The development and use of a stained chlamydial elementary body agglutination (EBA) antigen for detecting antibody activity in avian sera is described. Examples of serologic results on serum samples from various types of birds indicate the usefulness of EBA, latex agglutination (LA), and direct complement fixation (DCF) in diagnosing avian chlamydiosis. Results of tests on 10 cockatiels examined in clinics indicate that a combination of serology, culture, and/or antigen-detection enzyme-linked immunosorbent assay may be helpful when testing this type of bird. Agreement between EBA, LA, and DCF was 81.8% when testing 407 serum samples from cockatiels of unknown health status. The relationship between positive (> or = 10 titer) antibody activity and known health status of 77 cockatiels revealed that agreement between the two criteria was only 59.7%. Of 13 Chlamydia-inoculated cockatiels, seven birds seroconverted from negative to positive by EBA; five seroconverted by DCF. Only the five birds that seroconverted by both EBA and DCF were culture-positive for chlamydiae. None of 15 sham-inoculated control cockatiels developed detectable antibody activity, and none of 10 cultured were positive. In tests with column-separated IgM and IgG, EBA detected only IgM activity, LA detected IgM and IgG activity, and DCF detected only IgG activity.

No abstract

Chlamydiosis, caused by Chlamydia psittaci, an obligate intracellular bacterium, is a serious health concern for various types of birds and their owners. Disease can occur as a chronic inapparent, subacute, acute, or peracute infection.Culture for chlamydiae is considered the "gold standard" for evaluating other methods, even though it probably does not have 100% sensitivity nor 100% specificity in live human subjects, 9 and culture of multiple sample types from birds may increase the sensitivity. 1 Multiple cultures may increase chances of recovering chlamydiae from live birds, but it probably is not advisable to delay treatment of birds critically ill with signs consistent with chlamydiosis.Monoclonal antibodies that detect chlamydial lipopolysaccharide 3 have allowed development of an enzyme-linked immunosorbent assay (ELISA) for rapid detection. 6 Two commercially available kits have been evaluated for use in birds. 1,7 Microscopic examination of impressions of uncut surfaces of bird organ tissues and inoculated cell culture monolayers using the Gimenez staining method 3 (GSM) also was reported as a rapid diagnostic procedure for examining dead birds. 4 The purpose of this study was 2-fold: 1) to expand the comparison of 1 commercially available ELISAB with chlamydial isolation attempts from live birds and 2) to compare results of antigen-detection ELISA and the GSM with culture results on dead birds.Samples used in this study were submitted to the Texas Veterinary Medical Diagnostic Laboratory (TVMDL) for chlamydial testing. Cloacal swabs, choanal slit swabs, and fecal samples from live birds were submitted in chlamydial transport medium. Some fresh fecal samples also were submitted.Fresh fecal material was placed into chlamydial transport medium to make an approximate 20% (w:v) suspension, which was centrifuged at 1,000 x g for 15 minutes and then diluted 1:2 for isolation attempts. For the ELISA testing, 1.0 ml of fecal suspension was centrifuged at 3,000 x g for 20 minutes, and the supematant fluid was discarded. A swab was then used to collect the pellet.Visceral and other organs from dead birds necropsied by referring veterinarians or TVMDL personnel were processed for the GSM, ELISA, and isolation procedures. Impressions were most often made of liver and spleen uncut surfaces and occasionally of the pericardium, air sacs, or lungs. These impressions were stained by the GSM and examined microscopically. 4,5 Suspensions (20%, w:v) of pooled organ tissues for cell monolayer inoculation were made in cell culture growth medium and centrifuged for 15 minutes at 1,000 x g. For ELISA of tissues, swabs were inserted into spleen and liver and then air dried.

Abstract.A 2 x 2 contingency table was constructed to demonstrate the relationships between detectable Chlamydial antibody activity and clinical health status of tested birds. The table revealed that 65.5% of clinically ill birds were antibody positive by elementary body agglutination (EBA) (≥10 titers) and 59.0% were antibody positive by latex agglutination (LA). Thus, EBA was slightly more sensitive than LA in detecting antibody activity. Of the clinically normal birds, 96.7% were antibody negative (< 10 titers) by EBA and 98.3% were antibody negative by LA. Individual serum or plasma samples from a group of mixed types of psittacine birds and cockatiels were tested as a separate group, and relationships between EBA-detectable antibody activity and health status were obtained from a 2 x 2 contingency table. Sixty-six percent of birds clinically ill with signs of chlamydiosis in the mixed-type group were antibody positive, whereas only 32.3% of clinically ill cockatiels were antibody positive. Statistical analysis of the contingency table using a chi-square test demonstrated that the EBA test differentiates between individual birds on the basis of health status (P < 0.001). When testing paired serum or plasma samples by EBA, LA, and direct complement fixation (DCF), the highest percentage of significant (≥ 4-fold change) titer decreases was detected by LA, and the highest percentage of significant titer increases was detected by DCF. Examples of EBA, LA, and DCF titers in paired and multiple serum or plasma samples are presented to show the variety of responses that can occur. Results reflected variations seen in individual testing of birds with titer variability seen in the first sample tested. Additional types of testing believed necessary for confirming or ruling out an infectious process in birds are outlined. The current interpretations of serologic results are given.

Latex agglutination detection of chlamydial antibody activity in psittacine bird sera was significantly more sensitive when an improved protocol was followed than was a test using the previously used protocol. Titers of antibody-positive sera were fourfold to 32-fold higher by the improved protocol than titers by the previously used protocol, whereas antibody-negative sera were negative by both protocols. Column chromatography was used to separate immunoglobulins in psittacine bird serum. Immunoglobulin M was reactive in latex agglutination (LA) but non-reactive in direct complement fixation (DCF). Immunoglobulin G was reactive in LA and DCF. The fifth supernatant fluid of serum multiply absorbed with latex beads was non-reactive in LA and DCF. The immunoglobulins reactive in LA had high avidity and affinity for latex beads. Prolonged storage at 4 C to 6 C preserved LA reactivity of absorbed immunoglobulins better than storage at 21 C to 23 C.