Comparison of the Gimenez Staining Method and Antigen Detection ELISA with Culture for Detecting Chlamydiae in Birds

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, and culture of multiple sample types from birds may increase the sensitivity. 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 have allowed development of an enzyme-linked immunosorbent assay (ELISA) for rapid detection. Two commercially available kits have been evaluated for use in birds. Microscopic examination of impressions of uncut surfaces of bird organ tissues and inoculated cell culture monolayers using the Gimenez staining method (GSM) also was reported as a rapid diagnostic procedure for examining dead birds. 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.