Feline Alpha-1-acid glycoprotein (AGP)

Feline Alpha-1-acid glycoprotein (AGP)

Andy Pachikerl, Ph.D

Introduction

Alpha-1-acid glycoprotein (AGP) surges in cats’ blood when they fall in victim of feline infectious peritonitis (FIP), a lethal disease caused by feline coronavirus (FCoV). The diagnosis of feline infectious peritonitis (FIP) is often tough and not very viable at times. The clinical suspicion of FIP might be supported by the detection of effusions if any are present (Hartmann, et al., 2003; Paltrinieri, Parodi, & Cammarata, In Vivo Diagnosis of Feline Infectious Peritonitis by Comparison of Protein Content, Cytology, and Direct Immunofluorescence Test on Peritoneal and Pleural Effusions, 1999). The only way to conclusively test FIP to be positive is through histology followed by immunohistochemical or immunofluorescent detection of feline coronavirus (FCoV) within intralesional macrophages (Addie, Paltrinieri, & Pedersen, 2004; Barlough & Stoddart, 1990). Various studies suggested the implementation of biopsies for the confirmation of FIP in veterinary practices and it turns out to be quite useful with just a few downside (Alessia, Paltrinieri, Bertazzolo, Milesi, & Parodi, 2005).

The application of biopsies in vivo is usually restricted due to anaesthetic risks especially via surgical biopsy and the relatively high percentage of unsuitable or falsely negative tru-cut or fine-needle aspiration biopsies (Alessia, Paltrinieri, Bertazzolo, Milesi, & Parodi, 2005). Serology and polymerase chain reaction techniques are not suitable for FIP diagnosis because they do not differentiate between the widespread low pathogenic FCoVs and the mutant pathogenic FCoV strains (Addie, Paltrinieri, & Pedersen, 2004; Herrewegh, et al., 1997). A previous study (Stoddart, Whicher, & Harbour, 1988) reported high levels of a1-acid glycoprotein (AGP) in cats with experimentally induced FIP. This finding was confirmed by another study, (Duthie, Eckersall, Addie, Lawrence, & Jarrett, 1997) which proposed the possible use of serum AGP as a diagnostic tool for FIP. Serum AGP is now widely used in diagnostic profiles for FIP.1 However, serum AGP levels increase in inflammatory disorders other than FIP (Duthie, Eckersall, Addie, Lawrence, & Jarrett, 1997; Kajikawa, Furuta, Onishi, Tajima, & Sugii, 1999; TerWee, Lauritzen, Sabara, Dreier, & Kokjohn, 1997; TerWee, et al., 1998), neoplasia (Correa, Mauldin, Mauldin, & Mooney, 2001), and asymptomatic but FCoV-positive cats. This lack of specificity limits the diagnostic potential of serum AGP as a diagnostic test for FIP. For more information on FIP and FCoV can be found in previously published report article (link).

FIP clinical diagnosis through feline AGP

AGP has been used extensively, particularly in Europe, as an indicator test for FIP. AGP was found almost a decade ago to be hyposialylated in cats with FIP, but not in normal cats or in cats with other pathologies (Fabrizio, Claudia, Alessia, Vanessa, & Saverio, 2004). This study confirmed that serum AGP is a powerful discriminating marker for FIP, but only when coupled with other high risk factors (Saverio, Giordano, Tranquillo, & Guazzetti, 2007). A Bayesian approach demonstrated that, when the pre-test probability of FIP was high based on history and clinical signs, moderate serum AGP levels (1.5–2 μg / mL) could discriminate cats with FIP from others. However, only high serum AGP levels (>3 μg / mL) were highly suggestive of FIP in cats with a low pre-test probability of disease (Saverio, Giordano, Tranquillo, & Guazzetti, 2007).

Giori, et al. (2011) had shown specificity and sensitivity of several tests in 12 cats, four of which have the absence of FIP via histopathology and immunohistochemistry, and eight cats with FIP confirmed via histopathology and immunohistochemistry. Results from serum protein electrophoresis, analysis of effusions, anti-feline coronavirus serology, serum AGP concentrations and histopathology were then compared with the confirmed diagnosis. No concordance was found for serology and analysis of effusions, poor concordance was noted for histopathology, fair concordance for serum electrophoresis and perfect concordance for AGP. Their study proved that immunohistochemistry is always required to confirm FIP and, if immunohistochemistry is not feasible, they concluded that histopathology is not definitive, whilst elevated AGP concentrations might support the diagnosis of FIP. However, the small numbers of cats in this study make it difficult to validate such conclusions and the earlier study of Saverio, et al. (2007) is probably a more accurate assessment of AGP testing for FIP. Like most indirect tests for FIP, the positive predictive value increases with the number of other risk factors that are present.

Saverio, et al. (2007) also investigated the levels of leukocyte bound AGP in normal cats and cats with diseases including FIP by flow cytometry using an anti-feline AGP antibody. A total of 32 healthy cats (19 feline coronaviruses seropositive), 13 cats with FIP (presumably all coronavirus seropositive) and 12 cats with other diseases (six coronaviruses seropositive) were studied. The proportion of cats with AGP-positive leucocytes in each group or in cats with different intensities of inflammatory response (as measured by CBC, serum electrophoresis and serum AGP levels) was compared. AGP positive leucocytes were found in 23% of cats; most were diseased, but a small number were healthy. AGP positive leukocyte staining was associated with inflammation and not with leucocytosis per se. Staining among healthy cats was unrelated to coronavirus antibody status. Cats with FIP were more likely to have positive staining leukocytes than healthy cats, but not as likely as cats with other diseases. It was concluded that AGP positive leucocytes are present in feline blood, especially during inflammation. Staining leukocytes for AGP binding do not appear to have any value over serum AGP testing, especially when considering the potential cost and effort involved in this method.

A previous study by Paltrinieri, et al. (2007) showed positive correlation between AGP and FIP cats. In their study, they used 2 different groups of cats with FIP or non-FIP along with others contracted with other diseases such as FCoV. Their schematic experiment is as follows:

Group 1. FIP group. This group was composed of 58 cats that had clinical signs and laboratory findings confirmatory of effusive FIP (n 5 53) or dry FIP (n 5 5). Haematology and serum biochemistry in these cats revealed nonregenerative anaemia, neutrophilia, lymphopenia, increased total proteins, and total-, a2- and c-globulins. Cytology and protein analysis of the effusion in the effusive cases supported the diagnosis of FIP. All the cats were serologically negative for feline immunodeficiency virus (FIV) and feline leukaemia virus (FeLV) infection. Anti-FCoV serology, performed by an immunofluorescence test produced at the University of Zurich according to Osterhaus et al., was positive in 40 of the 44 cats in which this test was performed. All the suspected dry forms and 46 effusive cases had the diagnosis confirmed by necropsy, histological, and immunohistochemical findings. In the 7 effusive cases in which necropsy was not performed, FIP was confirmed using immunofluorescence staining of cells obtained by cytocentrifuging the effusions.

Group 2. Non-FIP group. This group was composed of 104 cats in which the diagnosis of FIP was definitively excluded. Based on the clinic-pathological findings, the clinical records, and the follow-up, these cats were further divided into the following subgroups:

Group 2a. Inflammatory processes other than FIP. This subgroup was composed of cats (n 5 26) in which there was a clinical suspicion of FIP, based on clinical signs consistent with the wet (n 5 6) or dry (n 5 20) form of the disease. Specifically, the clinical suspicion of FIP was due to the presence of intracavitary effusions in 6 cats. Cytology and or post-mortem examination in these cats excluded the diagnosis of FIP and led to the final diagnosis of epithelial tumours (n 5 4), lymphoma (n 5 1), and cholangioepathitis (n 5 1). In the remaining 20 cases, the clinical suspicion of FIP was due to the presence of fever (n 5 14) or neurological signs (n 5 6). Nine of the 14 febrile cats recovered after appropriate antibiotic therapy and were still alive 2 to 5 yr. later, thus excluding a diagnosis of FIP. The remaining 5 febrile cats developed gastrointestinal signs consistent with feline panleukopenia, confirmed by post-mortem examination. Of the 6 cats with neurological signs, a diagnosis of toxoplasmosis (confirmed by positive serology and response to specific therapy) was made in 4 cats, which recovered from neurological signs and were still asymptomatic 3 to 5 yr. later; in the remaining 2 cats, intracranic tumours were confirmed at necropsy.

Group 2b. Asymptomatic FCoV infection (n 5 49). These were FCoV-positive, FIV-negative, and FeLV negative cats living in breeding catteries known to have a high prevalence of FIP. All these cats were asymptomatic at the time of serum sampling and remained asymptomatic over a follow-up period ranging from 18 mo. to 5 yr.

Group 2c. Injection site sarcoma (n 5 19). These cats were sampled before surgical removal of feline injection site sarcoma as part of another study.

Group 2d. Postvaccination (n 5 7). These were clinically healthy shelter cats aged approximately 3 mo. and were serologically negative for FIV, FeLV, and FCoV (n 5 7). Serum was collected from these cats approximately 20 days after vaccination for feline panleukopenia, calicivirus, and herpesvirus A.

Group 2e. Specific pathogen–free (SPF) cats. Serum samples were collected from 3 clinically healthy, untreated SPF cats.

Rivalta test

The Rivalta test is widely touted, especially in Europe, and has been long used for diagnosing FIP-associated exudates (Hartmann, et al., 2003). The test involves placing a few drops of ascites or thoracic fluid into a tube containing a weak acetic acid solution. The appearance of a white flocculent material is seen in a positive test. A positive Rivalta test was once believed to be highly specific for FIP fluid. In a study of 497 cats with effusions, 35% of which had confirmed FIP, the Rivalta test had a sensitivity of 91% and a specificity of 66%, with a positive predictive value of 58% and a negative predictive value of 93% (Fischer, Sauter‐Louis, & Hartmann, Diagnostic accuracy of the R ivalta test for feline infectious peritonitis, 2012). As would be expected, these values increased when cats with lymphosarcoma or bacterial infections were excluded, or when only cats of ≥2 years of age were considered. The Rivalta test appears to be reproducible in samples stored for 21 days at room, refrigerator or freezer temperatures, and with some modifications of acid concentration (Fischer, Weber, Sauter-Louis, & Hartmann, 2013). However, reading of the test is subjective and results are therefore somewhat dependent on the evaluator.

Reference

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