Markers of inflammation in free‐living African elephants (Loxodonta africana): Reference intervals and diagnostic performance of acute phase reactants

Abstract

Introduction Acute phase reactants (APRs) have not been investigated in free‐living African elephants (Loxodonta africana), and there is little information about negative APRs albumin and serum iron in elephants. Objectives We aimed to generate reference intervals (RIs) for APRs for free‐living African elephants, and to determine the diagnostic performance of APRs in apparently healthy elephants and elephants with inflammatory lesions. Methods Stored serum samples from 49 apparently healthy and 16 injured free‐living elephants were used. The following APRs and methods were included: albumin, bromocresol green; haptoglobin, colorimetric assay; serum amyloid A (SAA), multispecies immunoturbidometric assay, and serum iron with ferrozine method. Reference intervals were generated using the nonparametric method. Indices of diagnostic accuracy were determined by receiver‐operator characteristic (ROC) curve analysis. Results Reference intervals were: albumin 41–55 g/L, haptoglobin 0.16–3.51 g/L, SAA < 10 mg/L, and serum iron 8.60–16.99 μmol/L. Serum iron and albumin concentrations were lower and haptoglobin and SAA concentrations were higher in the injured group. Serum iron had the best ability to predict health or inflammation, followed by haptoglobin, SAA, and albumin, with the area under the ROC curve ranging from 0.88–0.93. Conclusions SAA concentrations were lower in healthy African vs Asian elephants, and species‐specific RIs should be used. Serum iron was determined to be a diagnostically useful negative APR which should be added to APR panels for elephants.

Acute phase reactants in nondomesticated mammals—A veterinary clinical pathology perspective

“…Performance goals derived from estimates of biological variation are available for selected APRs in dogs, and total allowable error based on analytical performance and consensus between the two authors of this review has been suggested for SAA (Table 5). 34,49 Although not ideal, these could be useful when evaluating validation results for APR assays in zoo and wildlife species 34,36 …”

“…It has been recommended that 20–30 animals be included in each group, and that the presence or absence of inflammation is clear, based on clinical signs, other laboratory testing, or postmortem findings 10 . Evaluation of overlap performance is commonly performed in nondomesticated mammal APR studies, as noted in Tables 1‐4 13,14,16,22,23,32–37,39,40,42–46,50 . We recommend that to optimize this stage, clinical pathologists work closely with clinicians to gather information about the nature of the putative inflammatory disease and sample handling, such as sample type, storage conditions, number of freeze‐thaw cycles, diagnosis, chronicity of disease, a sample taken before or during treatment, availability of repeated samples from one animal.…”

“…A colorimetric peroxidase assay (PHASE Haptoglobin, Tridelta Ireland) is commonly used as it has the ability to measure haptoglobin across a range of species, and has been partially validated for use in bongo, capybara, red deer, bottlenose dolphin, European bison, elephants, Iberian ibex, koala, Pronghorn antelope, Rhesus macaques, white rhinoceros, and Grant's zebra (Table 2). 14,22,30,32–34,36–40,42,46 This assay shows good linearity and repeatability, but reproducibility did not meet performance goals in some studies 34,58 . A major function of haptoglobin is to bind hemoglobin, and in vitro hemolysis in samples will cause falsely low results for haptoglobin as the protein is bound up in haptoglobin‐hemoglobin complexes.…”

“…Iron is a negative APR that has shown a good ability to discriminate between health and inflammation in domesticated animals like horses and cattle 72–75 . Serum iron has been measured in several nondomesticated mammals; results from studies that included iron measurement in the context of inflammatory disease can be found in Table 4 23,33,34,36,76 . Serum iron measurement is available on most automated wet chemistry systems, using the spectrophotometric ferrozine zinc method.…”

“…Serum iron measurement is available on most automated wet chemistry systems, using the spectrophotometric ferrozine zinc method. Decreases in serum iron in animals with inflammatory disease compared with apparently healthy groups were investigated and found to be present in African elephants, white rhinoceros, and Rhesus macaques 33,34,36 …”