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PubMed1994

Inhibition of C3 deposition on Streptococcus equi subsp. equi by M protein: a mechanism for survival in equine blood.

The effect of the M protein of Streptococcus equi subsp. equi on complement deposition, complement degradation, and bacterial survival in equine whole blood was examined in vitro. Preincubation of bacteria with rabbit M protein-specific immunoglobulin G (IgG) inhibited the survival of the M+ strain in whole blood by 20-fold (P < 0.01). In addition, preincubation of bacteria with M protein-specific F(ab')2 fragments inhibited the survival of M+ cells in whole blood by 3.8-fold (P < 0.01). In the absence of specific antibody, an M+ strain (CF32) of S. equi subsp. equi survived 100-fold better in whole blood than an M- isolate (strain 19) (P < 0.01). Complement inactivation by cobra venom factor significantly enhanced the ability of the M- and M+ strains of S. equi subsp. equi to survive in whole blood, the latter in the presence or absence of M protein-specific IgG. The major opsonic forms of C3, C3b and iC3b, were present on both M- and M+ cells after opsonization in nonimmune plasma. However, colloidal gold staining indicated that the M- strain bound four times as much C3 as the M+ strain (P < 0.02) and that preincubation of the M+ strain with M protein-specific IgG or F(ab')2 fragments also enhanced the amount of C3 deposited by a factor of 4 (P < 0.02). Therefore, at least part of the M protein's ability to enhance bacterial survival in equine whole blood may be related to its ability to interfere with the deposition of equine complement on the bacterial surface.

Animals↗
PubMed2006

Relatedness of Streptococcus equi subsp. zooepidemicus strains isolated from harbour seals (Phoca vitulina) and grey seals (Halichoerus grypus) of various origins of the North Sea during 1988-2005.

The present study was designed to identify 15 beta-hemolytic streptococci isolated during a period between 1988 and 2005 from nine harbour seals and six grey seals from various origins of the North Sea. All isolates were identified as Streptococcus equi subsp. zooepidemicus. The bacteria were additionally investigated for relatedness by restriction fragment length polymorphism analysis of PCR amplified 16S-23S rDNA intergenic spacer region and gene szp and by macrorestriction analysis of chromosomal DNA of the strains by pulsed field gel electrophoresis. The molecular analysis yielded identical or closely related patterns within the strains of the present study and with the S. equi subsp. zooepidemicus strains isolated from harbour seals of German North Sea which were investigated previously [Akineden, O., Hassan, A.A., Alber, J., El-Sayed, A., Estoepangestie, A.T.S., Lämmler, C., Weiss, R., Siebert, U., 2005. Phenotypic and genotypic properties of S. equi subsp. zooepidemicus isolated from harbor seals (Phoca vitulina) from the German North Sea during the phocine distemper outbreak in 2002. Vet. Microbiol. 110, 147-152]. This indicates that this single or closely related bacterial clone existed during both phocine distemper virus epidemics in 1988 and 2002 and that a direct transmission of the strains has occurred between two seal species and between seal populations of far distant regions possibly with grey seals as a vector.

Animals↗
PubMed1991

Field evaluation of a commercial M-protein vaccine against Streptococcus equi infection in foals.

A double-blind randomized clinical trial was undertaken to determine the value of parenterally administered Streptococcus equi M-protein vaccine in foals during an epizootic of strangles. Weaned mixed-breed foals (n = 664) housed on 2 adjacent feed-lots (A and B) arrived over a 5-day period, 2 weeks before primary vaccination. Foals in lot B (n = 114) were randomly administered vaccine (n = 59) or saline solution (placebo; n = 55) on 3 occasions at biweekly intervals. Foals in lot A (n = 450) were given 1 dose of vaccine (n = 225) or placebo. The following clinical observations were scored blindly by a single observer for all foals in lot B and for 120 (randomly sampled) foals in lot A on a single day, 2 (Lot B) and 6 (lot A) weeks after final vaccination: cervical lymphadenopathy, type of bilateral nasal discharge, and palpable swelling at injection site(s). Bacteriologic culture of nasal swab specimens or lymph node aspirates from selected foals with clinical disease yielded S equi. Cervical lymphadenopathy was observed in 17 of 59 (29%) vaccinates and 39 of 55 (71%) nonvaccinated controls in lot B and in 32 of 60 (53%) vaccinates and 29 of 60 (48%) controls in lot A. Contingency chi 2 analysis confirmed significantly lower cervical lymphadenopathy rate (chi 2 = 18.5; P less than 0.001) and prevalence of mucopurulent nasal discharge (chi 2 = 11.4; P less than 0.01) for vaccinates in lot B only. Swelling(s) at the vaccine injection site were palpated in 44% of lot B and 29% of lot A vaccinates vs less than 2% of placebo controls.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed2006

Effects of a Streptococcus equi infection--mediated nutritional insult during mid-gestation in primiparous Thoroughbred fillies. Part 1: placental and fetal development.

REASONS FOR PERFORMING STUDY: There is a paucity of information on factors that influence placental development in the horse. HYPOTHESIS: Changes in nutrition, particularly around the time of proliferative placental growth, can affect development of the placenta and fetal growth. OBJECTIVE: To investigate the effects of 2 planes of nutrition and an unforeseen infection-mediated nutritional insult on placental and fetal development in the mare. METHODS: Twenty maiden Thoroughbred fillies, age 3 or 4 years, mated to one Thoroughbred stallion, were maintained on either High or Moderate food intake throughout pregnancy. In mid-gestation all the mares unexpectedly became infected with Streptococcus equi and suffered varying degrees of weight loss as a consequence. Gross and stereological measurements of the placenta and measurements of foal birthweight, crown-rump length, ponderal index and plasma IGF-1 concentration were made at term. RESULTS: Gains in bodyweight during gestation were significantly higher in the High vs. the Moderate nutrition groups. Placental and fetal growth parameters were not influenced by the plane of nutrition. However, transient weight loss in mid-gestation from acutely reduced food intake resulted in morphological changes to the allantochorion and decreased foal birthweight. CONCLUSIONS: Excessive weight gain during gestation in maiden Thoroughbred fillies does not result in nutrient partitioning to the dam at the expense of the fetus. However, sudden weight loss in mid-gestation has detrimental effects on placental development which results in reduced fetal growth. POTENTIAL RELEVANCE: Clinicians should appreciate the likely effects of maternal infection and resulting weight loss on pregnancy outcome.

Animal Nutritional Physiological Phenomena↗
PubMed1982

Lymphocyte stimulation response in horses against phytohaemagglutinin and M protein of Streptococcus equi using whole blood.

Lymphocyte stimulation was observed in whole equine blood in the presence of phytohaemagglutinin and M protein extracted from a typical strain of Streptococcus equi. Blood samples were collected from several healthy horses and horse and pony foals and cultured in vitro with varying concentrations of phytohaemagglutinin and M protein for several days. Phytohaemagglutinin was found to induce lymphocyte stimulation in these animals. Highest mean stimulation indices in horse foals (49.3 +/- 24.4) and pony foals (54.7 +/- 32.0) were observed with 0.625 and 1.25 micrograms/mL phytohaemagglutinin, respectively, at either 72 or 96 hours of incubation. Significantly higher radioactive counts per minute in horse and pony foals were recorded in blood cultures incubated with 0.625 and 1.25 micrograms/mL phytohaemagglutinin. M protein induced a dose related stimulation response in adult horses. Maximum stimulation indices were observed against 125 micrograms/mL M protein at 96 hours. These stimulation indices were higher in adult horses (40.0 +/- 2.2) than observed in pony foals (14.4 +/- 15.7). Higher stimulation levels in adult horses indicated either nonspecific stimulation against M protein or previous exposure of these animals to S. equi.

Age Factors↗
PubMed1985

Serum bactericidal responses to Streptococcus equi of horses following infection or vaccination.

An indirect test based on horse blood was used to study bactericidal responses of the horse to Streptococcus equi following infection or vaccination. Bactericidal antibody appeared in convalescent sera between two and four weeks and high titres were usually attained by eight weeks. Infection without clinical evidence of abscessation was also effective in eliciting strong bactericidal responses. Serum bactericidal activity of horses either recovered from strangles or immunised with commercial bacterin had declined eight months after vaccination. However, horses that developed strangles eight to 10 months after vaccination exhibited rapid and substantial increases in serum bactericidal activity. Groups of yearlings immunised with commercial S equi vaccines consisting either of M protein or bacterin developed clinical strangles within six months of vaccination although the majority of the animals had exhibited strong serum bactericidal activity a few weeks before occurrence of the disease. Similarly, a group of seven yearling ponies hyperimmunised with experimental vaccine, rich in M protein, were found to be highly susceptible to an intranasal challenge of 5 X 10(8) colony forming units of S equi, although their sera exhibited strong bactericidal activity at the time of challenge. These observations suggest that the role of serum bactericidal antibody in protection of the horse against strangles has been overrated.

Animals↗
PubMed1993

Production of monoclonal anti-idiotype antibodies which mimic an M-like protein of Streptococcus equi.

Rat monoclonal anti-idiotype antibodies (mAb2) were raised against two mouse monoclonal antibodies (mAb1), 1D10 and 2A6, with specificity for the M-like protein of Streptococcus equi. The capacity of the mAb2 to inhibit the binding between the corresponding mouse mAb1 against which the mAb2 were raised and the M-like protein was investigated in an inhibition EIA. One of the ten mAb2 examined, namely 5D1 (anti-mAb1 1D10), was able to inhibit this binding. The mAb2 5D1 bound to the mAb1 1D10 in such a way as to completely inhibit the subsequent binding of the M-like protein antigen to the paratope of the mAb1 1D10. The mAb2 5D1 is likely to represent a true image of the M-like protein antigen and may thus be described as an Ab2 beta anti-idiotype antibody.

Animals↗
PubMed1991

Protection against Streptococcus equi infection by monoclonal antibodies against an M-like protein.

We have developed an in vivo passive transfer assay using mice to identify monoclonal antibodies (mAbs) which offer protection against Streptococcus equi infection. The assay was developed using serum antibodies collected from horses convalescing from strangles. In this study, we show that a preparation of M-like protein, acid-extracted from S. equi, affords 80% protection to mice immunized with it. A number of mouse mAbs directed against a preparation of M-like protein were then assessed for their ability to passively protect mice against challenge with a lethal dose of the bacteria. Two mAbs, 1D10 and 2A6, were shown to be highly protective. It was also demonstrated, by means of a competitive enzyme immunoassay, that these mAbs recognized different epitopes in the preparation. Examination of a dose-response curve for mAbs 1D10 and 2A6 revealed that optimal levels of protection were achieved using 1 mg of either 1D10 or 2A6, or 0.5 mg 1D10 and 0.5 mg 2A6 given together. Immunological reactivity of these mAbs with a preparation of M-like protein showed that the antigens they recognized were comparable in size to some of the antigens recognized by convalescent horse serum antibodies. The role of immunoglobulin isotype in conferring protection is discussed.

Animals↗
PubMedDate not supplied

Unusual patterns of serum antibodies to Streptococcus equi in two horses with purpura hemorrhagica.

An enzyme-linked immunosorbent assay (ELISA) was developed for use in horses to determine serum titers of antibodies of the immunoglobulin classes IgA, IgG, and IgM to Streptococcus equi M-like protein and culture supernatant protein antigens. Serum antibodies were determined in 28 adult horses, including 9 horses with recent S. equi infections, 17 horses without known exposure to S. equi, but without a history of respiratory disease in the preceding 4 months, and 2 horses with clinical purpura hemorrhagica. Serum IgA titers to culture supernatant protein antigen were highest in recently infected horses (P less than 0.001). Serial determinations of antibody titers in the horses with purpura showed that IgG antibodies to both S. equi M-like protein and culture supernatant protein antigens were undetectable initially, but later rose coincidental with clinical recovery from the disease. Possible mechanisms for these findings are discussed.

Animals↗
PubMed1983

Experimental Streptococcus equi infection in the horse: correlation with in vivo and in vitro immune responses.

Fourteen young outbred horses, divided into 2 groups on the basis of 18- or 24-hour skin-test reactions to Streptococcus equi, were inoculated nasopharyngeally with virulent S equi. Animals (n = 6, group I) with evidence of previous exposure to S equi (positive dermal response and existing serum antibodies), with one exception, developed minimal or no signs of disease after inoculation. In contrast, S equi skin-test negative and seronegative horses (n = 8, group II) developed predictable and severe clinical signs of infection after their inoculation, including shedding of the organism from nasal discharges and ruptured mandibular lymph nodes. Results of the present study indicate that resistance to virulent S equi infection is correlated with existing humoral and cellular immune responses to streptococcal antigens. In susceptible horses, recovery from infection was accompanied by the appearance of humoral antibodies and the acquisition of a positive skin-test response to S equi antigens.

Animals↗
PubMed1985

Immune complexes in purpura hemorrhagica of the horse contain IgA and M antigen of Streptococcus equi.

Purpura hemorrhagica is an acute disease of the horse characterized by edema of the head and limbs, leucocytoclastic vasculitis, petechial hemorrhages in mucosae, musculature and viscera, and sometimes glomerulonephritis. It is usually associated with strangles, an upper respiratory tract disease of the horse caused by Streptococcus equi. We have detected and characterized immune complexes in the sera of horses with poststrangles purpura hemorrhagica by using PEG precipitation and Western blot analysis. The immune complexes contained IgA and S. equi-specific antigens similar to those found in acid extracts. We propose that purpura hemorrhagica is an immune complex-mediated disease.

Animals↗
PubMed2000

Analysis of immunoreactivity to a Streptococcus equi subsp. zooepidemicus M-like protein To confirm an outbreak of poststreptococcal glomerulonephritis, and sequences of M-like proteins from isolates obtained from different host species.

The etiologic agent of a large 1998 outbreak of poststreptococcal acute glomerulonephritis (PSGN) in Nova Serrana, Brazil, was found likely to be a specific strain of Streptococcus equi subsp. zooepidemicus from contaminated cheese (S. Balter et al., Lancet 355:1776-1780, 2000). In the present study, we used a serologic screen for a known surface-exposed virulence factor to confirm the epidemiologic findings. Using primers flanking a previously characterized M-like protein gene (J. F. Timoney et al., Infect. Immun. 63:1440-1445, 1995), we amplified and sequenced the M-like protein (designated Szp5058) gene and found it to be identical among four independent acute-phase PSGN patient isolates. Convalescent-phase sera from 33 of 44 patients in the PSGN outbreak were found to contain antibodies highly reactive to a purified Szp5058 fusion protein, compared with 1 of 17 control sera (P < 0. 0001), suggesting that Szp5058 was expressed during infection and further implicating this strain as the cause of the PSGN outbreak. The predicted signal sequence and cell wall association motif of Szp5058 were highly conserved with the corresponding sequence from S. equi subsp. zooepidemicus SzpW60, while the predicted surface-exposed portions differed markedly between these two proteins. The 5' end of the szp5058 gene, including its variable region, was identical to the szp gene from another strain associated with a previous PSGN outbreak in England (M. Barham et al., Lancet i:945-948, 1983), and the corresponding szp sequence found from the Lancefield group C type strain isolated from a guinea pig. In addition, the hypervariable (HV) portion of szp5058 was identical to a previously published HV sequence from a horse isolate (J. A. Walker and J. F. Timoney, Am. J. Vet. Res. 59:1129-1133, 1998). Three other strains of S. equi subsp. zooepidemicus, including another strain previously associated with a PSGN outbreak, were each found to contain a distinct szp gene. Two of these szp genes had HV regions identical to szp regions from isolates recovered from different host species.

Amino Acid Sequence↗
PubMed1986

Characteristics of an R antigen common to Streptococcus equi and zooepidemicus.

An R antigen of the group C streptococcus S. equi that cross reacts with a similar antigen of S. zooepidemicus has been identified and characterized. It is acid, heat and trypsin resistant, but pepsin sensitive and has an isoelectric point of 4.8. The amino acids in highest concentration are glutamic, aspartic, alanine, leucine, and valine. Bacterial components released in a French Press contain large amounts of R antigen, which is present also in culture supernatants and acid extracts. It has a molecular weight of about 82,000. Trypsin extraction of cells yields molecules of predominantly 56,000 and 25,000 molecular weight that appeared by immunoblotting to be similar to those obtained by trypsinization of purified R protein from preparations derived from the French Press. Although horses naturally infected with S. equi or S. zooepidemicus develop cross reacting R antibodies, the role of the R antigen in pathogenesis is unknown. Purified R protein does not stimulate bactericidal antibody in horses nor is it protective for mice. Its occurrence in antigen preparations in assays for S. equi antibody could be a source of interpretive errors because of the presence in many sera of antibody to the immunologically similar R antigen of S. zooepidemicus, a normal nasopharyngeal commensal of Equidae.

Animals↗
PubMed1990

Studies of antigenic components in acid extracts of group C streptococci with special reference to Streptococcus equi.

For the determination of a species-specific antigen of Streptococcus (S.) equi, acid extracts of group C streptococcal strains from horses (S. equi, S. zooepidemicus, S. equisimilis) were investigated using polyacrylamide gel electrophoresis and the immunoblotting technique. Using sera of horses suffering from strangles as well as sera from horses with respiratory infection of unknown etiology, Western blotting yielded more or less multiple banding reactions with bands in the 70, 54, 42, 40, and 31-28 kd molecular weight ranges against extracts of all of the 3 different bacterial species. However, an antigen found in this study at 18-16 kd which was highly sensitive to trypsin, proved to react specifically and regularly only with the serum of horses exposed to S. equi. The specificity of the reaction was assured by antisera of rabbits and horses vaccinated against S. equi, S. zooepidemicus or S. equisimilis, respectively, and by cross absorption of a serum originating from a mare recovered from strangles with Lancefield group C and group G streptococci as well as a strain of S. pyogenes. According to Western blot results on 180 serum pairs from horses with clinical signs of respiratory infection, 15.6% of which gave positive reactions, the 18-16 kd antigen appears as a marker suitable for qualitative testing of horse sera for antibodies to S. equi.

Absorption↗
PubMed2006

Use of leukocytes as treatment for endometritis in mares experimentally infected with Streptococcus equi subsp. zooepidemicus.

This study compared four treatments for bacterial endometritis in mares experimentally infected with Streptococcus zooepidemicus. Twenty-five mares were used, 20 resistant and five susceptible to endometritis. Mares would be in estrus when infected. Twenty-four hours after inoculation, clinical, bacteriological and cytological examinations were performed and repeated until the first occurrence: negative cytology and no Streptococcus growth or the seventh day post-infection. All mares showed clinical signs of endometritis and were assigned to one of the following treatments: (1) intrauterine infusion of fresh leukocytes; (2) intrauterine infusion of frozen-thawed leukocytes; (3) intrauterine infusion of lysed leukocytes; (4) intrauterine infusion of recombinant human interleukin-8 (rhIL-8); (5) control. Mares were submitted to all treatments, with at least a 14-day interval between treatments in a Latin square design. Treatment did not affect (P=0.121) time needed for resistant mares to eliminate bacteria. Time needed for elimination of bacteria was similar in susceptible mares treated with fresh and frozen leukocytes (P=0.333). Susceptible mares treated with frozen leukocytes also did not differ from those treated with lysed leukocytes (P=0.227) for time to eliminate bacteria, but were significantly different (P>0.02) from those treated with rhIL-8 and control. In resistant mares, physical clearance ability was probably the responsible for bacterial elimination. Intrauterine infusions in susceptible mares with viable or lysed leukocytes associated or not to opsonizing factors, reduced the time to elimination of bacteria. Infusions with bactericidal effect (functional neutrophils and granules) was likely effective and responsible for the more rapid elimination of bacteria in susceptible mares.

Animals↗