Bartonella spp antibodies and DNA in aqueous humour of cats.

Bartonella spp antibodies were measured in the serum and aqueous humour of cats with and without uveitis and polymerase chain reaction (PCR) for Bartonella spp DNA was performed on aqueous humour from most of the cats. Serum and aqueous humour were assayed from 49 client-owned cats with uveitis, 49 healthy shelter cats, and nine cats experimentally inoculated with either B henselae or B clarridgeiae, 454 days after inoculation. An aqueous antibody coefficient (C value) was calculated for cats positive for Bartonella spp antibodies in the aqueous humour. Ocular production of Bartonella spp IgG (C value >1) was detected in seven of 49 cats with uveitis, none of 49 healthy shelter cats, and four of nine experimentally inoculated cats. The organism was detected by PCR in the aqueous humour of three of 24 cats with uveitis, one of 49 healthy shelter cats, and four of nine experimentally inoculated cats. Bartonella spp infect the eyes of some cats following natural exposure or experimental inoculation and may cause uveitis in some cats.     

Enzyme-linked immunosorbent assays for the detection of Toxoplasma gondii-specific antibodies and antigens in the aqueous humor of cats.

Serum and aqueous humor samples, collected from 14 clinically normal cats and 96 cats with clinical evidence of intraocular inflammation, were assayed with ELISA for Toxoplasma gondii-specific immunoglobulin M (IgM), T gondii-specific IgG, T gondii-specific antigens, total IgG, and total IgM. Additionally, serum was assayed with ELISA for feline leukemia virus p27 antigen and antibodies against the feline immunodeficiency virus as well as with an immunofluorescent antibody assay for antibodies against feline coronaviruses. Calculation of the Goldmann-Witmer coefficient (C-value) for the T gondii-specific antibodies detected in aqueous humor established the likelihood of local antibody production. Serologic evidence of present or prior infection by an infectious agent was found in 81.9% of the clinically affected cats from which serologic results were available (77/94 cats). Seropositive results for toxoplasmosis were found in 74.0% of the clinically affected cats. Anterior segment inflammation was found in 93.1% (81/87 cats from which information was available) of the clinically affected cats, most of which were older males. Toxoplasma gondii-specific antibodies were not detected in the aqueous humor of 6 seropositive, clinically normal cats. The C-values for aqueous T gondii antibodies were greater than 1 in 44.8% of the cats and greater than 8 in 24.0% of the cats. Response to treatment with clindamycin HCl was positive in 15/20 (75%) of the T gondii-seropositive, clinically affected cats treated with this drug. In 13/15 (86.7%) T gondii-seropositive, clinically affected cats having a C-value greater than 1, response to treatment with clindamycin HCl was positive.(ABSTRACT TRUNCATED AT 250 WORDS)     

Tears and aqueous humor from horses inoculated with Leptospira contain antibodies which bind to cornea

An antigenic relationship between Leptospira interrogans and equine cornea was previously described by us. An enzyme-linked immunosorbent assay was employed in the present work to investigate the existence of anti-leptospira and anti-cornea antibodies in tears, aqueous humor and serum from horses inoculated i.m. with those antigens. Ten days after a booster by the same route, antibodies that bind to microtiter plates, coated with an homogenate of either equine cornea or leptospira, were detected in those fluids and in the sera. At the same time, the corneas of the horses began to develop a diffuse opacity. This finding of anti-leptospira antibodies in equine tears and aqueous humor shows the pathway along which they arrive at the cornea and bind to it.     

Detection of anti-lens crystallin antibody in dogs with and without cataracts

Objective To determine if antilens crystallin (ALC) serum and aqueous humor antibodies were present in normal dogs and dogs with cataracts, whether antibody incidence varied with stage of cataract, and whether antibody titer had a relationship to the presence of lens‐induced uveitis. Methods Serum and aqueous humor samples were obtained from normal dogs and dogs with cataracts. Lens crystallin was separated by SDS‐polyacrylamide gel electrophoresis (SDS‐PAGE), and antilens crystallin antibodies were detected by Western immunoblot analysis. An indirect ELISA using crystallin protein as antigen was also used to detect antilens crystallin antibodies in serum and aqueous humor. Test groups included normal, incipient, immature, mature, hypermature and diabetic cataract. Results SDS‐PAGE identified bands with molecular weights of lens crystallin subunits. Western immunoblotting demonstrated reaction between canine serum and these protein bands. The five canine serum samples that reacted with crystallin subunits on Western blots had corresponding reactivity on the ELISA. All aqueous humor samples (30) were negative. Serum ALC antibodies were detected in 59.3% (16/27) of controls, 66.7% (16/24) of incipients, 50.0% (10/20) of immatures, 37.9% (11/29) of matures, 28.6% (6/21) of hypermatures, and 26.7% (4/15) of diabetics. Serum ALC antibodies were detected in 43.1% (47/109) of all cataract samples. There was a statistically significant negative association between the presence (P = 0.004) and maturity (P = 0.004) of cataract and presence of ALC serum antibodies. In the immature and hypermature cataract groups, there was a statistically significant negative association between ALC serum antibody titer and severity of uveitis (95% confidence interval). Conclusions There is a negative association between the presence (P = 0.004) and maturity (P = 0.004) of cataract and presence of ALC serum antibodies.     

Bartonella spp infection as a possible cause of uveitis in a cat

A 6-year-old castrated mixed-breed cat was evaluated because of unilateral anterior uveitis. The cat was seronegative for antibodies to Toxoplasma gondii, coronaviruses, and feline immunodeficiency virus, and antigens for FeLV p27 and Cryptococcus neoformans. Antibodies to Bartonella spp were detected in serum and aqueous humor. The antibody coefficient (C value) for IgG antibodies to Bartonella spp in the aqueous humor was 4.42; values > 1 suggest ocular production of antibodies and supports a diagnosis of ocular infection. Topical administration of prednisolone and oral administration of prednisone failed to induce a response; however, the uveitis resolved rapidly after the cat was given doxycycline orally. Clinical or laboratory evidence of immunodeficiency in this cat was not detected. Detection of a serum IgG antibody titer to Bartonella spp and ocular production of IgG antibodies to Bartonella spp, exclusion of other causes of uveitis, and response to doxycycline suggests that the cat may have had bartonellosis resulting in uveal tract inflammation.     

Composition of a polysaccharide from mycoplasma (F-38) recognised by antibodies from goats with contagious pleuropneumonia

A polysaccharide was extracted by warm aqueous phenol from the F-38 strain of mycoplasma which causes contagious caprine pleuropneumonia (CCPP). After acid hydrolysis, the polysaccharide was found to be composed of the neutral sugars glucose, galactose, mannose and fucose and the amino sugars galactosamine and glucosamine. All the sugars were present in approximately equal quantities. Unmodified goat erythrocytes bound the polysaccharide readily and the sensitised cells reacted with antibodies in sera from goats with experimental or natural CCPP. The unique composition of the F-38 polysaccharide and the specific reactivity of polysaccharide-sensitised red cells with antibodies from CCPP infected animals suggests that the polysaccharide should be useful for identification of F-38 organisms and diagnosis of the disease.     

Role of bacteria in the pathogenesis of recurrent uveitis in horses from the southeastern United States

OBJECTIVE: To determine the role of intraocular bacteria in the pathogenesis of equine recurrent uveitis (ERU) in horses from the southeastern United States by evaluating affected eyes of horses with ERU for bacterial DNA and intraocular production of antibodies against Leptospira spp. SAMPLE POPULATION: Aqueous humor, vitreous humor, and serum samples of 24 clinically normal horses, 52 horses with ERU, and 17 horses with ocular inflammation not associated with ERU (ie, non-ERU inflammation). PROCEDURES: Ribosomal RNA quantitative PCR (real-time PCR) assay was used to detect bacterial DNA in aqueous humor and vitreous humor from clinically normal horses (n = 12) and horses with chronic (> 3-month) ERU (28). Aqueous humor and serum were also evaluated for anti-Leptospira antibody titers from clinically normal horses (n = 12), horses with non-ERU inflammation (17), and horses with confirmed chronic ERU (24). RESULTS: Bacterial DNA was not detected in aqueous humor or vitreous humor of horses with ERU or clinically normal horses. No significant difference was found in titers of anti-Leptospira antibodies in serum or aqueous humor among these 3 groups. Only 2 horses, 1 horse with ERU and 1 horse with non-ERU inflammation, had definitive intraocular production of antibodies against Leptospira organisms. CONCLUSIONS AND CLINICAL RELEVANCE: In horses from the southeastern United States, Leptospira organisms may have helped initiate ERU in some, but the continued presence of the organisms did not play a direct role in the pathogenesis of this recurrent disease.     

Antibody response in bovine pharyngeal fluid following foot-and-mouth disease vaccination and, or, exposure to live virus

Samples of pharyngeal fluid and serum were collected from cattle after exposure to live foot-and-mouth disease (FMD) virus (with or without prior vaccination) or after subcutaneous vaccination with inactivated virus. The pharyngeal fluid samples were examined for FMD neutralising activity and specific anti-FMD IgG, IgM and IgA antibodies. The neutralising activity of the serum was also monitored. A peak of neutralising activity which occurred in the pharyngeal fluid of unvaccinated cattle seven days after virus exposure corresponded to a rise in specific IgM and IgA antibodies. This peak appeared to be due to serum and tissue fluid escaping from the damaged mucosa during the acute inflammatory phase of infection. At later stages (20 to 60 days after virus exposure) the pharyngeal fluid neutralising activity corresponded to a rise in specific IgA antibodies, suggesting that active local antibody production was taking place. The pharyngeal fluid neutralising activity detected after revaccination with oil emulsion or aqueous vaccines, without exposure to live virus, corresponded to a rise in specific IgG and IgM antibody levels and this may have been due to serum transudation.     

Occurrence of various immunoglobulin isotopes in horses with equine recurrent uveitis (ERU)

We investigated 30 healthy eyes and 41 eyes with ERU from 57 horses. The total immunoglobulin titers and titers of IgGa, IgGb, IgM were measured in aqueous humour, vitreous and serum using different ELISA techniques. Every sample investigated contained detectable amounts of immunoglobulins. Compared to control eyes significantly increased titers were found in the aqueous humour and vitreous of the ERU eyes for all immunoglobulin isotypes studied (p < or = 0.01). While IgM was detected in only 2 out of thirty aqueous humour and in none of the thirty vitreous samples of healthy eyes, 79.6% of samples of ERU eyes revealed considerable IgM titers. Changes of the IgGa/IgGb ratio in the eye as compared to that in the autologous serum was more frequent in affected than in healthy eyes. In contrast to the intraocular immunoglobulins there were no significant differences in immunoglobulin serum titers in healthy horses and those affected with ERU (p > 0.05). In conclusion, the results argue for a physiological appearance of immunoglobulins in the healthy eye. The increased titers of immunoglobulins in eyes stricken with ERU might be signs either of a local ocular production of antibodies and/or an increased permeability of intraocular barriers.     

Dermatophagoides farinae-specific IgG responses in atopic dogs undergoing allergen-specific immunotherapy with aqueous vaccines

The molecular and immunologic mechanisms associated with successful allergen-specific immunotherapy (ASIT) have not been completely elucidated. The aim of this study was to characterize the changes in Dermatophagoides farinae -specific IgG in atopic dogs undergoing ASIT using aqueous vaccines. Fifteen atopic dogs with a positive skin test reaction to D. farinae were treated with aqueous vaccines for a minimum of 2 months following a standard protocol. Serum samples were collected before and during therapy and used to probe Western blots containing separated proteins of D. farinae . IgG responses were detected using a polyclonal goat anticanine IgG antibody and a chromogenic substrate 3,3'-diaminobenzidine. The blots were analysed using a semiquantitative digital image analysis system that evaluated the number and molecular weight of bands, as well as their intensity, which was related to IgG concentration. Prior to ASIT, all dogs showed allergen-specific IgG responses to various antigens of D. farinae . During ASIT, there was a significant increase in the total quantity of D. farinae -specific IgG antibodies to various antigens from the mite ( P  = 0.015). Significant increases were observed for a 98-kDa band ( P  = 0.015), likely to be Der f 15; bands with molecular weights between 50 and 70 kDa ( P  = 0.012); and bands between 30 and 45 kDa ( P  = 0.035). These findings provide support for the hypothesis that ASIT induces IgG blocking antibodies to allergens known to be relevant in canine atopic dermatitis.     

Concurrent Infection with Toxoplasma gondii and Feline Leukemia Virus: Antibody Response and Oocyst Production

Four adult cats (two testing positive and two negative for feline leukemia virus FeLV) were fed Toxoplasma gondii tissue cysts collected from the brains of mice. Two control cats (1 FeLV+, 1 FeLV-) were not fed cysts. The cats infected with T. gondii shed thousands of oocysts but remained clinically and physically normal, with hemograms and clinical chemistry values essentially unchanged irrespective of their FeLV status. Infection with FeLV did not increase the duration of oocyst shedding. At necropsy no significant lesions were found. T. gondii antibodies were detected by three serologic tests in the cats fed tissue cysts. The time necessary for an antibody response to T. gondii was not altered by the FeLV infection. Indirect hemagglutination (IHA) was the least reliable of the serologic tests studied; it detected antibodies later in the infection, and titers were less than in the other tests. Latex agglutination (LA) detected antibodies a few days before IHA, but titers were less than in modified direct agglutination (MAT). MAT detected antibodies earliest in the infection and also measured antibodies in aqueous humor and cerebrospinal fluid.     

Detection of foot-and-mouth disease virus infection in vaccinated cattle

The aim of this study was to evaluate the value of commercially available kits for the detection of foot-and-mouth disease (FMD) virus infection in vaccinated cattle. The cattle were vaccinated with a commercial aqueous FMD vaccine type A24 and subsequently challenged 28 days post vaccination with homologous FMD virus. Seven of eight animals were protected from clinical disease and all became carriers. They were bled sequentially for up to 130 days post infection and samples of sera were tested with three ELISA kits: CHEKIT FMD-3ABC, Ceditest FMDV-NS and SVANOIR FMDV 3ABC-Ab ELISA. The Ceditest kit appears to be relatively higher sensitive than the others. When examined with this ELISA, all cattle developed of FMDV nonstructural proteins (NSPs) antibodies and remained positive throughout the period of the experiment. The response of antibodies against 3ABC antigen delayed in two cattle challenged with FMDV A24 virus. One of the cattle reacted negatively in Svanoir ELISA kit and sera from two animals were found negative in CHEKIT ELISA. It can be concluded that all tested kits can be a promising tool for FMD control and eradication campaigns in situation where emergency vaccination was applied.     

免疫胶体金法检测磺胺甲噁唑残留的研究

应用胶体金免疫层析技术（GICA）进行磺胺甲嗯唑（SMZ）的残留检测研究。制备抗磺胺甲嗯唑多克隆抗体，并采用柠檬酸三钠法制备20nm胶体金颗粒。胶体金标记多抗后喷涂到玻璃纤维膜上，特异性抗原（SMZ-OVA）以线状包被在硝酸纤维素膜上，制成快速检测试纸条。滴加检测溶液后，若呈现一条红色条带则为阴性，无红色条带则为阳性。该方法对SMZ标准溶液的灵敏度达到50ng／mL，试纸条显色结果清晰，反应灵敏，整个检测反应在5～10min内完成，稳定性及重复性良好。     

Detection of feline herpesvirus-specific antibodies and DNA in aqueous humor from cats with or without uveitis.

OBJECTIVE: To determine whether uveitis in cats was associated with intraocular production of feline herpesvirus type 1 (FHV-1)-specific antibodies or with detection of FHV-1 DNA in aqueous humor (AH). ANIMALS: 44 cats with idiopathic uveitis, 29 cats with uveitis attributed to Toxoplasma gondii infection, 13 FHV-1 seropositive cats without uveitis, and 9 FHV-1 seronegative cats without uveitis. PROCEDURE: ELISA were used to detect FHV-1-specific antibodies and total IgG antibodies in serum and AH, and the Goldmann-Witmer coefficient (C-value) for intraocular antibody production was calculated. A polymerase chain reaction assay was used to detect FHV-1 DNA in AH. RESULTS: FHV-1 seroprevalence among cats with uveitis was not significantly different from seroprevalence among cats without uveitis. Intraocular FHV-1 antibodies were never detected in cats without uveitis. Significantly more cats with idiopathic uveitis (22/44) or with toxoplasmic uveitis (11/29) had evidence of intraocular antibody production (C-value > 1) than did cats without uveitis. Only cats with idiopathic uveitis had FHV-1 C-values > 8. Among cats with evidence of intraocular antibody production, cats with idiopathic uveitis had a significantly higher median FHV-1 C-value (9.61) than did cats with toxoplasmic uveitis (2.56). Overall, FHV-1 DNA was detected in AH from 12 cats, 11 of which had uveitis. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggest that FHV-1 can infect intraocular tissues of cats and that intraocular FHV-1 infection may be associated with uveal inflammation in some cats.     

Ophthalmologic and oculopathologic findings in red-tailed hawks and Cooper's hawks with naturally acquired West Nile virus infection

OBJECTIVE: To assess ophthalmologic features and ocular lesions in red-tailed hawks and Cooper's hawks naturally infected with West Nile virus (WNV). DESIGN: Original study. ANIMALS: 13 hawks. PROCEDURES: All hawks underwent complete ophthalmic examinations including slit lamp biomicroscopy and binocular indirect ophthalmoscopy. Eleven hawks were euthanized because of a grave prognosis; complete necropsies were performed. Eyes, brain, heart, and kidneys were processed for histologic and immunohistochemical examinations. Pooled tissue homogenates and aqueous humor samples were assessed for WNV nucleic acid via PCR assay, and anti-WNV antibody titers in aqueous humor and plasma were determined. RESULTS: All birds had similar funduscopic abnormalities including exudative chorioretinal lesions and chorioretinal scarring in a geographic or linear pattern. Eleven birds were euthanized, and 2 birds were released. Plasma from both released hawks and plasma and aqueous humor of all euthanized hawks that were evaluated contained anti-WNV antibodies. Except for 1 hawk, all euthanized hawks had WNV-associated disease (determined via detection of WNV antigen or nucleic acid in at least 1 organ). Histopathologic ocular abnormalities, most commonly pectenitis, were detected in all euthanized birds; several birds had segmental choroiditis, often with corresponding segmental retinal atrophy. West Nile virus antigen was detected in the retinas of 9 of the euthanized birds. In 2 hawks, WNV antigen was detected in the retina only. CONCLUSIONS AND CLINICAL RELEVANCE: Results indicated that funduscopically detectable chorioretinal lesions appear to be associated with WNV disease in hawks. Detection of ocular lesions may aid in antemortem or postmortem diagnosis of this condition.     

Evaluation of IFA, MAT, ELISAs and immunoblotting for the detection of anti-Toxoplasma gondii antibodies in paired serum and aqueous humour samples from experimentally infected pigs

The study evaluated the efficiency of diagnostic laboratory methods to detect anti-Toxoplasma gondii antibodies in paired serum and aqueous humour samples from experimentally infected pigs. 18-mixed breed pigs were used during the experiment; these were divided into two groups, G1 (infected group, n=10) and G2 (uninfected group, n=8). Infection was performed with 4 x 10(4) VEG strain oocysts at day 0 by the oral route in G1 animals. All pigs were euthanized at day 60, when retina, aqueous humour, and blood samples were collected. Anti-T. gondii antibody levels were assessed in serum (s) and aqueous humour (ah) by indirect immunofluorescence assay (IFA), modified agglutination test (MAT), m-ELISA (using crude membranes from T. gondii tachyzoites as antigen) and r-ELISA (using rhoptries from T. gondii tachyzoites as antigen). Polymerase chain reactions (PCR) of samples from the retina were performed by using Tox4 and Tox5 primers. Antibody titers of G1 animals ranged from 128 to 1024 and from 16 to 256 in serum and aqueous humour, respectively. There were differences in the correlation coefficients between IFA(s) x IFA (ah) (r=0.62, P=0.05), MAT(s) x MAT (ah) (r=0.97, P<0.0001); however, there was no significant difference between r-ELISA(s) x r-ELISA (ah) (r= 0.14, P=0.7). Antibodies present in serum and aqueous humour recognized similar antigens. Samples of retina were positive by PCR in 30% (3/10) of infected pigs. G2 animals remained without antibody levels and were PCR negative throughout the experiment. These results suggest that the use of a combination of tests and immunoblotting for paired aqueous humour and serum samples could improve the sensitivity and specificity for the diagnosis of ocular toxoplasmosis.     

Immediate hypersensitivity to leaf extracts of Callisia fragrans (inch plant) in a dog

Contact with members of the plant family Commelinaceae, which includes wandering jew (Commelina spp. formerly called Tradescantia spp.) and inch plant (Callisia fragrans), can cause cell-mediated contact dermatitis in dogs. However, reports of canine IgE-mediated hypersensitivity to these plants have not been published. The purpose of this study was to discover whether IgE antibodies specific for extractable components of C. fragrans could be identified in serum from a dog that had anaphylactic shock after exposure to the plant and after skin patch testing with the sap from a leaf of C. fragrans. Separate aqueous extracts of leaves and flowers of C. fragrans were subjected to sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and Western blotting. Serum from a dog with no history or symptoms of any allergies showed no specific IgE antibodies against the leaf extract. Serum from a dog with clinical symptoms of delayed, but not immediate hypersensitivity to leaf sap from C. fragrans, showed only minor IgE recognition of a single 65 k component in sap extracted from leaves harvested in summer but not in winter. However, IgE antibodies to a serum dilution of 1:200 specific for several components of the leaf extract were seen in serum from the dog that had anaphylactic shock after exposure to sap. The molecular weights of these molecules were in the range 51 k to 83 k. The bands on the immunoblots did not match with prominently stained protein bands in the gel, but instead identified molecules in a lightly stained area of the gel with diffuse bands. Testing for glycans indicated that the carbohydrate side chains of glycoproteins contributed significantly to the immunoreactivity of the putative allergens. All three dog sera failed to show any immunoreactivity against the extract from the flowers of C. fragrans.     

Clinical, cytologic and histopathologic manifestations of protothecosis in two dogs

Two cases of sudden onset of blindness associated with ocular protothecosis in dogs are reported. Both were adult, spayed female, mixed-breed dogs that lacked the usual clinical signs of systemic infection with Prototheca species. Physical abnormalities at the time of presentation were limited to the affected eyes which had serous discharge, hyperemic conjunctiva, and aqueous flare. The pupillary light reflexes were slow, and the menace reflexes were absent. Both dogs had glaucoma. Results of complete blood counts and serologic titres for antibodies to Blastomyces dermatitidis and Histoplasma capsulatum were within reference intervals. Protothecosis was diagnosed by cytologic analysis of vitreous humor and was confirmed at necropsy. These two cases were unusual because of their presenting signs and prolonged course of disease progression.     

旋毛虫53kuES重组蛋白单克隆抗体的制备及其免疫金标试纸条的研制

为建立一种简易、快速检测动物血清中的旋毛虫排泄分泌(Excretory-secretory,ES)抗原的胶体金免疫层析试纸法,本研究选用旋毛虫53 ku ES重组蛋白免疫BALB/c小鼠,应用杂交瘤细胞技术制备单克隆抗体(MAb).利用辛酸-饱和硫酸铵法和亲和层析柱联合法纯化MAb及兔抗旋毛虫53 ku ES多克隆抗体,利用枸橼酸三钠还原法制备的胶体金颗粒标记MAb,制备免疫金标试纸条.结果表明,获得的2株IgG1型杂交瘤细胞系4D10和5A2能够稳定分泌抗体,且效价高;胶体金标记MAb 4D10所制备的试纸条具有重复性好、灵敏性和特异性高的优点;该试纸条4℃条件下可密封保存6个月以上.     

Passive protection of mice with antiserum to neuraminidase fromPasteurella multocida serotype A:3

Antiserum to a partially purified neuraminidase fromPasteurella multocida, type A:3, was adsorbed with protease-digestedP. multocida type 3 lipopolysaccharide (LPS) to remove LPS immunoreactivity. The LPS-adsorbed antineuraminidase caused a 77% reduction in the neuraminidase activity of homologousP. multocida in anin vitro enzyme neutralization test. All 14 mice passively immunized with the adsorbed antineuraminidase were protected against challenge infection with homologousP. multocida in a mouse protection test. Ten out of 14 mice in one group that received antisera containing antibodies to both neuraminidase and LPS were protected. In contrast, only 1 out of 14 mice that were immunized with pre-immune serum survived the challenge. These results suggest that antiserum toP. multocida neuraminidase was, at least partly, responsible for the protection observed in this study. Neuraminidase may be one of the immunogenic protective proteins present in aqueous extracts ofPasteurella multocida.