Analyze Diet

Topic:Immunology

The equine immune system is a complex network of cells, tissues, and organs that work collaboratively to defend against pathogens and maintain homeostasis. It consists of innate and adaptive components, each with distinct functions and mechanisms. The innate immune system provides the first line of defense through physical barriers, phagocytic cells, and the complement system. The adaptive immune system involves lymphocytes, such as B cells and T cells, which generate specific responses to antigens and provide immunological memory. Research in equine immunology explores the interactions between these components, the impact of genetic and environmental factors on immune function, and the development of vaccines and therapeutics. This page gathers peer-reviewed studies and scholarly articles focusing on the mechanisms, regulation, and clinical applications of the equine immune system in health and disease.
Cytokine RNA expression in an equine CD4+ subset differentiated by expression of a novel 46-kDa surface protein.
Veterinary immunology and immunopathology    May 1, 1997   Volume 56, Issue 3-4 191-204 doi: 10.1016/s0165-2427(96)05752-2
Byrne KM, Davis WC, Holmes MA, Brassfield AL, McGuire TC.Two monoclonal antibodies (MAb), HB65A (IgG2a) and HB86A (IgGI), recognize a unique cell surface molecule on equine T-lymphocytes. The molecule, designated EqWC4, identified by these MAbs is present on a subpopulation of CD4+ equine lymphocytes (6.3-10.2% of Arabian lymphocytes CD4+ WC4+) and a smaller population of CD8+ lymphocytes (0.5% to 1.2% of Arabian lymphocytes CD8+ WC4+). EqWC4 is absent from B-lymphocytes, granulocytes, and macrophages. Both MAbs bound to a 46-kDa protein following immunoprecipitation reactions with lysates of surface labeled thymocytes. Immunoaffinity purification u...
Agonist-induced adherence of equine eosinophils to fibronectin.
Veterinary immunology and immunopathology    May 1, 1997   Volume 56, Issue 3-4 205-220 doi: 10.1016/s0165-2427(96)05740-6
Foster AP, McCabe PJ, Sanjar S, Cunningham FM.Eosinophils are believed to play an important part in the pathogenesis of equine diseases such as helminth infestation and the allergic skin disease, sweet itch. It has been shown that adherence of human eosinophils to the connective tissue matrix protein fibronectin enhances cell activation and survival time. If adherence causes similar changes in the properties of equine eosinophils, cell-induced tissue damage at a site of parasitic infestation or allergic response would be exacerbated. However, investigation of this hypothesis requires identification of mediators that cause equine eosinophi...
Serodiagnosis of Babesia equi infection–a comparison of Dot-ELISA, complement fixation test and capillary tube agglutination test.
Veterinary parasitology    May 1, 1997   Volume 69, Issue 3-4 171-176 doi: 10.1016/s0304-4017(96)01124-7
Kumar S, Malhotra DV, Dhar S.The present study aimed to develop Dot-ELISA, complement fixation test (CFT) and capillary tube agglutination test (CAT) for serodiagnosis of Babesia equi infection and to compare their sensitivity with each other. For this study, sequential serum samples were collected from four donkeys experimentally infected with B. equi up to 90 days post infection (P.I.). B. equi antigen was prepared from the blood of a donkey showing more than 80% parasitaemia. Dot-ELISA, CF and CA tests were standardized as per the standard method. While performing CFT, it was observed that CFT standardized for the donk...
Effect of antigen challenge on the activation of peripheral blood neutrophils from horses with chronic obstructive pulmonary disease.
Research in veterinary science    May 1, 1997   Volume 62, Issue 3 253-260 doi: 10.1016/s0034-5288(97)90200-9
Marr KA, Foster AP, Lees P, Cunningham FM, Page CP.The effect of antigen challenge on the state of activation of peripheral blood neutrophils from horses with chronic obstructive pulmonary disease (COPD) has been determined by measuring neutrophil superoxide anion formation. Prior to a seven-hour antigen challenge superoxide anion production by neutrophils from asymptomatic horses with COPD and normal horses in response to platelet activating factor (PAF) (with and without cytochalasin B), serum treated zymosan (STZ) and phorbol myristate acetate (PMA) was similar. Agonist-induced superoxide production by neutrophils from symptomatic COPD and ...
Characterization of five monoclonal antibodies specific for swine class II major histocompatibility antigens and crossreactivity studies with leukocytes of domestic animals.
Developmental and comparative immunology    May 1, 1997   Volume 21, Issue 3 311-322 doi: 10.1016/s0145-305x(97)00008-6
Bullido R, Doménech N, Alvarez B, Alonso F, Babín M, Ezquerra A, Ortuño E, Domínguez J.A set of five monoclonal antibodies (mAb) against porcine major histocompatibility complex (MHC), or swine leukocyte antigens (SLA), class II molecules has been characterized. These mAbs appear to recognize monomorphic determinants on SLA-DR (2F4, 1F12 and 2E9/13) and SLA-DQ (BL2H5 and BL4H2) molecules, as assessed by flow cytometry and immunoprecipitation. By Western blot, the 2F4, 1F12, BL2H5 and BL4H2 epitopes were located on the beta-chains of these molecules. mAbs 2F4 and 1F12 crossreact with leucocytes of dog, cattle, horse and human; mAbs 2E9/13, BL2H5 and BL4H2 bind leucocytes of cattl...
Melatonin protects mice infected with Venezuelan equine encephalomyelitis virus.
Cellular and molecular life sciences : CMLS    May 1, 1997   Volume 53, Issue 5 430-434 doi: 10.1007/s000180050051
Bonilla E, Valero-Fuenmayor N, Pons H, Chacín-Bonilla L.We investigated whether the administration of melatonin (MLT) reduces the death rate and evolution of the disease in mice infected with Venezuelan equine encephalomyelitis (VEE) virus. Our results show that, MLT protects mice infected with the virus. The mortality rate was reduced from 100% to 16% merely by increasing the dose from 0 to 1000 micrograms/MLT per kg body weight MLT significantly postponed the onset of the disease and death by several days. In surviving mice very high titres of VEE virus IgM antibodies were found seven weeks after virus inoculation. MLT significantly reduced VEE v...
Cloning, expression, purification, and characterization of the major core protein (p26) from equine infectious anemia virus.
Biochimica et biophysica acta    April 25, 1997   Volume 1339, Issue 1 62-72 doi: 10.1016/s0167-4838(96)00215-4
Birkett AJ, Yélamos B, Rodríguez-Crespo I, Gavilanes F, Peterson DL.The gene coding for the major core protein (p26) of the lentivirus equine infectious anemia virus (EIAV) was cloned from EIAV infected serum, expressed in E. coli, and the resultant protein purified to electrophoretic homogeneity. The protein was expressed in a soluble form and was purified by conventional protein separation methods. When analyzed by SDS-PAGE, under both reducing and non-reducing conditions, the purified protein migrated as a 26 kDa monomer. Recombinant p26 (rp26), therefore, does not contain any intermolecular disulfide bond. Gel filtration chromatography also indicated that ...
Equine arteritis virus.
Theriogenology    April 15, 1997   Volume 47, Issue 6 1275-1295 doi: 10.1016/s0093-691x(97)00107-6
Glaser AL, Chirnside ED, Horzinek MC, de Vries AA.Equine arteritis virus (EAV) is a small, enveloped, positive-stranded RNA virus, in the family Arteriviridae , W.H.ich can infect both horses and donkeys. While the majority of EAV infections are asymptomatic, acutely infected animals may develop a wide range of clinical signs, including pyrexia, limb and ventral edema, depression, rhinitis, and conjunctivitis. The virus may cause abortion and has caused mortality in neonates. After natural EAV infection, most horses develop a solid, long-term immunity to the disease. Marzz and geldings eliminate the virus within 60 days, but 30 to 60% of acut...
Control of equine infectious anemia virus is not dependent on ADCC mediating antibodies.
Virology    April 14, 1997   Volume 230, Issue 2 275-280 doi: 10.1006/viro.1997.8502
Tschetter JR, Byrne KM, Perryman LE, McGuire TC.Horses infected with equine infectious anemia virus (EIAV) have recurrent episodes of viremia which are eventually controlled, but the immune mechanisms have not been identified. Antibodies were detected to the surface of EIAV-infected cells within 1 month postinfection and remained for at least 3.5 years postinfection. These antibodies recognized cell surface-exposed envelope (Env) glycoproteins, but could not mediate antibody dependent cellular cytotoxicity (ADCC) using EIAV-WSU5-infected equine kidney (EK) cells as targets and peripheral blood mononuclear cells (PBMC) or polymorphonuclear c...
A novel family of viral death effector domain-containing molecules that inhibit both CD-95- and tumor necrosis factor receptor-1-induced apoptosis.
The Journal of biological chemistry    April 11, 1997   Volume 272, Issue 15 9621-9624 doi: 10.1074/jbc.272.15.9621
Hu S, Vincenz C, Buller M, Dixit VM.Molluscum contagiosum virus proteins MC159 and MC160 and the equine herpesvirus 2 protein E8 share substantial homology to the death effector domain present in the adaptor molecule Fas-associated death domain protein (FADD) and the initiating death protease FADD-like interleukin-1beta-converting enzyme (FLICE) (caspase-8). FADD and FLICE participate in generating the death signal from both tumor necrosis factor receptor-1 (TNFR-1) and the CD-95 receptor. The flow of death signals from TNFR-1 occurs through the adaptor molecule tumor necrosis factor receptor-associated death domain protein (TRA...
Insertions, duplications and substitutions in restricted gp90 regions of equine infectious anaemia virus during febrile episodes in an experimentally infected horse.
The Journal of general virology    April 1, 1997   Volume 78 ( Pt 4) 807-820 doi: 10.1099/0022-1317-78-4-807
Zheng YH, Nakaya T, Sentsui H, Kameoka M, Kishi M, Hagiwara K, Takahashi H, Kono Y, Ikuta K.We have studied a horse which exhibited typical clinical signs of disease when experimentally infected with a non-adapted virulent strain of equine infectious anaemia virus (EIAV), designated V70. Five viruses (F1V, F2V, F3V, F4V and F5V) were recovered during periodic febrile episodes. Cross-neutralization tests revealed that all of these variants and the parental V70 were antigenically distinct. Sequencing of their full-length env gp90 genes and gp45 5' sequences revealed novel mutations at a limited number of nucleotide positions, consisting of insertions and duplications in the gp90 princi...
Study of the duration and distribution of equine influenza virus subtype 2 (H3N8) antigens in experimentally infected ponies in vivo. Sutton GA, Viel L, Carman PS, Boag BL.The purpose of this experiment was to study the duration and distribution of equine influenza virus in actively infected ponies over a 3 wk period. Pony foals (6-8 mo old) were infected experimentally by nebulizing equine influenza subtype-2 virus ultrasonically through a face mask. Successful infection was clinically apparent as each of the foals (n = 6) had a febrile response, a deep hacking cough and mucopurulent nasal discharge for 7 to 10 d. The virus was isolated from nasopharyngeal swabs of all the ponies 3 and 5 d after infection and all the ponies seroconverted to the virus. Samples w...
Transforming growth factor-beta induced by live or ultraviolet-inactivated equid herpes virus type-1 mediates immunosuppression in the horse.
Immunology    April 1, 1997   Volume 90, Issue 4 586-591 doi: 10.1046/j.1365-2567.1997.00202.x
Charan S, Palmer K, Chester P, Mire-Sluis AR, Meager A, Edington N.Up to 21 days after exposure to live or ultraviolet-inactivated equid herpesvirus type-1 (EHV-1) autologous serum from ponies caused an immunosuppressive effect if incorporated into T-cell proliferation assays to EHV-1. The suppressive factor in the sera of ponies also inhibited T-cell response to phytohaemagglutinin. Increased levels of circulating activated transforming growth factor-beta 1 (TGF-beta 1) were detected, and the suppressive activity of the serum could be reversed by antibody to TGF-beta 1. In a challenge experiment the ponies which exhibited circulating TGF-beta 1 activity succ...
Identification of noncytopathic equine rhinovirus 1 as a cause of acute febrile respiratory disease in horses.
Journal of clinical microbiology    April 1, 1997   Volume 35, Issue 4 937-943 doi: 10.1128/jcm.35.4.937-943.1997
Li F, Drummer HE, Ficorilli N, Studdert MJ, Crabb BS.Equine rhinovirus 1 (ERhV1) is a recognized cause of acute febrile respiratory disease in horse, although the virus is rarely isolated from such animals, despite seroprevalence rates as high as 50% in some horse populations. Recently, ERhV1 has been shown to be most closely related to foot-and-mouth disease virus, raising questions as to its disease associations in horses. We report that ERhV1 infection was the likely cause of two separate outbreaks of severe febrile respiratory disease which involved more than 20 horses. Attempts to isolate ErhV1 from nasopharyngeal swabs by conventional cell...
Localization of regulatory peptides in the male urogenital apparatus of domestic equidae: a comparative immunohistochemical study in Equus caballus and Equus asinus.
Histology and histopathology    April 1, 1997   Volume 12, Issue 2 297-310 
Arrighi S, Domeneghini C.An immunohistochemical study was carried out on specimens of testis, excurrent duct including the male accessory glands and urethra in its various tracts in the horse and the donkey, in order to localize nine regulatory peptides. Immunoreactivities were tested by means of Labelled Strept Avidin-Biotin (LSAB) method. The study has shown that Equine male genitalia are supplied by many peptide immunoreactive nerves containing NPY-, VIP-, leu- and met-Enkephalin-, Substance P-, CGRP- and Bombesin/GRP-like peptides, each of them having a characteristic distribution pattern. These neurotransmitters ...
Antigenic polymorphism of the lipopolysaccharides from human and animal isolates of Bordetella bronchiseptica.
Microbiology (Reading, England)    April 1, 1997   Volume 143 ( Pt 4) 1433-1441 doi: 10.1099/00221287-143-4-1433
Blay KL, Gueirard P, Guiso N, Chaby R.Six monoclonal antibodies (mAbs) against lipopolysaccharides (LPS) from Bordetella pertussis (P1P3, 60.5), B. parapertussis (PP2, PP6, PPB) and B. bronchiseptica (BRg1) were used to examine the presence of antigenic determinants of LPS on B. bronchiseptica cells. Forty-eight clinical isolates of this Gram-negative bacterium (4 canine, 3 equine, 6 porcine, 4 rabbit and 31 human) were examined. Significant cross-reactivities with the heterologous anti-pertussis and anti-parapertussis mAbs were observed. The isolates also exhibited marked antigenic polymorphism. The 48 isolates could be classifie...
Phenotypic characterization of lymphocyte subpopulations in horses affected with chronic obstructive pulmonary disease and in normal controls.
Veterinary pathology    March 1, 1997   Volume 34, Issue 2 108-116 doi: 10.1177/030098589703400203
Watson JL, Stott JL, Blanchard MT, Lavoie JP, Wilson WD, Gershwin LJ, Wilson DW.The alterations in lymphocyte subsets in chronic obstructive pulmonary disease (COPD) in the horse were investigated by using monoclonal antibodies to identify CD3+, CD4+, CD8+, and surface immunoglobulin positive (sIg+) lymphocytes in peripheral blood, bronchoalveolar lavage fluid (BALF), and pulmonary biopsy frozen tissue sections. COPD-affected horses (n = 5) and normal controls (n = 5) were sampled prestabling and 14 days poststabling, at which time the COPD-affected horses wee exhibiting clinical signs of COPD. The peripheral blood absolute CD4+ lymphocyte count was significantly elevated...
Monoclonal gammopathy in a Dutch warmblood mare.
The veterinary quarterly    March 1, 1997   Volume 19, Issue 1 29-32 doi: 10.1080/01652176.1997.9694734
Geelen SN, Bernadina WE, Grinwis GC, Kalsbeek HC.A 15-year-old Dutch warmblood mare was presented because of lethargy, which had been present for several weeks, and severe anaemia. Total protein was high and serum electrophoresis revealed a monoclonal peak in the alpha-2 region. Monoclonal immunoglobulin, IgG(T), was detected by immuno-electrophoresis in serum and urine. Postmortem examination revealed a relatively large number of plasmacytoid cells in the bone marrow and a monotonous population of plasmacytoid cells in the spleen. These findings are suggestive of a plasma cell myeloma.
[A case of cutaneous epitheliotropic malignant lymphoma (mycosis fungoides) in a horse].
Berliner und Munchener tierarztliche Wochenschrift    March 1, 1997   Volume 110, Issue 3 86-89 
Hilbe M, Meier D, Feige K.This article describes an eleven year old mare with apathy, fever, enlarged mandibular lymph nodes, skin lesions on the upper lip and edematous, grey-red mucous membranes in the nose, mouth and vulva. Histopathology revealed infiltrates with atypical lymphocytes forming Pautrier's microabscesses. The neoplastic cells had large, often indented nuclei. Immunohistology showed that some cells were CD3-positive (Pan T-cell-marker). The diagnosis of cutaneous epitheliotropic malignant lymphoma (Mycosis fungoides) was made. The etiology in the horse is unknown.
Placental localization of relaxin in the pregnant mare.
Placenta    March 1, 1997   Volume 18, Issue 2-3 121-128 doi: 10.1016/s0143-4004(97)90083-7
Klonisch T, Mathias S, Cambridge G, Hombach-Klonisch S, Ryan PL, Allen WR.In situ hybridization employing a cRNA probe derived from a 428-bp fragment of equine relaxin was used to localize relaxin mRNA, and immunocytochemistry was used to localize relaxin itself, in tissues of the placenta-endometrium interface recovered between 33 and 153 days of gestation from mares carrying intraspecific horse, interspecific mule and extraspecific donkey conceptuses. Immunocytochemical staining was also used to localize trophoblast-specific and class I major histocompatibility complex (MHC) antigens on some specimens. Relaxin mRNA and relaxin were both present in the single-cell ...
Methods for the isolation, culture and characterisation of equine pulmonary artery endothelial cells.
Research in veterinary science    March 1, 1997   Volume 62, Issue 2 147-152 doi: 10.1016/s0034-5288(97)90137-5
MacEachern KE, Smith GL, Nolan AM.Equine endothelial cells were isolated from the pulmonary artery by enzymatic digestion and grown to confluency. The cells were characterised by positive immunofluorescent staining for von Willebrand factor and NADPH-diaphorase staining for nitric oxide synthase. Measurements of endothelins indicated that there were significant release rates from the cells for up to six hours. Measurements of intracellular calcium concentration showed that the application of bradykinin caused a transient increase in calcium concentration with similar characteristics to those observed in other endothelial cell ...
Medical management of a full-thickness tear of the retroperitoneal portion of the rectum in a horse with hyperadrenocorticism.
Journal of the American Veterinary Medical Association    March 1, 1997   Volume 210, Issue 5 665-667 
Mazan MR.A 30-year-old Morgan-Quarter Horse gelding with hyperadrenocorticism was referred for treatment of a full-thickness tear of the retroperitoneal portion of the rectum. In older horses, the caudal end of the peritoneal space may be farther cranial than is commonly thought. Thus, there is a greater chance that full-thickness rectal tears will involve the retroperitoneal, rather than the peritoneal, portion of the rectum. This horse had a quick recovery and good outcome, despite underlying hyperadrenocorticism that would be expected to impair healing. Although relatively little is known about mana...
Antibody directed against plasma membrane components of equine spermatozoa inhibits adhesion of spermatozoa to oviduct epithelial cells in vitro.
Biology of reproduction    March 1, 1997   Volume 56, Issue 3 720-730 doi: 10.1095/biolreprod56.3.720
Thomas PG, Ball BA, Ignotz GG, Dobrinski I, Parks JE, Currie WB.Before fertilization, equine spermatozoa adhere to oviduct epithelial cells (OEC) of the mare. The biochemical basis for this adhesion has not been determined. Our objective was to produce an antiserum to block this interaction. Ejaculated spermatozoa were subjected to nitrogen cavitation and spermatozoal plasma membranes enriched by sucrose density gradient centrifugation; membrane enrichment was confirmed by comparative alkaline phosphatase analysis, electron microscopy, and one- and two-dimensional PAGE. Periacrosomal plasma membrane was used as an immunogen for the production of an antiser...
Feed intake, body weight, body condition score, musculation, and immunocompetence in aged mares given equine somatotropin.
Journal of animal science    March 1, 1997   Volume 75, Issue 3 755-760 doi: 10.2527/1997.753755x
Malinowski K, Christensen RA, Konopka A, Scanes CG, Hafs HD.Sixteen 20- to 26-yr-old mares were given 0, 6.25, or 12.5 mg/d equine somatotropin (eST) to determine whether aged mares respond to ST with changes in feed intake, body weight, body condition score (based mostly on fat cover), or immunocompetence. Neither dry matter intake, body weight, nor body condition scores were altered during the 6 wk of eST injection. However, based on photographs taken to evaluate musculation before and after treatment (scores 0 to 4), mares given eST developed greater (P < .07) muscle definition (1.8 +/- .6 and 2.5 +/- .6 for 6.25 and 12.5 mg eST/d, respectively) ...
Frequency dependence of forced oscillatory respiratory mechanics in horses with heaves.
Journal of applied physiology (Bethesda, Md. : 1985)    March 1, 1997   Volume 82, Issue 3 983-987 doi: 10.1152/jappl.1997.82.3.983
Young SS, Tesarowski D, Viel L.The effect of measurement frequency on respiratory mechanics was investigated in six horses with reversible allergic airway disease. Total respiratory impedance was measured at 1.5, 2.0, 3.0, and 5.0 Hz by using the forced oscillation technique with the horses in remission, after acute antigenic challenge producing clinical heaves, and with heaves but after the administration of 2 mg fenoterol by inhalation. The slopes of the magnitude (magnitude of Zrs) and real part (R) of total respiratory impedance over the frequency range 1.5-3 Hz changed significantly after antigenic challenge and fenote...
Amantadine in man and horse–can we learn from each other?
Equine veterinary journal    March 1, 1997   Volume 29, Issue 2 89-91 doi: 10.1111/j.2042-3306.1997.tb01647.x
Oxford JS.The research examines the impact of administering amantadine to horses and humans to combat influenza A, speculating on potential benefits of dual-field research between human and animal health. The study […]
Serum antibodies to Sarcocystis neurona–half the horses in the United States have them!
Journal of the American Veterinary Medical Association    February 15, 1997   Volume 210, Issue 4 482-483 
MacKay RJ.No abstract available
Consideration of the optimum pH for the analysis of serum p-phenylenediamine oxidase activity in thoroughbred horses.
The Journal of veterinary medical science    February 1, 1997   Volume 59, Issue 2 137-139 doi: 10.1292/jvms.59.137
Okumura M, Asano M, Fujinaga T.The optimum pH for the measurement of serum p-phenylenediamine oxidase (Ox) activity was given (pH 6.6), and the relationship between serum ceruloplasmin (Cp) concentration and its Ox activity was established in healthy adult horses. In adult horses, serum antigenic Cp concentrations were measured by the single radial immunodiffusion (SRID) method with the affinity-purified antibody to equine plasma Cp and compared with its Ox activity. Efficient co-relation between Cp concentration and Ox activity in the sera (r = 0.93) and its Ox/Cp ratio were given. These results might contribute to the cal...
Case report: field-acquired subclinical Babesia equi infection confirmed by in vitro culture.
Journal of clinical microbiology    February 1, 1997   Volume 35, Issue 2 474-476 doi: 10.1128/jcm.35.2.474-476.1997
Holman PJ, Hietala SK, Kayashima LR, Olson D, Waghela SD, Wagner GG.A horse with no prior clinical history of equine piroplasmosis tested negative for Babesia caballi and Babesia equi in the complement fixation test before importation into the United States from France. After 5 years in residence in the United States, the animal tested serologically positive for B. equi by the complement fixation test, the immunofluorescent antibody test, and Western blot analysis. The carrier status of the horse was confirmed by culture of B. equi parasites. In vitro culture offers an efficient and comparatively inexpensive method to determine the carrier status of horses sus...
Experimental induction of equine protozoal myeloencephalitis in horses using Sarcocystis sp. sporocysts from the opossum (Didelphis virginiana).
Veterinary parasitology    February 1, 1997   Volume 68, Issue 3 199-213 doi: 10.1016/s0304-4017(96)01112-0
Fenger CK, Granstrom DE, Gajadhar AA, Williams NM, McCrillis SA, Stamper S, Langemeier JL, Dubey JP.Sarcocystis sp. sporocysts isolated from eight feral opossums (Didelphis virginiana) were pooled and fed to 18 commercially reared budgerigars (Melopsittacus undulatus), 14 wild-caught sparrows (Passer domesticus), one wild-caught slate-colored Junco (Junco hyemalis) and five weanling horses (Equus caballus). All budgerigars died within 5 weeks post inoculation (wpi). Histologic examination revealed meronts within the pulmonary epithelia and typical Sarcocystis falcatula sarcocysts developing in the leg muscles. Sparrows were euthanized 13 and 17 wpi and their carcasses were fed to four labora...