Cross protection between an encapsulated strain of Staphylococcus hyicus and encapsulated strains of Staphylococcus aureus

Strain ST67P of Staphylococcus hyicus grew diffusely in regular serum-soft agar. With the addition of rabbit antisera prepared with Staph. aureus strains, Smith, NS58D or NS41D, capsular type A, B or C, respectively, the organisms converted to compact type growth. Mice immunized with heat-killed vaccine of strain ST67P showed significant resistance against either homologous or heterologous strains, Smith, NS58D and NS41D. Passive protective activities in rabbit antisera prepared with strains Smith, NS58D and NS41D were absorbed out with either homologous cell surface polysaccharide fraction or cell surface polysaccharide fraction extracted from strain ST67P. Well-defined large capsules were observed in ultra-thin sections treated with rabbit antiserum prepared with homologous strain conjugated with ferritin. Also, the capsule surrounded by ferritin granules was shown in ultra-thin sections treated with ferritin conjugated with antisera prepared with those heterologous strains although the capsular size was significantly smaller than those observed by homologous antiserum.     

Conversion of biological and immunological properties during a process of decapsulation in a strain of Staphylococcus hyicus

E. YOSHIDA, Y. ICHIMAN, M. SUGANUMA AND K. YOSHIDA. 1991. Repeated subculture at 42°C of Staphylococcus hyicus strain ST67P, which exhibits streaming-type growth in a soft-agar medium, yielded three variants, ST67L, ST67S and ST67C, which had different colonial morphologies; small compact colonies possessing long and short tails and perfect compact colonies. The parent strain and ST67L respectively gave strong and weak positive intensity when stained by rabbit antisera prepared by capsular type I and II strains of Staph. epidermidis conjugated with fluorescein isothiocyanate. Variant ST67L gave a positive result with antiserum prepared by capsular type I strain and no staining was observed with variants ST67S and ST67C against these antisera preparations. Strain ST67C had the lowest virulence although no remarkable difference was shown between the parent strain and variants ST67L and ST67S. The cell volume index of the parent strain was 1.35, 2.43 and 3.71 times larger than those of ST67L, ST67S and ST67C, respectively. The converting activity of rabbit anti-ST67P strain serum absorbed by strain ST67C required four times more of the organisms than strain ST67P, changing the colonial morphology of the strain from diffuse to compact type by the addition of antiserum to soft agar medium. Positive coagulase and false positive clumping factor reaction were shown in variants ST67C, but no remarkable alteration was observed with 19 biochemical properties determined by a conventional identification kit. In ulta-thin sections of the parent strain labelled with rabbit anti-strain ST67P serum conjugated with ferritin, large capsules surrounded by ferritin granules were demonstrated by electron microscopy. In variants ST67L and ST67S, but not ST67C medium size capsules surrounded by ferritin granules and only ferritin granules located around the cell wall, respectively, were observed.     

Conversion of biological and immunological properties during a process of decapsulation in a strain of Staphylococcus hyicus

Repeated subculture at 42 degrees C of Staphylococcus hyicus strain ST67P, which exhibits streaming-type growth in a soft-agar medium, yielded three variants, ST67L, ST67S and ST67C, which had different colonial morphologies; small compact colonies possessing long and short tails and perfect compact colonies. The parent strain and ST67L respectively gave strong and weak positive intensity when stained by rabbit antisera prepared by capsular type I and II strains of Staph. epidermidis conjugated with fluorescein isothiocyanate. Variant ST67L gave a positive result with antiserum prepared by capsular type I strain and no staining was observed with variants ST67S and ST67C against these antisera preparations. Strain ST67C had the lowest virulence although no remarkable difference was shown between the parent strain and variants ST67L and ST67S. The cell volume index of the parent strain was 1.35, 2.43 and 3.71 times larger than those of ST67L, ST67S and ST67C, respectively. The converting activity of rabbit anti-ST67P strain serum absorbed by strain ST67C required four times more of the organisms than strain ST67P, changing the colonial morphology of the strain from diffuse to compact type by the addition of antiserum to soft agar medium. Positive coagulase and false positive clumping factor reaction were shown in variants ST67C, but no remarkable alteration was observed with 19 biochemical properties determined by a conventional identification kit. In ulta-thin sections of the parent strain labelled with rabbit anti-strain ST67P serum conjugated with ferritin, large capsule surrounded by ferritin granules were demonstrated by electron microscopy.(ABSTRACT TRUNCATED AT 250 WORDS)     

Induction of Resistance in Mice by the Capsular Polysaccharide Antigens of Staphylococcus aureus

Mice actively immunized with capsular polysaccharides extracted from capsular type strains A, B, C, and D, determined by the serum-soft agar technique, were protected against lethal infection by homologous strains, but no animals survived infection by heterologous substance immunization even with at high doses. Passive protective antibody in rabbit antisera prepared using these strains was absorbed out only by homologous capsular polysaccharide in mice. These results indicated that resistance was specific for capsular polysaccharide. The substance contained mainly neutral sugar, small amounts of hexosamine, methyl-pentose, and phosphate although these amounts varied depending on the capsular types strains.     

Demonstration of capsule in a strain of Staphylococcus hyicus by an electron microscope

A compact type variant designated as strain ST67V was isolated by a high-temperature subculture method from strain ST67P of Staphylococcus hyicus , which was a diffuse type in serum-soft agar. The parent strain was relatively virulent in mice and resisted ingestion by mouse peritoneal cells. The variant strain, however, was avirulent in mice and no antiphagocytic activity was observed in the peritoneal cavity. In ultra-thin sections of the organisms treated with anti-ST67P rabbit anti-serum conjugated with ferritin, the outermost layer of the cell wall of the parent strain was covered with a well-defined capsule while no capsule was shown in the variant strain.     

The Relationship of Capsular-type of Staphylococcus epidermidis to Virulence and Induction of Resistance in the Mouse

Biochemical, immunological and morphological properties of mouse virulent Staphylococcus epidermidis strains ATCC-31432, SE-360 and SE-10 isolated from clinical specimens were compared. Heat-killed organisms and cell surface polysaccharides extracted from cell surface fractions induced resistance in mice only against challenge with the homologous strain. Hyperimmune rabbit serum prepared with these strains passively protected mice against challenge infection only with the homologous strain. Protective activity in immune sera was absorbed by homologous whole cell and cell surface polysaccharide, but not by heterologous organisms and cell surface polysaccharide. In agar diffusion tests, cell surface polysaccharides from strains ATCC-31432, SE-360 and SE-10 produced single precipitin lines only with the homologous antiserum. The outermost layer of ultra-thin sections of the three strains was labelled by homologous but not by heterologous ferritin-conjugated serum. Biochemical analysis of the cell surface polysaccharides showed that they were composed of hexosamine, glycerol, phosphorous, alanine, glycine and phenylalanine. The three strains ATCC-31432, SE-360 and SE-10 were regarded as different from each other. Thirteen of 300 fresh isolates of Staph. epidermidis randomly selected from human clinical specimens proved to be virulent for mice. With ATCC-31432, SE-360 and SE-10 tentatively designated as capsular-type I, type II and III, respectively, a majority of mouse virulent strains belonged to capsular-type II.     

Demonstration of capsule in a strain of Staphylococcus hyicus by an electron microscope

A compact type variant designated as strain ST67V was isolated by a high-temperature subculture method from strain ST67P of Staphylococcus hyicus, which was a diffuse type in serum-soft agar. The parent strain was relatively virulent in mice and resisted ingestion by mouse peritoneal cells. The variant strain, however, was avirulent in mice and no antiphagocytic activity was observed in the peritoneal cavity. In ultra-thin sections of the organisms treated with anti-ST67P rabbit anti-serum conjugated with ferritin, the outermost layer of the cell wall of the parent strain was covered with a well-defined capsule while no capsule was shown in the variant strain.     

Antigenic specificity of opsonophagocytic antibodies in rabbit anti-sera to group B streptococci.

An opsonophagocytic assay has been developed which requires human polymorphonuclear leukocytes, immune serum, and complement for optimal killing of Group B streptococci. Only with all three of these components was killing of greater than 1.0 log10 of the initial inoculum achieved, using rabbit antisera directed to homologous strains of each of the five known serotypes of Group B streptococci. Titers of specific antisera which opsonized the strains and resulted in greater than 1 log 10 reduction of colony-forming units, ranged from 1:100 (serotype Ib) to 1:3200 (serotype Ia). Cross-reactions between serotype-specific sera and heterologous strains were seen in certain instances. Type Ic strain and serotype Ic antiserum demonstrated cross-reactions with types Ia and Ib which were explainable by known shared antigens among these types. The only other cross-reaction which resulted in greater than 1 log 10 reduction in colony-forming units was when unabsorbed antiserum to strain Ia was used to opsonize a strain of serotype III. Opsonization of 10 serotype III strains was demonstrated with a single type III antiserum. Killing of nine of these strains required polymorphonuclear leukocytes, complement, and antiserum, but one strain, D136C, the reference strain, could be killed (greater than 1 log 10 reduction in colony-forming units) without either complement or specific antiserum. Inhibition studies were performed utilizing large m.w. polysaccharide antigens extracted from each serotype. These antigens inhibited opsonization of homologous strains by homologous antisera with 50% inhibition points ranging between 0.5 and 4 mug.     

Cross protection between a strain of Staphylococcus epidermidis and eight other species of coagulase-negative staphylococci

Passive protective activity of rabbit antiserum prepared by a representative capsular type II strain of Staphylococcus epidermidis in mice was absorbed out with homologous capsular type strains of S. simulans, S. cohnii, S. xylosus, S. hominis, S. capitis, S. hyicus, S. haemolyticus, and S. saprophyticus in addition to the homologous strain. The minimum amount of the strains required for absorption differed greatly, depending upon the strain. No absorption of the activity was shown with a strain of capsular type I and III of S. epidermidis, S. simulans, and S. cohnii, and a strain of capsular type III of S. hominis. These results suggest a possible capsular type specificity in the cross protection between strains of S. epidermidis and other species of coagulase-negative staphylococci.     

Mouse Virulent Strain of Staphylococcus epidermidis

Using 10⁹ or 10⁷ colony-forming units of a strain of Staphylococcus epidermidis (strain 1142) in saline or 5% mucin, respectively, 90 to 100% of mice died within 24 to 48 hr after intraperitoneal challenge infection. These organisms gradually multiplied in the peritoneal cavity when injected intraperitoneally into mice, while the mouse avirulent strain (strain 1124) rapidly decreased and no organisms were found there 20 hr after injection. This strain was capable of inducing resistance against challenge with homologous strains. The resistance appeared as early as the first week and disappeared the 4th week after the immunization. However, no resistance was induced with strain 1124 against challenge with strain 1142. Also, hyperimmune rabbit serum prepared with strain 1142 passively protected against challenge with homologous strain in mice. The protective antibody was absorbed out with homologous organisms but not with strain 1124. Subsequently, a surface substance was obtained from strains 1142 or 1124 by the method of Morse. The 1142 surface substance was capable of inducing a resistance against challenge with the homologous strain but not with the 1124 surface substance. Also, this substance absorbed the protective antibody in hyperimmune rabbit serum prepared with the homologous strain but not with the 1124 surface substance nor with the Smith surface antigen extracted from the Smith strain of Staphylococcus aureus. Conversely, the protective antibody in rabbit anti-Smith strain serum against challenge with the homologous strain was absorbed with the Smith surface antigen but not with the 1142 surface substance. In the agar diffusion test, the 1142 surface substance and the Smith surface antigen produced single precipitin lines only against homologous antisera. Biochemical analysis of the 1142 surface substance showed that the substance contained neither nucleic acids nor proteins but is composed of hexosamine, glycerol, phosphorus, alanine, glycine and phenylalanine.     

Mouse virulent strain of Staphylococcus epidermidis. Relation of antiphagocytic activity to the protection-inducing antigen.

Using 10(9) or 10(7) colony-forming units of a strain of Staphylococcus epidermidis (strain 1142) in saline or 5% mucin, respectively, 90 to 100% of mice died within 24 to 48 hr after intraperitoneal challenge infection. These organisms gradually multiplied in the peritoneal cavity when injected intraperitoneally into mice, while the mouse avirulent strain (strain 1124) rapidly decreased and no organisms were found there 20 hr after injection. This strain was capable of inducing resistance against challenge with homologous strains. The resistance appeared as early as the first week and disappeared the 4th week after the immunization. However, no resistance was induced with strain 1124 against challenge with strain 1142. Also, hyperimmune rabbit serum prepared with strain 1142 passively protected against challenge with homologous strain in mice. The protective antibody was absorbed out with homologous organisms but not with strain 1124. Subsequently, a surface substance was obtained from strains 1142 or 1124 by the method of Morse. The 1142 surface substance was capable of inducing a resistance against challenge with the homologous strain but not with the 1124 surface substance. Also, this substance absorbed the protective antibody in hyperimmune rabbit serum prepared with the homologous strain but not with the 1124 surface substance nor with the Smith surface antigen extracted from the Smith strain of Staphylococcus aureus. Conversely, the protective antibody in rabbit anti-Smith strain serum against challenge with the homologous strain was absorbed with the Smith surface antigen but not with the 1142 surface substance. In the agar diffusion test, the 1142 surface substance and the Smith surface antigen produced single precipitin lines only against homologous antisera. Biochemical analysis of the 1142 surface substance showed that the substance contained neither nucleic acids nor proteins but is composed of hexosamine, glycerol, phosphorus, alanine, glycine and phenylalanine.     

Monoclonal IgM antibody protection in mice against infection with an encapsulated strain of Staphylococcus epidermidis.

Passive protective activities of three different classes of monoclonal antibodies in mice against challenge with strain ATCC 31432 (capsular type I) of Staphylococcus epidermidis were examined. Monoclonal IgM antibody passively protected mice against challenge with the homologous strain, whereas monoclonal IgG1 and IgG2b antibodies did not. The protective activity of IgM was absorbed by the cell surface antigen extracted from the homologous strain but not by the antigen from heterologous strains. Rapid reduction of viable cells took place in the peritoneal cavity of mice immunized with monoclonal IgM as early as 6 h after the challenge with the homologous strain. An enzyme-linked immunosorbent inhibition assay showed there was remarkable inhibition with the homologous cell surface antigen but not with heterologous preparations from other strains. Results suggest that in the mouse the major passive protection against the S. epidermidis strain is provided by the IgM antibody to the cell surface antigen.     

Protective antibodies in human sera against encapsulated strains of Staphylococcus epidermidis

Passive protective antibodies in 100 samples of normal human sera against challenge with three representative capsular type strains of Staphylococcus epidermidis in mice were examined. Six of them passively protected mice against capsular type I; 17 protected against capsular type II; one against capsular type III; and one against both capsular types I and II. The activities were sensitive to 2-mercaptoethanol and were absorbed out either with rabbit anti-human IgG serum, rabbit anti-human IgA serum or rabbit anti-human IgM serum. Also, the sera activities absorbed out with cell surface polysaccharide extracted from three representative capsular type strains. These results indicate that the protective activities were specifically related to three major immunoglobulins against the above cell surface polysaccharides.     

Protective antibodies in human sera against encapsulated strains of Staphylococcus epidermidis

Passive protective antibodies in 100 samples of normal human sera against challenge with three representative capsular type strains of Staphylococcus epidermidis in mice were examined. Six of them passively protected mice against capsular type I; 17 protected against capsular type II; one against capsular type III; and one against both capsular types I and II. The activities were sensitive to 2-mercaptoethanol and were absorbed out either with rabbit anti-human IgG serum, rabbit anti-human IgA serum or rabbit anti-human IgM serum. Also, the sera activities absorbed out with cell surface polysaccharide extracted from three representative capsular type strains. These results indicate that the protective activities were specifically related to three major immunoglobulins against the above cell surface polysaccharides.     

Application of the Laurell immunoelectrophoresis technique to the study of serological relationships between granulosis viruses

The Laurell technique of two-dimensional immunoelectrophoresis was used to distinguish between isolates of granulosis virus (GV) from Plodia interpunctella (GV strains A and B), Ephestia cautella, Spodoptera littoralis (GV strain 65), Pieris brassicae, and Cydia pomonella. Granules, alkali-soluble proteins, and virus particles of P. interpunctella GV strain A and granules of P. interpunctella GV strain B were used as sources of antigens. They were reacted with the immunoglobulins of antisera prepared against whole granules of each strain of virus. Peaks of precipitation were most clearly defined when antigens were pretreated with 0.1 m Na₂CO₃, 2% Triton X, and succinic anhydride, Granules and alkali-soluble proteins of P. interpunctella GV strain A treated in this way exhibited at least one peak of precipitation when reacted with each of the antisera studied. Four peaks were observed in both the homologous reaction and in the heterologous reaction with antiserum prepared against granules of P. interpunctella GV strain B. Four different peaks were present in the homologous reaction between immunoglobulins and virus particles of P. interpunctella GV strain A. Two peaks were present in the heterologous reaction with antiserum prepared against granules of P. interpunctella GV strain B and one in that with the antiserum prepared against granules of S. littoralis GV strain 65.     

Evidence for a Multiplicity of Capsular Types Among Staphylococcus aureus Strains

The Smith diffuse variant and the wound mucoid strain of Staphylococcus aureus were shown to exhibit serologically distinct capsules. The Welwood and K-6 strains of S. aureus were tested to determine their capsular types. Both Welwood and K-6 were found to be representative of the Smith capsular type. An additional 13 isolates of S. aureus from mice were tested. Gel double-diffusion tests and immunoelectrophoresis of staphylococcal antigens disclosed the possible existence of at least two additional capsular types. Passive hemagglutination tests carried out with cells sensitized with 1 mg of antigen per ml showed a multiplicity of cross-reacting antigens. However, cells sensitized either with 0.1 or 0.05 mg of antigen per ml and reacted with antisera absorbed with 10 or 1 mug/ml showed the presence of a specific antigen in each strain of S. aureus. Corroborative evidence for a multiplicity of capsular types was obtained by the specific capsular reaction. At least four capsular types of S. aureus were found. The prototypic strains for these antigens are the RLM or wound strain, the Smith diffuse strain, and mouse strains designated 36T and 43R. We propose to designate these types 1, 2, 3, and 4, respectively.     

Neutralization of activity of two different heat-stable enterotoxins (STh and STp) of enterotoxigenic Escherichia coli by homologous and heterologous antisera

Abstract Two distinct heat-stable enterotoxins (ST) produced by enterotoxigenic Escherichia coli , ST_p and ST_h, were purified and antisera against the purified ST_p and ST_h were prepared by immunizing rabbits with the purified ST's coupled with bovine serum albumin. Neutralizing activity of each antiserum was examined and it was found that both antisera neutralized not only homologous ST but also heterologous St. These data indicate that the two anti-ST antisera are raised against the region of common amino acid sequence of the two ST's.     

The effect of specific antiserum on the resistance of Neisseria gonorrhoeae to intracellular killing by phagocytes of human blood.

The high natural resistance of gonococci showing a characteristic 'double highlight' (DH) colonial morphology (Penn, Veale & Smith, 1977b) to intracellular killing by human phagocytes was markedly reduced by addition of rabbit antiserum to the phagocytosis medium or by preincubation of organisms with antiserum. Antisera raised to three different DH gonococcal strains showed a complex pattern of specificity in phagocytosis tests with the homologous organisms and three other DH strains. The effect of antiserum could be neutralized by adsorption with intact organisms or with extracts, prepared ultrasonically, of the homologous strain. Antiserum also promoted the intracellular killing of a strain which had a 'single highlight' colonial morphology (Penn et al., 1977b) and a low natural resistance to phagocytic killing, but adsorption with this strain neutralized the antiserum less consistently than the DH strain. The neutralization of antiserum-mediated promotion of intracellular killing by extracts of organisms naturally resistant to such killing may provide an assay for the aggressins responsible for this resistance.     

The characterization of antibacterial antibodies in bovine immune sera to Staphylococcus aureus

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. These studies suggest that IgG as well as IgM antibody might play an important role in protection against infection with encapsulated strains of S. aureus in cows.     

不同动物制备的抗血清对病毒抗原免疫反应的差异

血清学技术是病毒诊断、鉴定、分类及亲缘关系分析的重要手段。一般常用以制备抗病毒血清的动物是家兔,但也有采用其它动物的,如蛙、羊、豚鼠、鸡及小鼠等。本文比较了Balb/c小鼠、昆明种小鼠和新西兰大白兔对长叶车前花叶病毒上海分离株(RMVsh)和烟草花叶病毒普通株(TMVc)的免疫反应特征。