuclear cells and basophils, respectively, in the animals immunized with BGG in CFA and those immunized with BGG in IFA. d) Uptake of 3H‐thymidine by lymphocytes was increased remarkably in the presence of BGG when cells were obtained at early stages after immunization by both methods. e) Macrophage migration inhibition was strongly positive in animals immunized with BGG in CFA but weakly positive in those immunized with BGG in IFA. Increased lymphocyte transformation preceded the appearance of a positive migration inhibition. f) After immunization with BGG in CFA, Jones‐Mote hypersensitivity appeared to precede the development of tuberculin‐type hypersensitivity.
The Characterization of Antibacterial Antibodies in Bovine Immune Sera to Staphylococcus aureus
Authors: Yokomizo Yuichi , Isayama Yasuro
Published: November 2013

Humoral antibody responses to the encapsulated Smith diffuse strain of Staphylococcus aureus were examined in cows immunized with the killed vaccine via different systemic routes. The sequential appearance of the antibody within different immunoglobulin classes in the sera during the course of immunization was followed by passive hemagglutination (PHA) and precipitation (PC) reactions and the mouse passive protection test. Repeated intravenous injections with the killed vaccine suspended in buffered saline stimulated production of IgM antibody exclusively during the whole period of immunization. On the contrary, following intramuscular administration with the vaccine incorporated in Freund's incomplete adjuvant, the antibodies appeared predominantly in IgG fractions of the sera.

Specific antibody to the homologous strain used for vaccination was prepared from bovine immune sera by an absorption and elution process. The mouse passive protective activity of the antibody preparation was removed by absorption with the capsular polysaccharide antigen as well as by the whole cell adsorbent of the Smith diffuse strain, but not by the Smith compact and Cowan I strains of S. aureus. IgM, IgG1 and IgG2 proteins were isolated from the purified antibody and were compared, on a weight basis, with respect to their biological activities. Slightly higher activity of the IgG over the IgM antibody was demonstrated both in the mouse passive protection test and PC reaction, whereas in the PHA reaction, IgM antibody was shown to possess a significantly higher activity than IgG antibody.
In Vitro Cytotoxicity of Peritoneal Macrophages Activated with Mycobacterium smegmatis
Authors: Yamamoto Saburo , Tokunaga Tohru
Published: November 2013

Incorporation of 3H‐TdR into EL4 leukemic cells in vitro was inhibited by peritoneal exudate cells (PEC) harvested from syngeneic C57BL/6J mice given an intraperitoneal (i.p.) injection of 1 × 107 viable Mycobacterium smegmatis ATCC 607 (Smeg) 4 days before. This phenomenon was also observed in the following five systems of PEC from animals and syngeneic tumor cells: C57BL/6J mice and B16 melanoma; DBA/2 mice and P815 mastocytoma; SWM/Ms mice and K5 fibrosarcoma; BALB/c, nu/nu mice and KKN‐1 fibrosarcoma; and strain 2 guinea pigs and line‐10 hepatoma. The in vitro cytotoxicity of the PEC activated by viable Smeg was much higher than those activated by dead‐Smeg, viable BCG or proteose peptone. The activity of the adherent fraction of the PEC was stronger than that of the nonadherent one, and not influenced by either anti‐θ or anti‐mouse lymphocyte rabbit sera. The PEC induced with Smeg 4 days before contained a large population of mononuclear cells (88.9%) and a significant level of polymorphonuclear cells (PMN) (3.2%), and showed a much higher cytotoxicity than the PEC induced with Smeg 3 hr before, which contained a much larger population of PMN (71.9%), suggesting that PMN were not the effector cells in this system. In vitro and in vivo treatment with macrophage‐inhibitors such as carrageenan, trypan blue and cytochalacin B, reduced the activity of the PEC. All of these facts suggested macrophages as the effector. Viable macrophages were required for the growth inhibition of EL4 in vitro: gamma‐ray irradiated or freeze‐thawed macrophages were ineffective. Kinetic studies revealed that inhibition of 3H‐TdR incorporation into EL4 cells started within 3 hr of incubation together with the activated macrophages at an effector to target (E/T) ratio of 5, and the incorporation decreased gradually with the lapse of incubation time. On the other hand, 51Cr release from labelled EL4 was undetected when the E/T ratio was 5 but detected at on E/T of 10 or more. Even at the higher E/T ratio, at least 10 hr were needed until the release of 51Cr, suggesting that the activated macrophages produced growth inhibition of tumor cells followed by cell destruction.
Cell Wall Autolysis in Log Phase Cells of Micrococcus lysodeikticus (luteus)*
Authors: Monodane Toshihiko , Matsushima Yoshio , Hirachi Yoshiyuki , Kotani Shozo
Published: November 2013

Log phase cells of Micrococcus lysodeikticus (luteus) IFO 3333 autolyzed when incubated at 37 C in 0.01 m sodium‐phosphate buffer pH 7.5. The enzyme involved in the autolysis was recovered mainly in an aqueous phase from cytoplasmic membranes and cytoplasmic materials treated with n‐butanol, and proved to be an N‐acetylmuramyl‐L‐alanine amidase. The autolysis of log phase cells suspended in autolyzing buffer was depressed by the addition of trypsin to the buffer.
Demonstration of the Physiological Role of Autolysis by a Comparative Study with a Wild‐Type and its Non‐Autolytic Mutant of Micrococcus lysodeikticus (luteus) Cultivated with Externally Added Proteolytic Enzymes*
Authors: Monodane Toshihiko , Matsushima Yoshio , Kotani Shozo
Published: November 2013

The log phase cells of autolytic Micrococcus lysodeikticus (luteus) IFO 3333 did not autolyze when grown in the presence of trypsin although the growth curve and morphology of the cells were not influenced.

A non‐autolytic mutant was obtained by subculture of the wild‐type strain IFO 3333 on an agar slant containing 1% glucose. The mutant (strain MT) was arranged in tetrads or in clusters consisting of regular tetrads, in contrast to the wild‐type IFO 3333 which occurred singly or in irregular masses. The mutant MT grown in a culture medium containing trypsin caused remarkable alteration in cell morphology: large cell packets consisting of a number of “unit tetrads” arranged regularly in three dimensions were formed by the addition of trypsin to the medium. The findings suggest that inhibition of the separation of divided cells is brought about by inactivation or suppression of a cell wall autolytic enzyme which plays an important role in the separation step and is accessible to externally added trypsin in the mutant cells but not in the wild‐type cells.

The possibility that there are two kinds or phases of autolytic enzymes, “a physiological autolytic enzyme” and “a useless autolytic enzyme”, is discussed.
A New Method for Extraction of Extravasated Dye in the Skin and the Influence of Fasting Stress on Passive Cutaneous Anaphylaxis in Guinea Pigs and Rats*
Authors: Katayama Satoshi , Shionoya Hiroshi , Ohtake Shinzaburo
Published: November 2013

A method for quantitative extraction of extravasated dye from the skin was studied in guinea pigs and rats. A simple method with a low cost and good recovery was established as follows; A piece of the skin containing extravasated dye was soaked overnight in a stoppered glass tube containing 1 ml of 1 n KOH at 37 C. Then, 9 ml of a mixed solution of 0.6 n H3PO4 and acetone (5:13) was added to the tube. The tube was shaken vigorously for a few seconds and centrifuged at 3,000 rpm for 15 min. Absorbance of supernatant was measured at 620 nm. The recovery rate of the dye was about 95% both in guinea pigs and rats.

Using this method we observed that fasting stress significantly reduced the intensity of skin reactions induced by chemical mediators, heterologous PCA and especially homologous PCA in guinea pigs.
Improvement of the Micromethod for the Limulus Lysate Test
Authors: Okuguchi Shuji
Published: November 2013

Frauch's micro‐slide method was improved to facilitate the endpoint‐determination of the Limulus test. Two precise observations, by inverted phase contrast microscopy and with a staining procedure, were newly performed as additions to the slide test. The staining procedure was proposed as an improved method for the Limulus test since it is simple and convenient. In the staining method, bromophenol blue (BPB) solution was used as the staining solution. A negative (–), a strong positive (++) and a weak positive reaction (+) were characterized by a “ring” formation, a “cloud‐like” spread of gel and a “spot” in the “cloud,” respectively. Since the distinction between (–) and (+) reactions was obvious in the proposed method, determination of the endpoint was easier than in the ordinary tube and Frauch's method. The sensitivity of the present method was equal to or higher than that of other methods. Inverted phase contrast microscopy was utilized to confirm the findings obtained by the staining method. The volume of the lysate used in this method was as little as 1/10 of that used in the tube method.
Inhibition by Ammonium Ion of Germination of Unactivated Spores of Bacillus cereus T Induced by L‐Alanine and Inosine
Authors: Shibata Hirofumi , Takamatsu Hiroaki , Tani Isamu
Published: November 2013

Studies were carried out on the inhibitory effect of NH4+ on germination of spores of Bacillus cereus T induced by L‐alanine and inosine. Kinetic analysis showed that NH4+ inhibited the germination competitively. Its inhibitory effect was greater when the unactivated spores had been preincubated with L‐alanine. NH4+ did not inhibit the response of unactivated spores to L‐alanine during preincubation. These results suggest that L‐alanine sensitizes the spores to the inhibitory effect of NH4+.
Kinetics and Mechanisms of Macrophage Activation by Corynebacterium anaerobium
Authors: Hanazawa Shigemasa , Kato Hideo , Yamaura Koichi , Yamaguchi Yasuo
Published: November 2013

Kinetics and mechanisms of macrophage activation by heat‐killed Corynebacterium anaerobium (CA) in mice were investigated. The carbon clearance test revealed that the function of the reticuloendothelial system rose markedly on the 4th day after a single intravenous injection of CA and continued in a highly enhanced state until the 14th day. This activity declined gradually and dropped to a normal level around the 21st day. On the other hand, both lysosomal enzymes, β‐glucuronidase and acid phosphatase, of peritoneal macrophages decreased after the CA injection and then recovered, taking an almost inverse course to the function of the reticuloendothelial system. These results might be attributable to possible extracellular secretion of the lysosomal enzymes in accordance with macrophage activation by CA. A remarkable cytotoxicity of peritoneal macrophages, examined in vitro against L 929 cells, was detected on the 4th day following intraperitoneal administration of CA. It was maintained up to the 14th day and then declined rapidly. The mechanisms of macrophage activation by CA were also examined in vitro. CA‐homogenate, heat‐killed CA disrupted with an ultrasonicator, directly activated thioglycollate‐induced macrophages. The macrophages were also activated by simultaneous treatment with both CA‐homogenate and CA‐sensitized spleen cells. Furthermore, the supernatant obtained from the culture of CA‐sensitized spleen cells with CA‐homogenate was capable of inducing activation of the macrophages. Conversely, the culture supernatant of spleen cells from CA‐immunized athymic nude mice with CA‐homogenate was unable to activate them. It was ascertained from the above‐mentioned results that macrophages are activated initially by direct action of CA in a nonspecific way and subsequently by a soluble factor elaborated by CA‐sensitized lymphocytes in an immunological way.
Properties of the Erythrocyte Receptors for Influenza C Virus
Authors: Ohuchi Masanobu , Homma Morio , Muramatsu Michiko , Ohyama Shinobu
Published: November 2013

Properties of the receptor for influenza C virus were studied. Although the receptor for influenza C virus on chicken erythrocytes was destroyed by the homologous virion, neuraminidase activity could not be detected in any of the influenza C virus strains tested. The receptor activity of chicken erythrocytes for influenza C virus was diminished by formaldehyde treatment but not by periodate oxidation. There was a considerable variation in the pattern and the titer of hemagglutination of influenza C virus when human erythrocytes of different blood types were used; the virus agglutinated most type B erythrocytes but not type A erythrocytes. By using human type B erythrocytes, differences among strains of influenza C virus in the hemagglutinating activity were also demonstrated. These results showed that both the receptor for and the receptor‐destroying activity of influenza C virus were completely different from those of influenza A or B virus and also that carbohydrates were not involved in the receptor for influenza C virus.
Proliferation of Hemolysin‐Plaque‐Forming Cells in Nude Mice and Normal Littermates Subjected to Antigenic Competition
Authors: Taniguchi Toru , Nomoto Kikuo , Miake Shunji , Takeya Kenji
Published: November 2013

The increase of PFC per spleen and the development of hemolytic foci were examined to clarify the patterns of clonal expansion of B‐lymphocytes in athymic nude mice (nu/nu) and normal littermates (nu/+) subjected to the procedure for antigenic competition between horse erythrocytes (HRBC) and sheep erythrocytes (SRBC). In normal littermates without pretreatment with HRBC, a small number of PFC and hemolytic foci of small size were detected 2 days after the challenge with SRBC. The number of PFC increased progressively from day 2 to day 4, and hemolytic foci increased in the number and size during the period. In nude mice, a small number of PFC were detected on day 2 and the number increased only slightly from day 2 to day 4. No large hemolytic foci were detected during the period. In normal littermates subjected to the procedure for antigenic competition, the patterns of increase of PFC and development of hemolytic foci were similar to those in nude mice. In nude mice, the procedure for antigenic competition exerted almost no effect on the patterns.
Studies on Delayed Hypersensitivity to Protein Antigen
Authors: Nakashima Shoichi , Sakai Yasuo , Take Nobuaki , Umeda Yukio
Published: November 2013

It was shown in our previous paper that mice primed with chemically modified bacterial α‐amylase (BαA), which was neither cross‐reactive with anti‐BαA antibody nor able to induce a humoral anti‐BαA response, developed enhanced responses to a subsequent challenge with native BαA and that the magnitude of the immunological memory was closely related to the priming dose of modified BαA. This paper describes the experimental conditions for induction of delayed hypersensitivity (DH) by modified BαA in relation to the development of immunological memory for antibody response to native BαA.

Mice primed with either an intraperitoneal (i.p.) or subcutaneous (s.c.) injection of modified BαA in complete Freund's adjuvant (CFA) developed enhanced anti‐BαA responses to a subsequent challenge with BαA. In contrast, when mice were immunized with an s.c. injection of the modified BαA only, a significant level of DH to native BαA could be induced, as measured by the footpad reaction (FPR). The highest degree of DH was observed in mice given 50 μg of modified BαA. DH was detectable within 5 days and persisted for 25 days after immunization. In the reciprocal combination of native BαA as the immunogen and modified BαA as the eliciting antigen, the relationship of anti‐BαA responses to DH was examined. The primary anti‐BαA responses induced by an i.p. injection of large doses of BαA was markedly higher than those induced by an s.c. injection, while DH was exhibited only in mice given an s.c. injection of BαA in CFA.
Existence of 28R‐Antigen in a Certain Strain of Group F Streptococcus
Authors: Maekawa Shizue , Yaegashi Tazuko , Nakayama Yasuhiko
Published: November 2013

An inter‐Group common antigen was detected between Group A type 28 (Small)‐ and Group F (21/58/O'Mahoney, Colindale)‐streptococcal cells by the T‐typing agglutination reaction. The characteristics of this antigen coincide with those of the 28R‐antigen, which was first detected in the Group A type 28 (Small) cells by Lancefield in 1943, in the following points: 1) It can beextract ed from the cells with HCl at pH 2.0 at 100 C in a stable state; 2) It can be kept in a stable state by heating in an alkaline solution at pH 7.8; 3) The antigen on the heat‐killed cells was not affected by trypsin digestion at pH 7.8 but was destroyed by pepsin digestion at pH 2.0.
Studies on the Lipid Components of Endotoxin II. Chemical Analyses and Biological Properties of Neutral, Polar‐I and Polar‐II Subfractions
Authors: Konno Seishi
Published: November 2013

Lipid components obtained from Salmonella typhosa O‐901 endotoxin by acid hydrolysis were separated into neutral, polar‐I and polar‐II lipid fractions by silica gel column chromatography. These lipids were further separated by silica gel column and/or thin‐layer chromatography. The subfractions were analyzed by thin‐layer chromatography, gas chromatography and infrared spectrophotometry. Seven subfractions obtained from the neutral lipid fraction contained lauric, myristic, palmitic, 3‐OH‐myristic acid, artificial products of 3‐OH‐myristic acid, or a small amount of two unidentified fatty acids. These fatty acids and glucosamine were commonly detected in six subfractions obtained from the polar‐I lipid fraction. Fatty acids, glucosamine, and O‐phosphorylethanolamine were detected in all of the 13 subfractions obtained from the polar‐II lipid fraction. Chick embryo lethal activity, rabbit pyrogenicity and in vitro interferon inducing activity were found in three polar‐I lipid subfractions and five polar‐II lipid subfractions, but not in neutral lipids. The activities were highest in a polar‐II lipid subfraction, which contained smaller amounts of O‐phosphorylethanolamine and glucosamine than the other subfractions. However, no particular chemical constituent (s) related to the biological activities could be found. Prolonged acid hydrolysis of the polar‐II lipids gave rise to neutral and polar‐I lipids. Chemical and biological aspects of the lipid constituents of endotoxin are discussed.
Effects of Protamine Sulfate on Dengue Type 1 Viral Activities
Authors: Fujita Nobuya , Karabatsos Nick , Shope Robert E.
Published: November 2013

Protamine treatment of type 1 dengue‐infected mouse brain suspension resulted in precipitation of several viral specific activities. Complement‐fixation activity was almost completely precipitated by protamine. The complement‐fixation components recovered in the precipitate were comparable to a non‐precipitated reference dengue 1 antigen in their homologous and heterologous reactions. Dengue hemagglutinin was also precipitated by the same treatment. The precipitated hemagglutinin was resolved into three components by buoyant density centrifugation, whose densities were 1.236, 1.215, and 1.178 g/ml, respectively. Three similar HA components were detected in non‐protamine treated virus preparations. In both instances the highest‐titered HA fraction possessed a buoyant density of 1.21–1.22 g/ml. These HA components were tested in the hemagglutination‐inhibition reaction and were proved to be virus‐specific. Cesium chloride density gradient centrifugation was shown to be useful for removing possible inhibitor(s) of viral specific hemagglutinin.
Plasmid‐Mediated Sulfanilamide Resistance
Authors: Nagate Takatoshi , Inoue Matsuhisa , Inoue Kunio , Mitsuhashi Susumu
Published: November 2013

Dihydropteroate synthetase (DHPS) is specified by a substrain of Escherichia coli K12, ML1410. This enzyme activity is inhibited by sulfanilamides (Sa) and is known to be heat‐stable, i.e., an Sa‐sensitive normal enzyme. Another DHPS activity specified by E. coli ML1410 carrying drug resistance plasmids is Sa‐resistant but heat‐sensitive, i.e., an Sa‐resistant enzyme. Most plasmids encoding single Sa or double (Sa. Tc or Sa. Sm) (Tc, tetracycline; Sm, streptomycin) resistance mediate the formation of this type of DHPS. Therefore, E. coli carrying these plasmids becomes diploid for DHPS, i.e., an Sa‐resistant and an Sa‐sensitive normal enzyme. The biochemical mechanism of Sa resistance mediated by plasmids encoding triple (Cm.Sm.Sa; Tc.Sm.Sa) and quadruple (Cm.Tc.Sm.Sa) resistance is not due to the formation of an altered DHPS but probably due to the decrease in permeation of the drug into the cell. The evolutionary process of the formation of Sa‐resistance determinants on plasmids is discussed based on the presence of two types of Sa resistance mechanism.
Physico‐Chemical Properties of Mouse Hepatitis Virus (MHV‐2) Grown on DBT Cell Culture
Authors: Hirano Norio , Hino Shigeo , Fujiwara Kosaku
Published: November 2013

Some properties of a strain of mouse hepatitis virus, MHV‐2, grown on DBT cells were determined using a plaque assay on the cells. Viral growth was not inhibited by the presence of actinomycin D or 5‐iodo‐2‐deoxyuridine. MHV‐2 was completely inactivated by ether, chloroform, sodium deoxycholate or beta‐propiolactone, but showed a moderate resistance to trypsin. Heating at 56 C for 30 min did not completely abolish the virus infectivity. The virus was stable after heating at 50 C for 15 min in 1m‐MgCl2 or 1m‐MgSO4 as well as at 37 C for 60 min at pH 3.0 to 9.0. Infectivity was decreased to 1/100 and 1/400 after storing at 4 C for 30 days and 37 C for 24 hr, respectively. The virus passed through a 200‐nm but not a 50‐nm Sartorius membrane filter. The buoyant density of MHV‐2 was 1.183 g/cm3 in sucrose gradient, and the fraction contained coronavirus‐like particles measuring 70 to 130 nm in diameter. Survival rate was 10% after exposure to ultraviolet at 150 ergs/mm2. Freezing and thawing or sonication at 20 kc for 3 min did not affect the virus titer. No hemagglutinin was demonstrable with red blood cells of the chicken, Japanese quail, mouse, rat, hamster, guinea pig, sheep, bovine or human.
A New Microplate Neutralization Test for Typing of Herpes Simplex Virus
Authors: Tada Aiko , Yoshino Kamesaburo
Published: November 2013

A microplate serum neutralization test for estimation of complement‐requiring neutralizing (CRN) antibody was established as the first step for simplification of typing of herpes simplex virus (HSV). When guinea pigs were immunized with type 2 HSV, the late sera could mostly differentiate the types of HSV better than hyperimmune rabbit sera, the CRN titer against the heterologous type 1 HSV being much lower than the homologous titer. Sera of guinea pigs immunized with type 1 HSV showed about the same level of cross reaction against type 2 HSV as did rabbit antisera. Guinea pig sera having minimal levels of cross reaction were selected, and their high dilution (1:160) and complement were added to serial 10‐fold dilutions of virus in the microplate titration of virus infectivity. Selective reduction of virus titer by either antiserum could determine the type of HSV. No equivocal intermediate case was found among a number of stock strains including many fresh isolates. The typing result coincided with that determined by a modification of Yang et al's method based on virus titers obtained with Vero and primary chick embryo cells. The typing based on plaquing in chick embryo cells sometimes failed to identify type 1 HSV.
A Sensitive Solid Phase Radioimmunoassay for Secretory IgA
Authors: Shibata Rumiko , Onoue Kaoru , Goya Nagahide
Published: November 2013

A sensitive solid phase radioimmunoassay method was established for the specific quantitative determination of secretory IgA (sIgA) by taking advantage of the dual antigenicities of sIgA, one specific for α‐chain and the other for secretory component (SC). The sIgA and IgA in the sample were first bound by anti‐IgA antibodies coated on the polystyrene tube, then the amount of bound sIgA was quantified by the use of 125I‐labeled anti‐SC antibodies. This method is quite sensitive and allows us to distinguish sIgA from IgA and free SC which usually coexist in exocrine secretions. Linear relationship was observed between the bound radioactivity of radioiodinated anti‐SC and the amount of sIgA in the range of 5 to 60 ng of sIgA.

With this method, the urinary sIgA levels in normal children and adults were measured. Urinary sIgA was detected in half of infants within 7 days after birth, while it could be demonstrated in almost all of the infants by 14th day. Then, it gradually increased and reached about a half of the adult level by the age of puberty.

This method will be useful for the sensitive and specific measurement of sIgA in various exocrine secretions.
Serological Studies on the Major Histocompatibility Complex of New Inbred Strains of the Guinea Pig1
Authors: Chiba Joe , Otokawa Minoru , Nakagawa Masao , Egashira Yasuyuki
Published: November 2013

New inbred strains of guinea pigs, JY 1, JY 2, JY 3, JY 6, JY 9 and JY 10 have been established in this Institute. Serologic studies of guinea pig leukocyte antigens (GPLA antigens) were carried out in order to examine their major histocompatibility complex (GPLA complex). Antisera specific for Ia antigens were raised by cross‐immunization of NIH strain 2 (NIH 2) and NIH strain 13 (NIH 13) guinea pigs, well known inbred guinea pigs. The sera identified four distinct Ia specificities, which were designated as Ia.2a, Ia.2b, Ia.13a and Ia.13b. Six antigenic specificities different from the above Ia specificities were identified by sera obtained by appropriate immunization of the inbred guinea pigs and were designated as P.1, P.2, P.3, P.4, P.5 and P.6. Antigenic specificities of GPLA antigens recognized in inbred guinea pigs were: NIH 2 (Ia.2a, Ia.2b, P.1, P.2, P.3, P.4), NIH 13 (Ia.13a, Ia.13b, P.1, P.2, P.3, P.4), JY 1 (Ia.13a, Ia.13b, P.5), JY 2 (Ia.2b, Ia.13b, P.3, P.4, P.6), JY 3 (Ia.13a, Ia.13b, P.2, P.4, P.5), JY 6 (Ia.2b, Ia.13b, P.3, P.6), JY 9 (Ia.13a, Ia.13b, P.4, P.5), JY 10 (Ia.13a, Ia.13b, P.2, P.3, P.4, P.6). The correspondence of these specificities to those already reported was discussed and the P.2 or P.4 was considered to be an additional specificity of GPLA antigens that have not been reported yet. As the new inbred strains of guinea pigs were thus found to possess characteristic GPLA complex, which differ from each other and from those of NIH 2 and NIH 13 strain, they should be useful for studies of roles of the major histocompatibility complex in the immune system.
Concanavalin A‐Mediated Agglutination of 3T3 Cells After Exposure to UV‐Irradiated Herpes Simplex Virus
Authors: Taguchi Fumiaki , Hara Kazuya , Nagaki Daizo
Published: November 2013

Studies were made to determine the effect of UV‐irradiation of herpes simplex virus (HSV) on Concanavalin A (Con‐A)‐mediated agglutination of 3T3 cells. There were three different phases of agglutination by Con‐A of cells infected with HSV. The agglutinability began to increase from 3 or 4 hr, or 72 hr after exposure of cells to HSV. The early‐appearing agglutinability was further divided into two phases, based on its sensitivity to metabolic inhibitors. These were tentatively called “Early 1 or inhibitor sensitive”, “Early 2 or inhibitor insensitive” and “Late” agglutinability.

“Early 1” agglutination, detected from 3 hr post infection (pi), was induced by treating cells with HSV, either active or UV‐irradiated for less than 5 min and was inhibited when actinomycin D (1 μg/ml) or cycloheximide (50 μg/ml) was added to the cultures. “Early 2” agglutination began to increase from 4 hr pi when cells were inoculated with HSV irradiated for 7 to 20 min and was not affected by either inhibitor. HSV irradiated for 6 min failed to induce either agglutinability. “Late” agglutination, observed 72 hr pi, was detected in cultures which had been treated with HSV irradiated for 4 to 15 min. Among those, virus irradiated for 6 to 8 min was most efficient. HSV‐transformed cells were also agglutinated without exception by low concentrations of Con‐A.
Separation of RRBC‐RFC and Non‐Rosette Forming Cells (non‐RFC) of Guinea Pig T‐Cell Populations and Their Functional Difference
Authors: Yokomuro Kozo , Mabuchi Ayako , Kimura Yoshitami
Published: November 2013

Guinea pig lymph node cells suspension (LNC‐O) was filtered through a glass wool column and the effluent (LNC‐G) was further filtered through a nylon column. In this effluent (LNC‐NE) about 30 per cent of the lymphocytes was identified as non‐rosette forming cells (non‐RFC). The non‐RFC fraction was separated from LNC‐NE fraction by Ficol‐Conray specific gravity centrifugation of effluent cells reacted previously to rabbit red blood cells (RRBC). The upper layer after centrifugation, designated non‐RFC fraction, was separated. In this fraction 96% of the cells were lymphocytes and about 95% of them were non‐RFC, which lacked receptors for rabbit red blood cells (RRBC) or EAC and detectable surface immunoglobulin by conventional techniques.

Though the response of the lymphocytes in the non‐RFC fraction to mitogenic (Con‐A, LPS) or antigenic stimulation was lower in comparison with that in RFC‐rich fraction, the response of non‐RFC to ConA exceeded the response to LPS.

These facts suggest that at least a portion of the non‐RFC may be cells from the T‐cell line.
The Mode of Production of Endotoxin‐Induced Interferon in Rabbit Tissue Cells
Authors: Hashimoto Hiroshi , Shibukawa Nobuyuki , Kojima Yasuhiko
Published: November 2013

In vitro production of endotoxin‐induced interferon in rabbit tissue cell cultures could be enhanced by pretreatment with interferon. The enhancible state developed from the first hr of incubation at 37 C and a maximal priming effect was attained at 6 hr of incubation. Yields of interferon from unprimed cultures were usually 20–200 units/ml. In contrast, the primed cultures constantly yielded 1,000–2,500 units/ml of interferon. The pretreatment with interferon seemed to cause an earlier appearance of detectable interferon and the primed cells became more sensitive to endotoxin.

It turned out that 10–30 units/ml of rabbit interferon were enough to develop the maximal priming. Even when cells were pretreated with higher doses of rabbit interferon such as 1.0 × 104–1.0 × 105 units/ml, the same level of priming effect was always observed without diminution. Various types of homologous (rabbit) and heterologous (human and mouse) interferon preparations showed similar dose‐dependent enhancement of interferon production in proportion to the antiviral titers of these preparations as tested with RK‐13 cells of rabbit origin.
R Plasmids with Thermosensitive Transferability in Salmonella Strains Isolated from Humans
Authors: Yoshida Yoko , Terawaki Yoshiro , Nakaya Rintaro
Published: November 2013

Temperature dependence of transfer was examined with ten R plasmids originating from clinical isolates of Salmonella. Six of the plasmids were thermosensitive upon transfer, five of which were originally harbored in S. typhimurium and the remaining one in S. derby. One of these plasmids, pNR502, which conferred resistance to kanamycin, streptomycin (Sm) and tetracycline (Tc) on its host was stably maintained both in Salmonella and Escherichia coli at either 30, 37, or 43 C. Another plasmid, pNR516, which was resistant to chloramphenicol, sulfathiazole, Sm and Tc, was slightly unstable only at 43 C. The remaining four plasmids, pNR503, pNR510, pNR512 and pNR514, conferred resistance to Sm and Tc. Of these plasmids, the former two were stably maintained at both 30 and 37 C, but were unstable at 43 C. The latter two were slightly unstable at the lower temperatures and considerably unstable at 43 C. Kinetics of the transfer of the plasmid pNR503 revealed that the efficiency of transfer of the plasmid between E. coli strains was affected not only by the temperature of the conjugation but also by the preincubation temperature of the donor culture before the conjugation.
Leukocyte Migration Inhibition Test (LMIT) in Systemic Lupus Erythematosus
Authors: Tanaka Kohtaro , Yamauchi Yasutaka , Horimatsu Tohru , Isobe Takashi , Imura Hiroo
Published: November 2013

A leukocyte migration inhibition test (LMIT) utilizing the agarose gel technique was performed with native DNA as an antigen in ten patients with systemic lupus erythematosus (SLE) and five normal subjects. Irrespective of disease activity, supernatants obtained at different time intervals during lymphocyte culture in eight patients with SLE showed significant alteration of migration, either enhancement or inhibition, of normal leukocytes. However, supernatants in the control experiments produced no significant alteration of migration. Polyacrylamide gel electrophoresis of supernatants obtained from the SLE group revealed that the inhibitory activity was present in the albumin region, whereas the enhancement activity was found in the β‐globulin region. These results indicate that the hitherto employed estimation of the leukocyte migration inhibition test based on the total activity of these two factors is insufficient for accurate evaluation of chemical mediators from sensitized lymphocytes and that the separation of these two factors may be important for a greater understanding of cellular immunity.
Spontaneous Production of a C‐Type RNA Virus in a Cell Line Derived from Rat Glioma
Authors: Igarashi Koichi , Sasada Reiko , Niiyama Yasuhiko , Kozai Yoshio , Sugino Yukio
Published: November 2013

The spontaneous production of a rat C‐type RNA virus (ACV) in a cultured cell line (AC cells) established from a chemically induced rat glioma was studied. The characteristics of ACV were: morphology typical of C‐type RNA virus; buoyant density of 1.15 g/ml in a sucrose density gradient; RNA directed DNA polymerase activity; viral core with a density of 1.28 to 1.30 g/ml; 70S RNA with dimer structure; and structural protein composed of mainly four polypeptides. Kinetical analysis of DNA‐DNA hybridization revealed that DNA sequences homologous to DNA transcripts of RNA of ACV were present in rat cells. RNA directed DNA polymerase of ACV partially cross‐reacted with antiserum to the polymerase of Rauscher murine leukemia virus. These data suggest that ACV is an endogenous C‐type RNA virus of rat origin.
Physico‐ and Immunochemical Properties of Staphylococcal Protein A Extracellularly Produced by a Set of Mutants from Staphylococcus aureus Cowan I
Authors: Movitz Jan , Masuda Shogo , Sjöquist John
Published: November 2013

Mutant strains of Staphylococcus aureus have been isolated using a simple cosedimentation technique. While the parental strain contains predominantly a cell‐bound protein A, the mutant strains exclusively produce extracellular protein A. The mutant forms of protein A all have lower molecular weights than that of protein A from the parental strain. They showed the same antigenicity as the parental protein A and gave similar reactivity with immunoglobulin to the parental one except for one mutant. A conspicuous spur was observed between the parental protein A and that produced by the mutant against normal dog serum in the micro‐Ouchterlony immunodiffusion test.
Pathogenesis of Hobbs' Heat‐Sensitive Spore Forming Clostridium perfringens Type A Strain†
Authors: Chakrabarty A.K. , Narayan K.G.
Published: November 2013

Food poisoning in man due to heat‐sensitive strains of Cl. perfringens type A appeared to be mediated through enterotoxin synthesized in vivo during sporulation. A minimum of 2.0 × 105 vegetative cells suspended in sporulation medium was sufficient to elicit gut‐loop response in rabbits. The functional disturbance in the gut as well as the structural changes were progressive and proportional to the size of the inoculum up to a dose limit of 2.0 × 107 vegetative cells and beyond this the changes remained steady.
Detection of Human B Cells and Chronic Myelocytic Leukemic Cells by Rosette Formation with Sheep Erythrocytes and Fresh Human Sera
Authors: Chen Po‐Min , Yamanaka Noboru , Kikuchi Kokichi , Kitajima Koichi , Kimura Ikuro
Published: November 2013

A new method of detecting the C3b receptor is reported. A particular merit of this method is that anti‐RBC rabbit antiserum is not required. Rosettes were formed with human B lymphocytes, B lymphoblasts and granulocytes, using sheep erythrocytes (SRBC) sensitized with fresh human serum (FHS). T lymphocytes and T lymphoblasts did not form rosettes. The percentage of cells forming rosettes with this method approximated the percentage of rosettes formed with EACm. However, FHS coated SRBC did not react with most cells of B cell type chronic lymphocytic leukemia (CLL), whereas EACm rosette formations showed a definite reaction. On the other hand, 34–58% of cells of chronic myelocytic leukemia (CML) bound with the indicator red cells. SRBC sensitized with fresh rabbit or guinea pig serum formed rosettes with PBL, tonsil cells, B lymphoblasts and granulocytes. Complement and IgM antibody were required for this reaction, as in EAC rosette formation.
The Induction of Interferon by Vesicular Stomatitis Virus in Mouse L Cells
Authors: Nishiyama Yukihiro , Ito Yasuhiko , Shimokata Kaoru , Kimura Yoshinobu
Published: November 2013

Although no detectable interferon was produced when L cells were infected with wild‐type VSV (VSV‐o), considerable amounts of interferon were produced when cells were infected with UV‐irradiated VSV‐o at a multiplicity equivalent to 10 PFU/cell. Treatment of VSV‐o with UV‐light resulted in the marked reduction of the RNA synthesizing capacity and cytotoxity of the virus, and the UV‐irradiated virus had neither infectivity nor interfering activity against homologous viruses. The amount of interferon induced by UV‐VSV‐o was markedly influenced by multiplicity of infection and incubation temperature. Less‐virulent temperature‐sensitive mutants (VSV‐mp and VSV‐sp) derived from L cells persistently infected with VSV induced interferon in L cells without treatment of the viruses with UV‐light, but these viruses could not induce interferon if the infected cells were incubated at nonpermissive temperature, or if cells were infected at multiplicities of more than 10 PFU/cell. On the other hand, it was shown that treatment of cells with cycloheximide (100 μg/ml) delayed the expression of cell damage caused by non‐irradiated VSV‐o and resulted in the production of interferon when cycloheximide was removed from the cultures. These results indicate that VSV has intrinsically interferon‐inducing capacity in L cells and can induce interferon if the induction is carried out under such condition that cell damage caused by VSV are suppressed or delayed. Furthermore, the effect of pretreatment of cells by interferon and undiluted passage of VSV‐o on interferon induction was discussed in relation to persistent infection.
Different Effects of the Capsular Polysaccharide of Klebsiella pneumoniae on the Functions of Various Subpopulations of B Cells in the Immune Response of Mice to T‐Dependent Antigen
Authors: Nakashima Izumi , Yokochi Takashi , Nagase Fumihiko , Kato Nobuo
Published: November 2013

Using the capsular polysaccharide of Klebsiella pneumoniae (CPS‐K) as a polyclonal B‐cell activator (PBA) and sheep red blood cells (SRBC) as a T‐dependent antigen, we studied the effects of PBA on the functions of various subpopulations of B cells in the immune response of mice to T‐dependent antigen. Antibody‐forming cells (AFC) of IgM and IgG types were estimated as anti‐SRBC direct and indirect plaque‐forming cells (PFC), and the B cells with precursor activities involving generation of AFC and supplementing new B cells as rosette‐forming cells (RFC) of the B‐cell type. Stimulation of normal mice by CPS‐K caused a definite increase in the number of direct PFC but not in that of indirect PFC and RFC in the spleens. The responsiveness of spleen cells of CPS‐K‐treated mice to generate PFC and RFC responses to a subsequent injection of SRBC was lower than that of CPS‐K‐untreated normal mice. In this case, the responsiveness to generate RFC and indirect PFC was inhibited more strongly by CPS‐K than that to generate direct PFC. When CPS‐K was injected into normal mice simultaneously with SRBC, CPS‐K never decreased but increased the levels of PFC and RFC responses to SRBC. In the spleens of SRBC‐primed mice, the number of RFC was markedly decreased following injection of CPS‐K, the number of direct PFC was increased only slightly and the number of indirect PFC was increased very slightly. The responsiveness of spleen cells of these CPS‐K‐treated SRBC‐primed mice to generate secondary PFC and RFC responses to a subsequent injection of SRBC was much lower than that of CPS‐K‐untreated SRBC‐primed mice. In this case, the responsiveness to generate the secondary RFC and indirect PFC responses was more strongly inhibited by CPS‐K than that to generate the secondary direct PFC response. When CPS‐K was injected into SRBC‐primed mice simultaneously with the secondary injection of SRBC, there were marked decreases in the level of the secondary RFC response and slight decreases in that of the secondary indirect PFC response, but little change in that of the secondary direct PFC response. From these results it has been concluded that CPS‐K provides the positive signal (the minor action) and the negative signal (the major action) to various subpopulations of B cells functioning at various stages of the immune response to T‐dependent antigen in different ways, and acts to regulate the levels of B‐cell responses to the antigen‐mediated positive signal.
Dysfunction of Thymocytes of C3H/HeJ Mice in Blastogenic Response to Anti‐Thymocyte Serum In Vitro and Its Repair with Lipopolysaccharide Induced Mediator
Authors: Tanabe Masao J. , Nakano Masayasu
Published: November 2013

In vitro mitogenic responses of thymocytes to rabbit anti‐mouse thymocyte serum (ATS) have been compared in several mouse strains. The response of thymocytes of C3H/HeJ mice is about one‐third of those of thymocytes of C3H/He, ATL or ATH strains. Phenol‐extracted bacterial lipopolysaccharide (LPS) does not induce mitogenic response in cultured C3H/HeJ spleen cells, but the spleen cells of all other strains used are capable of responding to LPS. The low response of C3H/HeJ thymocytes to ATS is restored by adding “endotoxin soup” prepared from spleen cell cultures of LPS‐responder mice in the presence of LPS. Neither soup prepared from C3H/HeJ spleen cell cultures without the addition of LPS nor soup prepared from cell cultures with LPS show such restoration of the response of C3H/HeJ thymocytes to ATS. The molecular size of the active factor in “endotoxin soup” was estimated on a Sepharose CL‐4B column and determined to be about 20,000 daltons. The activity of “endotoxin soup” is destroyed by heating at 70 C for 30 min or 80 C for 10 min and diminished by digestion with trypsin. The mechanisms of restoration of low response of C3H/HeJ thymocytes to ATS by “endotoxin soup” are discussed.
Effect of Carbohydrates and Control of Culture pH on Beta Toxin Production by Clostridium perfringens Type C
Authors: Sakurai Jun , Duncan Charles L.
Published: November 2013

Clostridium perfringens type C strain CN 5384 produced a higher level of beta toxin in a controlled pH medium containing 1% glucose, starch, or sucrose than in media with dextrin, fructose, or raffinose. Toxin synthesis was not related to the growth yield.

The effect of glucose on beta toxin production by 11 strains was investigated with and without control of the culture pH at 7.5. Strain CN 5386 produced distinctly higher toxin when the pH of the culture was maintained at 7.5, compared with uncontrolled pH.
Mode of Cell Separation and Arrangement of Staphylococcus
Authors: Amako Kazunobu , Umeda Akiko
Published: November 2013

The process of cell separation and arrangement of Staphylococcus was investigated using a scanning electron microscope.

After two cycles of cell division, the Staphylococcal cells cultured on an agar medium were generally observed to be arranged in three morphological types: linear, square, and crooked arrangements. Results of the examination of cell surface structure revealed that separations had occurred in these clustered cells following two patterns. One type of second separation occurred parallel to the transversal axis of the preceding pair of the parental cells (X‐type) and the other occurred tangential to it (Y‐type). In the former type, the four daughter cells were usually arranged tetragonally after the separations, and in the latter type they were arranged either linearly or crookedly depending on the direction of the second separation. The final pattern of the cell arrangement was thus determined by the type of septal wall formation and the direction of cell separation. After several cycles of cell divisions, the cells were finally arranged in an irregular grape‐like cluster, even though the cross walls were formed regularly at the rectangular face of the preceding cross walls.
Effects of In Vivo Priming on In Vitro Induction of Cytotoxicity
Authors: Yamada Hiroyasu , Nomoto Kikuo , Takeya Kenji
Published: November 2013

In vivo presensitization of donor mice of responding cells with third party cellular antigens augmented in vitro generation of cytotoxic T lymphocytes in allogeneic and xenogeneic combinations. In vitro induction of detectable cytotoxicity in presensitized responding cells required the incubation period needed for in vitro primary response. However, such cytotoxic T lymphocytes were generated after in vitro stimulation with monolayers of methylcholanthrene‐induced tumor cells, UV‐irradiated or heated spleen cells which had proved to be effective in secondary but not in primary response. Presensitized responding cells exposed to 600R‐irradiation did not augment in vitro induction of cytotoxicity in normal responding cells. The augmenting effect of presensitized responding cells may be attributable to radiosensitive T cells which are in a transitional state in differentiation from typical unprimed cells to typical primed cells.