Psittaci Antigen Research Articles

Selection of a consortium of Chlamydia psittaci antigens for the construction of a multispecies vaccine against animal Chlamydia

Chlamydia of farm animals is an infectious disease that poses a potential threat to human health due to the possibility of disease as a result of contact with infected animals and birds. To date, vaccination remains the main method of prevention and control of infection. Currently, there is a question of developing preventive vaccines containing antigens in their composition, which make it possible to create stable immunity in most species of farm animals, which will expand the range of consumers due to economic benefits. The paper presents the results of studies on the selection of antigens from strains of the causative agent of chlamydia, which will serve as the basis for the design of a preventive drug that protects against infection. Antigens were obtained from strains isolated from various animal species with different forms of pathology. Chl. psittaci was cultured on developing chicken embryos, antigens were obtained from yolk membranes, inactivated and purified. The obtained antigens were studied for sterility, harmlessness, antigenic activity and immunogenicity for laboratory animals. The selection of the consortium was carried out on the basis of the conducted research. Antigens from strains 250, PC-85, AMK16 Chl. psittaci were selected as components for the future polyvid vaccine against chlamydia in farm animals.

Simultaneous Intramuscular And Intranasal Administration Of Chitosan Nanoparticles-Adjuvanted Chlamydia Vaccine Elicits Elevated Protective Responses In The Lung.

BackgroundChlamydia psittaci is a zoonotic bacteria closely associated with psittacosis/ornithosis. Vaccination has been recognized as the best way to inhibit the spread of C. psittaci due to the majority ignored of infections. The optimal Chlamydia vaccine was obstructed by the defect of single immunization route and the lack of availability of nontoxic and valid adjuvants.MethodsIn this study, we developed a novel immunization strategy, simultaneous (SIM) intramuscular (IM) and intranasal (IN) administration of a C. psittaci antigens (Ags) adjuvanted with chitosan nanoparticles (CNPs). And SIM-CNPs-Ags were used to determine the different types of immune response and the protective role in vivo.ResultsCNPs-Ags with zeta-potential values of 13.12 mV and of 276.1 nm showed excellent stability and optimal size for crossing the mucosal barrier with high 71.7% encapsulation efficiency. SIM-CPN-Ags mediated stronger humoral and mucosal responses by producing meaningfully high levels of IgG and secretory IgA (sIgA) antibodies. The SIM route also led to Ags-specific T-cell responses and increased IFN-γ, IL-2, TNF-α and IL-17A in the splenocyte supernatants. Following respiratory infection with C. psittaci, we found that SIM immunization remarkably reduced bacterial load and the degree of inflammation in the infected lungs and made for a lower level of IFN-γ, TNF-α and IL-6. Furthermore, SIM vaccination with CNPs-Ags had obviously inhibited C. psittaci disseminating to various organs in vivo.ConclusionSIM immunization with CNPs-adjuvanted C. psittaci Ags may present a novel strategy for the development of a vaccine against the C. psittaci infection.

Epidemiology of Chlamydia psittaci Infection in Racing Pigeons and Pigeon Fanciers in Beijing, China.

Over 3 million racing pigeons (Columba livia) are registered in Beijing City Center for gambling purposes. During 2008-2010, we evaluated the occurrence and prevalence of Chlamydia psittaci in racing pigeons as well as the possible zoonotic transmission to pigeon fanciers in six districts of Beijing where pigeon races are particularly popular. C. psittaci-specific serum antibody titres were obtained from 370 pigeons and 79 fanciers using enzyme-linked immunosorbent assay. In addition, 206 and 67 throat swabs were, respectively, collected from pigeons and fanciers and tested for the presence of chlamydial antigen using immunofluorescence. C. psittaci-specific serum antibody was detected in 37 of 370 pigeons and 19 of 79 fanciers. Of 206 pigeon clinical specimens, 55 were positive for C. psittaci antigen, while 16 of 67 swabs from the pigeon fanciers were positive. Based on ompA sequence analysis, the genotype of several avian and human isolates was genotype B. Thus, both high-titre C. psittaci-specific antibody and C. psittaci-specific antigen were found with relatively high frequency in the pigeon flocks as well as in the pigeon fanciers. Our study suggests that C. psittaci infection is prevalent among the racing pigeon population in Beijing. Moreover, detection of serum antibodies and antigen in pigeon fanciers suggests that exposure and possible zoonotic transmission of C. psittaci from racing pigeons to humans does occur. In view of the life-threatening respiratory illness C. psittaci may cause in humans, regulatory public health measures, to prevent further spread of the pathogen in avian populations and possible transmission to exposed humans, are urgently needed.

Evaluation of Chlamydophila psittaci infection and other risk factors for atherosclerosis in pet psittacine birds

To determine whether the presence of Chlamydophila psittaci antigen, plasma cholesterol concentration, diet, sex, species, and age are risk factors for the development of atherosclerosis in pet psittacine birds. Retrospective case-control study. 31 psittacine birds with atherosclerosis (study birds) and 31 psittacine birds without atherosclerosis (control birds). Necropsy reports were reviewed, birds with a histopathologic diagnosis of atherosclerosis were identified, and available medical records were reviewed. Signalment, history, clinicopathologic findings, and other relevant data were recorded and evaluated. Control birds did not have atherosclerosis and were chosen by both convenience sampling and population demographics. Histologic sections of great vessels from all birds (study and control birds) were reviewed and then submitted for immunohistochemical staining for the presence of C psittaci antigen. Result of immunohistochemical staining for C psittaci antigen in blood vessels was significantly associated with atherosclerosis. After adjusting for age, species origin, and type of illness, the odds of atherosclerosis was 7 times as high for birds with positive immunohistochemical staining for C psittaci antigen, compared with that of birds with negative immunohistochemical staining. Study birds and control birds differed significantly only with respect to plasma cholesterol concentrations. The median plasma cholesterol concentration of study birds (421 mg/dL) was significantly higher than that of control birds (223 mg/dL). Infection with C psittaci and a high plasma cholesterol concentration may be risk factors for developing atherosclerosis in pet psittacine birds.

Epidemiological study of Chlamydophila psittaci in pet birds in Croatia

A total of 411 samples from birds of different species originating from all counties of the Republic of Croatia have been tested for the presence of Chlamydophila psittaci. The sampling was conducted in pet stores, breeders' aviaries, in a specialized bird clinic and in zoos. The testing included 177 parrots, 169 pigeons, 58 canaries and 7 finches. For the detection of specific C. psittaci antigen a commercial ELISA kit was used- IDEIATM PCE Chlamydia (DAKO Cytomation Ltd., United Kingdom). The samples that were non-specifically positive or doubtful in the ELISA test (a total of 26 samples) were analyzed also by means of polymerase chain reaction (PCR). Diagnostic ELISA method found a total of 17.03% birds positive for chlamydiosis, and after additional testing by PCR a total of 12.65% positive ones were found. According to bird species, the most frequently positive ones were canaries and pigeons (15.52% and 13.02%), and according to the sampling location most of the positive birds were found in pet stores (16.52%), but a high percentage of positive samples were also found in breeders’ aviaries (11.76%). The average positive result for chlamydiosis in 12.65% of tested birds is alarming and it confirms the importance of monitoring bird health and of prescribed legal regulations when it comes to chlamydial diseases, as well as education of persons involved in breeding, keeping or selling birds.

HEALTH EVALUATION OF GALAPAGOS HAWKS (BUTEO GALAPAGOENSIS) ON SANTIAGO ISLAND, GALAPAGOS

Galapagos Hawks (Buteo galapagoensis), the only endemic, diurnal raptor species in Galapagos, are currently distributed on eight Galapagos Islands having been extirpated from three of the human-inhabited islands. In January 2009, we performed health assessments of 89 Galapagos Hawks on Santiago Island, Galapagos. Four of the 89 Galapagos Hawks (4%) evaluated had physical abnormalities. Blood parameters did not differ between males and females, except for aspartate transaminase values, which were significantly higher in females than males. No Galapagos Hawks tested positive for antibodies to avian encephalitis virus, Marek virus, and paramyxovirus-1 or to haemosporidian antigen. Chlamydophila psittaci antigen was detected in 2 of 86 Galapagos Hawks (2%), with 24 of 43 Galapagos Hawks (56%) antibody-positive for avian adenovirus-1 and 1 of 48 Galapagos Hawks (2%) antibody positive for Toxoplasma gondii. There were no significant differences in infectious disease results based on sex. This study contributes to the understanding of the health status of the Galapagos Hawk and to the establishment of baseline information for the species.

Chlamydophila spp. infection in horses with recurrent airway obstruction: similarities to human chronic obstructive disease

BackgroundRecurrent airway obstruction (RAO) in horses is a naturally occurring dust-induced disease mainly characterized by bronchiolitis which shows histological and pathophysiological similarities to human chronic obstructive pulmonary disease (COPD). In human COPD previous investigations indicated an association with Chlamydophila psittaci infection. The present study was designed (1) to clarify a possible role of this infectious agent in RAO and (2) to investigate the suitability of this equine disorder as a model for human COPD.MethodsClinico-pathological parameters of a total of 45 horses (25 horses with clinical signs of RAO and 20 clinically healthy controls) were compared to histological findings in lung tissue samples and infection by Chlamydiaceae using light microscopy, immunohistochemistry, and PCR.ResultsHorses with clinical signs of RAO vs. controls revealed more inflammatory changes in histology (p = 0.01), and a higher detection rate of Chlamydia psittaci antigens in all cells (p < 0.001) and bronchiolar epithelial cells alone (p < 0.001) by immunohistochemistry. The abundance of chlamydial inclusions increased with the severity of disease. PCR was positive in 60% of horses with RAO vs. 45% of the controls (p = 0.316). OmpA sequencing identified Chlamydophila psittaci (n = 9) and Chlamydophila abortus (n = 13) in both groups with no significant differences. Within the group of clinically healthy horses subgroups with no changes (n = 15) and slight inflammation of the small airways (n = 5) were identified. Also in the group of animals with RAO subgroups with slight (n = 16) and severe (n = 9) bronchiolitis could be formed. These four subgroups can be separated in parts by the number of cells positive for Chlamydia psittaci antigens.ConclusionChlamydophila psittaci or abortus were present in the lung of both clinically healthy horses and those with RAO. Immunohistochemistry revealed acute chlamydial infections with inflammation in RAO horses, whereas in clinically healthy animals mostly persistent chlamydial infection and no inflammatory reactions were seen. Stable dust as the known fundamental abiotic factor in RAO is comparable to smoking in human disease. These results show that RAO can be used as a model for human COPD.

Evaluation of two commercial assays for the detection of Chlamydophila abortus antibodies

Two commercial enzyme-linked immunosorbent assays (ELISA), the CHEKIT ®-CHLAMYDIA which uses inactivated Chlamydophila psittaci antigen, and the Chlamydophila abortus ELISA produced by the Institut Pourquier which uses a recombinant fragment of the 80–90 kDa protein, were evaluated with the objective to determine whether the new ELISAs would perform as improved alternatives to the complement fixation test (CFT) for the serological diagnosis of ovine enzootic abortion (OEA). The results were compared to those obtained by the CFT and the competitive ELISA (cELISA). The tests were assessed with a panel of 17 serum samples from specific pathogen-free (SPF) lambs experimentally infected with various subtypes of Chlamydophila pecorum, with sera from 45 C. abortus-infected pregnant sheep and from 54 sheep free of OEA. The C. abortus ELISA was identified as being more specific and sensitive than the other tests. The 4 assays were evaluated further with 254 sera from flocks with documented OEA, from flocks with no history of abortion and from animals after abortion of unknown cause. The C. abortus ELISA by the Institut Pourquier identified less OEA-positive sera than the other assays though it identified correctly 9 of 10 OEA-positive flocks. The basis of the discordant results is discussed.

Characterization of serum antibody response to chlamydiae in patients with sexually acquired reactive arthritis.

Sera from patients with sexually acquired reactive arthritis (SARA) with antibodies reacting with C. trachomatis and C. pneumoniae (group 1; n = 20) and also with C. psittaci (group 2; n = 19) were analyzed for antibody specificity. Sera from group 2 reacted significantly more often with C. trachomatis serotype E, H and K and had higher antibody titers to serotype E, as tested by microimmunofluorescence tests. Cross-reactivities occurring in microimmunofluorescence tests were related to the presence of antichlamydial lipopolysaccharide antibodies, adsorption of which by recombinant lipopolysaccharide removed microimmunofluorescence reactivity with C. psittaci antigen. In group 2, significantly more sera had antibodies to C. pneumoniae, remaining after lipopolysaccharide adsorption, as proved by adsorption with viable C. trachomatis and C. pneumoniae organisms. None of the sera had antibodies to Yersinia enterocolitica, Shigella flexneri, Sh. sonnei and Salmonella spp. It was observed that the frequency and titer of cross-reacting antibodies to chlamydial serotypes and species were related to the time period between the diagnosis of genital chlamydial infection and of SARA. Cross-reactivities were also related to the presence of lipopolysaccharide, but not heat shock protein 60- or neutralizing antibodies to chlamydiae. Antibody reactivity induced by antichlamydial lipopolysaccharide antibodies can be removed by lipopolysaccharide adsorption.

The prevalence of avian chlamydiosis (Chlamydophila psittaci) in Bosnia and Hezegovina

In the article are presented the results of our research on chlamydophilosis in parrots, free-living and breeding pigeons, and intensive breeding chickens in Bosnia and Herzegovina. For detection of the antigen two immunoenzyme tests for the detection of antibodies against Chlamydophila psittaci and a complement fixation test by a Kolmer and indirect immunofluorescence method (BioMerieux, France) were used. From a total of 275 samples of cloacal swabs the presence of Chlamydophila psittaci antigen was detected by ELISA (DAKO Ltd., United Kingdom) in 34.9% birds: 45.5% in intensive breeding chickens, 12.1% in free-living pigeons and 8.0% in parrots. By the same method the presence of Chlamydophila psittaci antigen in breeding pigeons was not detected. Sixty cloacal swabs from intensive breeding chickens and pigeons were tested by immunoenzyme test (Unipath Limited, England) and the presence of the pathogen was found in 6.7% cases. Fifty-eight sera from free-living pigeons and intensive breeding chickens were tested for the presence of specific antibodies to Chlamydophila psittaci by indirect immunofluorescence method and were found in 42.1% examined sera of pigeons, and in 27.6% pigeons from the total number of examined birds. The presence of specific antibodies was not found in sera of intensive breeding chickens. Using a complement fixation test, antibodies were not detected in the examined sera in pigeons nor in intensive breeding chickens. The results of this study show that the presence of antigens and antibodies for Chlamydophila psittaci is obvious in tested sera samples, but the clinical disease was not found in any of the examined birds.

Epidemiological investigation of Chlamydophila psittaci in pigeons and free-living birds in Croatia

During 2003, 278 adult pigeons (Columba livia) and 54 birds of 11 other free-living species were caught in the various locations in the City of Zagreb, Croatia. Sera from 182 pigeons were tested for the presence of antibodies against Chlamydophila (C.) psittaci by ELISA test and 174 of them (95.6%) were found positive. Because of the high positivity rate in sera, cloacal swabs of 278 pigeons as well as 54 other species of free-living birds were tested for the presence of C. psittaci antigen. Fourty-four of the 278 pigeons (15.83%) were antigen positive, whereas all 54 of the wild birds were negative. Antigen-positive pigeons were euthanised and examined pathomorphologically and cytologically. Findings of specific antibodies and antigen of C. psittaci confirmed the high rate of infection among urban pigeons in the City of Zagreb, fortunately not among other free-living birds. Although the pigeon serovars of C. psittaci are considered to be of moderate pathogenicity for humans, the identification of 15.8% antigen-positive birds represents a potential source of infection to humans, especially for elderly people and immunodeficient patients, as well as for poultry in the Zagreb city area.

Characterization of serum antibody response to chlamydiae in patients with sexually acquired reactive arthritis

Sera from patients with sexually acquired reactive arthritis (SARA) with antibodies reacting with C. trachomatis and C. pneumoniae (group 1; n=20) and also with C. psittaci (group 2; n=19) were analyzed for antibody specificity. Sera from group 2 reacted significantly more often with C. trachomatis serotype E, H and K and had higher antibody titers to serotypes E, as tested by microimmunofluorescence tests. Cross-reactivities occurring in microimmunofluorescence tests were related to the presence of antichlamydial lipopolysaccharide antibodies, adsorption of which by recombinant lipopolysaccharide removed microimmunofluorescence reactivity with C. psittaci antigen. In group 2, significantly more sera had antibodies to C. pneumoniae, remaining after lipopolysaccharide adsorption, as proved by adsorption with viable C. trachomatis and C. pneumoniae organisms. None of the sera had antibodies to Yersinia enterocolitica, Shigella flexneri, Sh. sonnei and Salmonella spp. It was observed that the frequency and titer of cross-reacting antibodies to chlamydial serotypes and species were related to the time period between the diagnosis of genital chlamydial infection and of SARA. Cross-reactivities were also related to the presence of lipopolysaccharide, but not heat shock protein 60- or neutralizing antibodies to chlamydiae. Antibody reactivity induced by antichlamydial lipopolysaccharide antibodies can be removed by lipopolysaccharide adsorption.

Localization of chronic Chlamydia psittaci infection in the reproductive tract of sheep.

Twelve sheep experimentally infected with Chlamydia psittaci during pregnancy either aborted or gave birth to weak, low-birth-weight lambs as a result of uteroplacental infection. Subsequently, these ewes excreted chlamydial antigen from their reproductive tracts during estrus. About 1 year after pregnancy failure, immunohistochemistry and polymerase chain reaction were used to examine sections of vagina, uterus, and oviduct for evidence of C. psittaci. Four noninfected control ewes were similarly examined. C. psittaci antigen or DNA was detected in vagina, uterus, and oviduct samples from chronically infected ewes. Endometrial cells in the basal stroma were the predominant site of infection. There was no obvious evidence of pathology associated with persistent infection, but increased numbers of plasma cells and intraepithelial lymphocytes were detected in the uterus. C. psittaci is a naturally occurring reproductive infection in sheep that persists following primary infection. Therefore, sheep provide an excellent model to study the host-parasite interactions that occur during natural infection and subsequent persistence.

Pathogenesis of Chlamydia psittaci infection in sheep: Detection of the organism in a serial study of the lymph node

Sixteen sheep were inoculated subcutaneously over the left prefemoral lymph node with an abortifacient strain of Chlamydia psittaci. Groups of four animals were killed after 3, 6, 12 and 18 days. Four of eight sheep which received a control inoculum were killed on day 6 and four on day 18. The left and right prefemoral lymph nodes were removed and weighed and portions taken from each for examination by the polymerase chain reaction (PCR), by culture, and by histopathological and immunohistochemical methods. The left prefemoral lymph nodes enlarged after the injection of C. psittaci, with the group mean weight on day 6 being the greatest and that on day 18 being normal. Examination by "nested" PCR showed samples from these nodes to be positive, except for one animal killed on day 3 and one on day 12. Live organisms, however, were not cultured from any of the samples collected. C. psittaci antigen was detected immunohistochemically in three of four nodes on day 3, in each of four on day 6, and in two of four on both days 12 and 18. Nodes from the contralateral side remained normal, as did those from unchallenged control sheep, and no antigen or DNA was detected in them.

Pathology of bovine abortion and newborn calf death caused by dual infection with Chlamydia psittaci and infectious bovine rhinotracheitis virus.

Nine aborted fetuses and one newborn calf, diagnosed as Chlamydia psittaci (C. psittaci) infection, were pathologically examined. The characteristic lesions in the liver were focal necrosis in 9 aborted fetuses and granulomatous necrotic foci in the newborn calf. Moderate numbers of intranuclear inclusion bodies were found in necrotic foci of the liver, spleen, thymus, lymph nodes, adrenal gland, kidney, lung and forestomach. Immunohistologically, a small number of C. psittaci antigens was demonstrated in necrotic foci of the liver and correlated with distribution of elementary bodies. Moderate numbers of infectious bovine rhinotracheitis (IBR) virus antigens were also detected in degenerating and necrotizing parenchymal cells in various organs and correlated with distribution of intranuclear inclusion bodies. Thus, these aborted bovine fetuses and newborn calf were interpreted as being dually infected with C. psittaci and IBR virus during pregnancy.

Detection of Chlamydia psittaci (Ovis) antigen in tissue sections and McCoy cells using streptavidin-biotin and the imagen ™ staining method

Two procedures for identifying chlamydial antigen in infected McCoy cells and tissue sections are described. Both the streptavidin-biotin method and the IMAGEN test clearly detected chlamydial antigen in ovine placental tissue sections and in infected McCoy cells before they could be detected by conventional stains. The streptavidin-biotin method is lengthy, but specific and sensitive, and the slides can be kept indefinitely for later examination. By contrast, the IMAGEN test is a single step procedure which requires fluorescent microscopy and slides can only be kept for up to 1 week before examination.

Purification of chlamydia psittaci antigen by affinity chromatography on polymyxin B agarose for use in the enzyme-linked immunosorbent assay (ELISA)

The purification of cell wall antigens of Chlamydia psittaci by affinity chromatography on polymyxin B agarose is described. Chlamydial cell wall antigens were prepared using different methods: heat treatment, ultrasonication and sodium deoxycholate treatment. The antigens were subsequently purified by gel chromatography. The highest amount of cell wall antigens was obtained by heat treatment of the chlamydiae at 90 degrees C and pH 8.5. The purified antigens showed molecular weights of 450 kDa to 700 kDa. Treatment of heat-extracted antigens with trypsin, pronase E or proteinase K did not destroy antigenic activity or alter the molecular weight. On the other hand, potassium metaperiodate treatment led to a significant decrease of both. Chlamydial cell wall antigens extracted by heat treatment and purified by affinity chromatography were used as ELISA-antigen. Using this ELISA and the complement fixation test (CFT), a total of 576 bovine sera was tested. 92 of these sera reacted positively in the ELISA and only 13 in the CFT. In addition, the prepared ELISA was compared with a commercial ELISA. 12 out of 15 sera tested were positive when using the ELISA described and 10 were positive when using the commercial ELISA.

Preparation and use of a monoclonal antibody to detect Chlamydia psittaci antigen in paraffin-embedded tissue sections

A murine monoclonal antibody prepared against an ovine abortion isolate of Chlamydia psittaci (A22/Teramo) revealed specific binding to a 57 kDa chlamydial antigen in immunoblotting studies. The monoclonal antibody was able to detect intracytoplasmic chlamydial inclusions and scattered elementary bodies in infected McCoy cell culture, and on formalin-fixed paraffin-embedded tissue sections both from experimentally infected mice and from fetal membranes of cases of ovine enzootic abortion.

Development of serodiagnostic kit "HITAZYME Chlamydia Ab" for Chlamydia trachomatis infections using extracted antigen

To develop EIA kit with low cross-reactivity for the quantitative detection of anti-chlamydial antibodies, we examined the preparation of trachomatis antigens and its specificity to mouse antisera and human sera. The chlamydial elementary body (EB) purified from C. trachomatis L2/434/Bu strain was treated by Sarkosyl, dithiothreitol and SDS by turns to obtain the soluble EB outer membrane (COMC). SDS-PAGE showed that the major components of the COMC were 96K, 60K and 39.5 KDa peptides. The reactivity of the COMC immobilized to 96 wells microtiter plate to mouse anti-serum to C. trachomatis was higher than the other two mouse anti-sera to C. psittaci and pneumoniae. In human sera, the cut off values were calculated from an average optical density plus its two-fold standard deviation obtained by the testing of 100 samples of healthy human sera. We evaluated the specificity of the kit to 17 anti-C. pneumoniae, 9 C. trachomatis and 4 C. psittaci antibodies positive patients' sera judged by the MFA method respectively. The results showed that the concordance ratio of IgG and IgA were 88%, 100% in anti-C. pneumoniae, 89%, 78% in anti-C. trachomatis and 50%, 50% in anti-C. psittaci respectively. From the results obtained in this study, we concluded that the HITAZYME method which had a very low cross-reactivity to C. pneumoniae is clinically useful in the serodiagnosis of C. trachomatis infections, even if it has a little common antigenicity with C. psittaci antigen.

Serological assessment of chlamydial infection in the koala by a slide EIA technique

A rapid and simplified slide enzyme immunosorbent assay (EIA) was developed for the diagnosis of chlamydial infection in the koala. HeLa 229 cells infected with koala strain Chlamydia psittaci were fixed on the surface of multiwell slides and used as the antigen. The assay consisted of first reacting koala antiserum with the fixed C psittaci antigen, followed by reaction with biotinylated rabbit anti-koala IgG, ABC reagent and substrate. The chlamydial EIA antibody titres obtained were compared with those of a complement fixation (CF) test using koala strain C psittaci as antigen. Of 35 koala sera tested, 16 CF positive sera (greater than or equal to 1:8) also had a positive titre (greater than or equal to 1:200) in the slide EIA test (sensitivity 93.8%, 15/16). Nineteen CF negative sera were also negative in the slide EIA (specificity 100%, 19/19). Sixty-eight samples of koala blood were collected by ear-prick using a sampling paper method and were assayed by both tests. Sensitivity of the slide EIA was 100% (15/15) and specificity of the test was 96.2% (51/53). To simplify the slide EIA for use as a practical screening test, a 3-point serum dilution series (1:100, 1:200, 1:400) was used. This 3-point slide EIA was compared with the CF test using sheep strain chlamydial antigen. Thirty-nine sera were assayed by both tests. The sensitivity of the 3-point method was 85.7% (6/7) and the specificity was 71.9% (23/32) as compared with the sheep antigen CF test.(ABSTRACT TRUNCATED AT 250 WORDS)