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Topic:Phagocytosis

Phagocytosis in horses refers to the process by which certain cells, known as phagocytes, engulf and digest foreign particles, bacteria, and cellular debris. This cellular mechanism is a part of the innate immune response, helping to protect horses from infections and maintain tissue homeostasis. Key phagocytic cells in horses include neutrophils and macrophages, which are responsible for identifying and eliminating pathogens. Research on phagocytosis in equines explores the efficiency, regulation, and impact of this process on overall health and disease resistance. This page includes peer-reviewed research studies and scholarly articles that investigate the mechanisms, cellular interactions, and implications of phagocytosis in equine immunology.
Immunological and haematinic consequences of feeding a standardised Echinacea (Echinacea angustifolia) extract to healthy horses.
Equine veterinary journal    July 11, 2002   Volume 34, Issue 3 222-227 doi: 10.2746/042516402776186001
O'Neill W, McKee S, Clarke AF.This study was undertaken to compile new data on the efficacy of Echinacea in stimulating the immune system of the horse. Use of Echinacea is becoming widespread in horses, despite an absence of controlled laboratory research into its effectiveness or safety. This paper documents results of a double-blind, placebo-controlled, cross-over trial investigating the effect of standardised Echinacea extract on 8 horses. Animals were supplemented with Echinacea or placebo for 42 days, and their response to supplements recorded. Treatment with Echinacea increased phagocytic ability of isolated neutroph...
Simultaneous flow cytometric analysis of phagocytosis and oxidative burst activity in equine leukocytes.
Veterinary research communications    April 2, 2002   Volume 26, Issue 2 85-92 doi: 10.1023/a:1014033016308
Flaminio MJ, Rush BR, Davis EG, Hennessy K, Shuman W, Wilkerson MJ.This paper describes a method for simultaneously measuring phagocytosis and oxidative burst activity in equine peripheral blood leukocytes by flow cytometry. Opsonized propidium iodide-labelled Staphylococcus aureus (PI-Sa) was used to measure the uptake of bacteria by equine phacocytes and the oxidative burst activity by oxidation of dihydrorhodamine 123. The requirements to achieve optimal activity of phagocytosis and oxidative burst are described. The advantage of the simultaneous technique is that it provides both independent and comparative values for phagocytosis and the oxidative burst,...
Opsonic capacity of foal serum for the two neonatal pathogens Escherichia coli and Actinobacillus equuli.
Equine veterinary journal    January 5, 2002   Volume 33, Issue 7 670-675 doi: 10.2746/042516401776249381
Gröndahl G, Sternberg S, Jensen-Waern M, Johannisson A.Two of the most commonly isolated foal pathogens are Escherichia coli and Actinobacillus equuli. The hypothesis tested in this study was that young foals carry a lower opsonic capacity for these bacteria compared to adult horses. A flow-cytometric method for the phagocytosis of these by equine neutrophils was established. The opsonic capacity of serum from healthy foals from birth to age 6 weeks was evaluated and related to the concentrations of IgGa and IgGb. Phagocytosis of yeast was used as a control. Serum was required for phagocytosis, with higher concentrations for E. coli than for A. eq...
Neutrophil functions and serum IgG in growing foals.
Equine veterinary journal    January 5, 2002   Volume 33, Issue 7 676-680 doi: 10.2746/042516401776249327
Demmers S, Johannisson A, Gröndahl G, Jensen-Waern M.The aim of this study was to investigate the phagocytic and killing capacities as well as expression of CD18 of neutrophils obtained from healthy foals from birth to age 8 months. Blood was taken from 6 Standardbred foals at 7 time-points between ages 2-56 days and thereafter once a month. For comparison, cells from 16 mature horses were evaluated. Neutrophil phagocytosis of yeast cells was assessed by flow cytometry after opsonisation with mature pooled serum, autologous serum or anti-yeast IgG. The killing capacity of the neutrophils, as indicated by the oxidative burst, was monitored by che...
Interaction between equine semen and the endometrium: the inflammatory response to semen.
Animal reproduction science    December 18, 2001   Volume 68, Issue 3-4 273-278 doi: 10.1016/s0378-4320(01)00164-6
Troedsson MH, Loset K, Alghamdi AM, Dahms B, Crabo BG.Insemination of mares with bacteria-free equine spermatozoa results in an influx of polymorphonuclear neutrophils (PMNs) into the uterine lumen. In vitro studies have demonstrated that equine spermatozoa activate complement, resulting in cleavage of factors C5a and C3b. Since uterine secretion is rich in complement, it is likely that an interaction between spermatozoa and uterine secretion results in C5a-mediated chemotaxis and migration of PMNs into the uterine lumen. Once in the uterine lumen, the PMNs phagocytize bacteria and spermatozoa, which is an important part of sperm elimination from...
The fibrinogen-binding protein (FgBP) of Streptococcus equi subsp. equi additionally binds IgG and contributes to virulence in a mouse model.
Microbiology (Reading, England)    December 12, 2001   Volume 147, Issue Pt 12 3311-3322 doi: 10.1099/00221287-147-12-3311
Meehan M, Lynagh Y, Woods C, Owen P.The major cell-wall-associated protein of the equine pathogen Streptococcus equi subsp. equi is an M-like fibrinogen-binding protein (FgBP) which binds equine fibrinogen (Fg) avidly, through residues located at the extreme N-terminus of the molecule. In this study, it is shown that FgBP additionally binds equine IgG-Fc. When tested against polyclonal IgG from ten other animal species, it was found that FgBP binds human, rabbit, pig and cat IgG, but does not bind mouse, rat, goat, sheep, cow or chicken IgG. Through the use of a panel of recombinant FgBP truncates containing defined deletions of...
A comparison of foal and adult horse neutrophil function using flow cytometric techniques.
Research in veterinary science    October 23, 2001   Volume 71, Issue 1 73-79 doi: 10.1053/rvsc.2001.0490
McTaggart C, Yovich JV, Penhale J, Raidal SL.Flow cytometric assays were used to compare phagocytic and oxidative burst activity of neutrophils from healthy foals less than 7 days of age with the activity of cells from healthy adult horses. The phagocytosis of Staphylococcus aureus by foal neutrophils was less than that observed for adult neutrophils when autologous serum was used as the source of opsonins in the assay. The use of adult serum did not significantly improve the ability of foal neutrophils to attach bacteria. The oxidative burst activity of foal neutrophils was equivalent to that of adult cells. However, when serum or plasm...
Effects of training on resting peripheral blood and BAL-derived leucocyte function in horses.
Equine veterinary journal    May 16, 2001   Volume 33, Issue 3 238-243 doi: 10.2746/042516401776249651
Raidal SL, Rose RJ, Love DN.In this study, the effects of prolonged, high intensity training on aspects of peripheral blood and bronchoalveolar lavage (BAL)-derived leucocyte function were evaluated in 8 horses. All horses undertook a 7 week endurance training programme, followed by 5 weeks of high intensity training (HIT). Thereafter, horses were divided into control (C) and overtraining (OT) groups. The frequency and intensity of training were increased more substantially for horses in the OT group. Training was terminated in week 32 when horses in the OT group demonstrated a significant performance reduction. Peripher...
The effect of high intensity exercise on the functional capacity of equine pulmonary alveolar macrophages and BAL-derived lymphocytes.
Research in veterinary science    July 6, 2000   Volume 68, Issue 3 249-253 doi: 10.1053/rvsc.1999.0373
Raidal SL, Love DN, Bailey GD, Rose RJ.The effect of strenuous exercise on the functional capacity of pulmonary alveolar macrophages (PAM) and bronchoalveolar lavage-derived lymphocytes was determined in eight horses prior to and after 7 weeks of training. Strenuous exercise had no effect on the total cell count or the percentage of live cells in bronchoalveolar lavage (BAL) samples prior to or following training. However, training was associated with a significant increase in the total cell count of pre-exercise BAL samples and a significant reduction in the percentage of live cells in post-exercise samples. Strenuous exercise was...
Streptococcus equi with truncated M-proteins isolated from outwardly healthy horses.
Microbiology (Reading, England)    June 10, 2000   Volume 146 ( Pt 6) 1361-1369 doi: 10.1099/00221287-146-6-1361
Chanter N, Talbot NC, Newton JR, Hewson D, Verheyen K.The M-protein genes of Streptococcus equi isolated from 17 outwardly healthy horses after 4 strangles outbreaks had ended, including a quarantined animal, were compared with those of S. equi isolates from 167 active cases of strangles across 4 countries. The healthy horses included 16 persistent S. equi carriers, at least one from each of the four outbreaks. These carriers, despite being outwardly healthy, had empyema of the guttural pouch(es), an enlargement of the equine Eustachian tube. A persistent carrier from two of these outbreaks, the quarantined animal and a healthy animal with normal...
Effect of single bouts of moderate and high intensity exercise and training on equine peripheral blood neutrophil function.
Research in veterinary science    April 11, 2000   Volume 68, Issue 2 141-146 doi: 10.1053/rvsc.1999.0349
Raidal SL, Love DN, Bailey GD, Rose RJ.The effects of single bouts of moderate (30 to 40 per cent VO(2)max) and high (115 per cent VO(2)max) intensity exercise on equine peripheral blood leucocyte function were evaluated by determining neutrophil phagocytosis and oxidative burst activity before and after treadmill exercise and training. Prior to all exercise tests, the possible effect of diurnal variation was evaluated in samples obtained from four resting horses. Subsequently eight horses underwent moderate and high intensity exercise protocols and then commenced a 17-week training period. High intensity exercise tests were repeat...
Sperm transport and survival in the mare: a review.
Theriogenology    March 29, 2000   Volume 50, Issue 5 807-818 doi: 10.1016/s0093-691x(98)00185-x
Troedsson MH, Liu IK, Crabo BG.After the deposition of semen in the mare's uterus, spermatozoa must be transported to the site of fertilization, be maintained in the female tract until ovulation occurs, and be prepared to fertilize the released ovum. Sperm motility, myometrial contractions, and a spontaneous post-mating uterine inflammation are important factors for the transport and survival of spermatozoa in the mare's reproductive tract. Fertilizable sperm are present in the oviduct within 4 h after insemination. At this time, the uterus is the site of a hostile inflammatory environment. Our data suggest that spermatozoa...
Sperm transport and survival in the mare.
Theriogenology    March 25, 2000   Volume 49, Issue 5 905-915 doi: 10.1016/s0093-691x(98)00040-5
Troedsson MH, Liu IK, Crabo BG.Following the deposition of semen in the mares uterus, spermatozoa must be transported to the site of fertilization, be maintained in the female tract until ovulation occurs, and be prepared to fertilize the released ovum. Sperm motility, myometrial contractions, and a spontaneous post-mating uterine inflammation are important factors for the transport and survival of spermatozoa in the mares reproductive tract. Fertilizable sperm are present in the oviduct within 4 hours after insemination. At this time, the uterus is the site of a hostile inflammatory environment. Our data suggest that sperm...
Characterization of peripheral blood and pulmonary leukocyte function in healthy foals.
Veterinary immunology and immunopathology    March 14, 2000   Volume 73, Issue 3-4 267-285 doi: 10.1016/s0165-2427(00)00149-5
Flaminio MJ, Rush BR, Davis EG, Hennessy K, Shuman W, Wilkerson MJ.Studies in infants and foals indicate an age-dependent maturation of peripheral lymphocyte subsets. The age-dependent relationship for maturation of cellular immune responses, such as phagocytosis and lymphocyte responses of the peripheral and pulmonary-derived leukocytes, has not been characterized in foals. Lymphocyte subpopulations, mitogen stimulation response of lymphocytes, lymphokine-activated killing cell activity, phagocytosis and oxidative burst activity, and serum immunoglobulin (Ig) classes G and M concentrations were determined in developing foals. This study illustrates age-depen...
Comparison of the phenotypes of Streptococcus zooepidemicus isolated from tonsils of healthy horses and specimens obtained from foals and donkeys with pneumonia.
American journal of veterinary research    February 24, 2000   Volume 61, Issue 2 162-166 doi: 10.2460/ajvr.2000.61.162
Anzai T, Walker JA, Blair MB, Chambers TM, Timoney JF.To determine whether streptococcal pneumonia is caused by strains of Streptococcus zooepidemicus similar to those obtained from the tonsils of healthy horses. Methods: 5 tonsils from healthy horses, 8 tracheal washes and 6 lung specimens from foals with pneumonia, and 5 nasopharyngeal swab specimens from donkeys with acute bronchopneumonia. Methods: Variable M-like protectively immunogenic SzP proteins of 5 isolates of S. zooepidemicus from each tonsil and clinical specimen were compared, using immunoblots. The SzP gene of 13 isolates representative of various SzP immunoblot phenotypes from 1 ...
Sequential changes in bronchoalveolar cytology after autologous blood inoculation.
Equine veterinary journal. Supplement    February 5, 2000   Issue 30 126-130 doi: 10.1111/j.2042-3306.1999.tb05202.x
McKane SA, Slocombe RF.Six horses, free of recent exercise-induced pulmonary haemorrhage (EIPH), each had 8 bronchial segments inoculated with 40 ml of autologous blood lavaged on Days 3, 5, 7, 10, 14 and 21 after inoculation. The bronchoalveolar lavage (BAL) fluid samples were analysed, and total leucocyte counts and differential cell percentages determined. Statistical analysis of the differences between the 8 post inoculation and 1 pre-inoculation (control) lavage sites demonstrated an early neutrophilic response, rising from < 5% of alveolar leucocytes to 10% by 24-48 h post inoculation. As this response wane...
The role of seminal plasma in post-breeding uterine inflammation.
Journal of reproduction and fertility. Supplement    January 1, 2000   Issue 56 341-349 
Troedsson MH, Lee CS, Franklin RD, Crabo BG.The effect of seminal plasma on polymorphonuclear neutrophil (PMN) chemotaxis, PMN phagocytosis and complement-induced cytolysis was determined using blood plasma pooled from four horses and seminal plasma pooled from two stallions. To investigate chemotaxis, complement in blood plasma was activated with E. coli lipopolysaccharide in the presence of 0-50% seminal plasma diluted with a standardized volume of McCoy's medium and placed in a chemotactic chamber. Chemotaxis of blood derived equine PMNs toward the chemoattractants was determined after incubation at 37 degrees C for 45 min. To invest...
Effects of Actinobacillus equuli culture supernatants on equine neutrophil functions and survival.
Zentralblatt fur Veterinarmedizin. Reihe B. Journal of veterinary medicine. Series B    December 22, 1999   Volume 46, Issue 9 595-602 doi: 10.1046/j.1439-0450.1999.00285.x
Sternberg S, Johannisson A, Magnusson U, Jensen-Waern M.After exposure of equine granulocytes from both foals and adult horses to culture supernatants from clinical isolates of Actinobacillus equuli, phagocytic capacity and respiratory burst was examined by flow-cytometry and a chemiluminescence assay, respectively. One haemolytic isolate of an equine Actinobacillus was also included in the study. An average decrease of 22% in total number of granulocytes, in the flow cytometric assay (P < 0.01), and an average decrease of 26% in light emission, in the chemiluminescence assay (P < 0.001), was seen after exposure to bacterial culture supernata...
Functional characterization of equine dendritic cells propagated ex vivo using recombinant human GM-CSF and recombinant equine IL-4.
Veterinary immunology and immunopathology    December 10, 1999   Volume 71, Issue 3-4 197-214 doi: 10.1016/s0165-2427(99)00094-x
Hammond SA, Horohov D, Montelaro RC.Naive T cells can be activated both in vivo and in vitro by specialized antigen presenting cells, dendritic cells (DC), with potent antigen-specific, immunostimulatory activity. Indeed, DC can provide an extremely powerful and important immunological tool by which to potentiate the immune response for specific recognition of foreign antigens. Until recently, the direct isolation of DC from PBMC required laborious procedures with extremely poor yields (<0.1%). Methods have been developed for the human, lower primate, and murine model systems to propagate large numbers of DC from PBMC or bone...
Morphological study on pigmented cells in the horse testis.
The Journal of veterinary medical science    November 24, 1999   Volume 61, Issue 10 1183-1186 doi: 10.1292/jvms.61.1183
Murabayashi H, Hondo E, Kitamura N, Furuoka H, Taguchi K, Nambo Y, Yamada J.One of the most attractive characteristics of a horse testis is the change of the weight during development. As the testicular weight changes and the number of Leydig cells decreases, pigments appear in interstitial tissues. In the present study, the characteristics of the pigments found in the interstitial tissues were examined histochemically and ultrastructurally. Specific stainings indicated that the pigmented granules showed almost all of the histological and histochemical characteristics of ceroid or ceroid-like pigment. The cells showed positive reaction for acid phosphatase while the p...
Erythrocyte associated haemato-biochemical changes in Babesia equi infection experimentally produced in donkeys.
Veterinary parasitology    September 17, 1999   Volume 85, Issue 4 319-324 doi: 10.1016/s0304-4017(99)00110-7
Ambawat HK, Malhotra DV, Kumar S, Dhar S.Equine babesiosis, caused by Babesia equi and transmitted by ticks is of major economic importance in India. The adverse effects which B. equi organism and its metabolites inflict on red blood cells have not been reported. Erythrocytes were analysed for red cell membrane phospholipids, proteins and haemoglobin (Hb) concentration and plasma for malondialdehyde (MDA) in B. equi carrier donkeys before splenectomy (50% parasitaemia. Before splenectomy the mean values of membrane protein, phospholipids, plasma MDA and Hb were found to be 1.63 +/- 0.12 mg/ml PCV, 2.28 +/- 0.9 mg/ml PCV, 3.63 +/- 0.3...
In vivo pathogenicity and resistance to phagocytosis of Streptococcus equi strains with different levels of capsule expression.
Veterinary microbiology    August 31, 1999   Volume 67, Issue 4 277-286 doi: 10.1016/s0378-1135(99)00051-6
Anzai T, Timoney JF, Kuwamoto Y, Fujita Y, Wada R, Inoue T.The glossy non-encapsulated strain of Steptococcus equi, NCTC 9682, was compared with the matt strain Hidaka/95/2 which expresses a medium sized capsule and with the mucoid CF32 which expresses a large sized capsule in phagocytosis assays and for virulence in inoculated horses. The three strains, NCTC 9682, Hidaka /95/2 and CF32 produced 2.0, 3.1, and 5.3 mg/g wet cells respectively after 3 h incubation, but similar amounts of M-like proteins, cytotoxin and mitogen. NCTC 9682 showed no resistance to phagocytosis by equine neutrophils regardless of the presence of opsonin while strains Hidaka /...
The effect of nonspecific immunostimulation of pregnant mares with 1,3/1,6 glucan and levamisole on the immunoglobulins levels in colostrum, selected indices of nonspecific cellular and humoral immunity in foals in neonatal and postnatal period.
Veterinary immunology and immunopathology    May 8, 1999   Volume 68, Issue 1 1-11 doi: 10.1016/s0165-2427(99)00006-9
Krakowski L, Krzyzanowski J, Wrona Z, Siwicki AK.The objectives of the studies were to evaluate the effect of levamisole and 1,3/1,6 glucan applied in pregnant mares on parameters of non-specific cellular and humoral immunity of foals. Eighteen mares in three experimental groups (six animals in each) and their progeny were examined. Multiparous mares, crossbreed of Polish, full-blood and Hannover lines (400-500 kg), 4-9 years old, originated from four different farms. They were kept under identical zoohygienic and nutritional conditions. The animals were randomly chosen in experimental groups. None of mares had been previously vaccinated. In...
The role of neutrophil chemotactic cytokines in the pathogenesis of equine chronic obstructive pulmonary disease (COPD).
Veterinary immunology and immunopathology    December 10, 1998   Volume 66, Issue 1 53-65 doi: 10.1016/s0165-2427(98)00178-0
Franchini M, Gilli U, Akens MK, Fellenberg RV, Bracher V.Chronic obstructive pulmonary disease (COPD) is defined as a chronic obstructive inflammatory disease affecting the small airways associated with hay dust exposure (Lowell, F.C., 1964. Observation on heaves. An asthma like syndrome in the horse, J. Allergy 35, 322-330). The disease corresponds histopathologically to a chronic bronchiolitis (Gerber, H., 1973. Chronic pulmonary disease in the horse, Equine Vet. J. 5, 26-33; Winder, N.C., Grünig, G., Hermann, M., Howald, B., von Fellenberg, R., 1989. Comparison of respiratory secretion cytology and pulmonary histology in horses, J. Vet. Med., A3...
The flow cytometric evaluation of phagocytosis by equine peripheral blood neutrophils and pulmonary alveolar macrophages.
Veterinary journal (London, England : 1997)    November 7, 1998   Volume 156, Issue 2 107-116 doi: 10.1016/s1090-0233(05)80036-x
Raidal SL, Bailey GD, Love DN.Flow cytometry was used to assess the phagocytosis of fluorescent-labelled bacteria by equine peripheral blood neutrophils and pulmonary alveolar macrophages. Cell populations were prepared from venous blood following ammonium chloride lysis and from washed bronchoalveolar lavage derived samples. Discrete clusters of cells, corresponding to different leucocyte groups, were readily identified on the basis of differing light scattering properties and could thus be discriminated, negating the need for prior cell separation. Cells able to associate with fluorescent-labelled bacteria (by attachment...
Immunologic function in horses after non-specific immunostimulant administration.
Veterinary immunology and immunopathology    July 10, 1998   Volume 63, Issue 4 303-315 doi: 10.1016/s0165-2427(98)00111-1
Flaminio MJ, Rush BR, Shuman W.Inactivated Propionibacterium acnes is a biologic response modifier for treatment of non-specific respiratory disease in horses. The objectives of this investigation were to determine alterations in phagocytic activity, phenotypic expression of lymphocyte subpopulations and lymphokine-activated killing cell response in healthy young horses. Samples were collected on day 0, 7 and 14 of the investigation. Blood samples were obtained via jugular venipuncture and pulmonary leukocytes were recovered via bronchoalveolar lavage (BAL). Commercially available P. acnes (Eqstim) was administered intraven...
Opsonic effect of equine plasma from different donors.
Veterinary microbiology    June 16, 1997   Volume 56, Issue 3-4 227-235 doi: 10.1016/s0378-1135(97)00091-6
Gröndahl G, Johannisson A, Jensen-Waern M.The ability of equine plasma from different donors to enhance phagocytic capacity was assessed in neutrophils obtained from seven foals, aged 7-8 days (Study A), and from seven adult horses (Study B). Neutrophils were allowed to phagocytize fluorescent yeast cells opsonized with plasma from one of three donors or with pooled serum, all previously frozen (-18 degrees C) and thawed. The results were analysed by flow cytometry. In study A, fresh autologous foal serum was also used for opsonization, and in study B, heat-inactivated plasma and pooled serum were used in addition to untreated samples...
The comparative biology of pulmonary intravascular macrophages.
Frontiers in bioscience : a journal and virtual library    June 1, 1997   Volume 2 d232-d241 doi: 10.2741/a186
Longworth KE.Pulmonary intravascular macrophages are an important part of the mononuclear phagocyte system in some species of mammals, mainly sheep and other ruminants, pigs, and horses. These cells phagocytize foreign particles, cell debris and pathogens that pass through the pulmonary circulation. Species with intravascular macrophages localize intravenously injected tracer particles and bacteria predominantly in the lung rather than the liver, and exhibit pulmonary hypertension when these cells are activated. Both in vivo and in vitro studies show that pulmonary intravascular macrophages have distinct s...
Effect of transportation on lower respiratory tract contamination and peripheral blood neutrophil function.
Australian veterinary journal    June 1, 1997   Volume 75, Issue 6 433-438 doi: 10.1111/j.1751-0813.1997.tb14349.x
Raidal SL, Bailey GD, Love DN.To evaluate the effect of transportation on lower respiratory tract contamination and peripheral blood neutrophil function in horses and to compare results from transported horses with those obtained in earlier experiments from horses confined with heads elevated. Methods: A prospective study. Methods: Six horses were transported by road for 12 h. Clinical and haematological examination, transtracheal aspiration and cell function studies were conducted before and after transportation. Results obtained after transportation were compared to pre-transportation values. Results: After transportatio...
Immunohistochemical detection of virulence-associated antigens of Rhodococcus equi in pulmonary lesions of foals.
Veterinary pathology    May 1, 1996   Volume 33, Issue 3 341-343 doi: 10.1177/030098589603300312
Madarame H, Takai S, Morisawa N, Fujii M, Hidaka D, Tsubaki S, Hasegawa Y.Rhodococcus equi was isolated from the lungs of six foals with bronchopneumonia. All isolates expressed 15-17-kd antigens by immunoblot analysis and contained a virulence-associated plasmid of 85 or 90 kb. Immunohistochemically, R. equi from all pulmonary lesions showed the expression of 15-17-kd antigens mainly in the phagocytic cells. The specific monoclonal antibody to 15-17-kd antigens of R. equi (MAb 10G5) may be an aid in the diagnosis of R. equi-induced pneumonia.