What If The Antigen Test Is Negative?

Publications suggest the sensitivity for Histoplasma or Blastomyces antigen in urine approaches 100% but the results of clinical testing indicate negative results occur perhaps in 5-10% of cases. The causes for false negative results include limited fungal burden which occurs in cases with focal lung lesions, skin lesions, ocular lesions, etc. Negative results may also occur in the first few weeks following onset of symptoms.

ANSWER: I recommend sending urine and serum with instructions to test the urine for either Histoplasma or Blastomyces antigen (Primary tests). There is no need to test for both Histoplasma and Blastomyces antigen since they are cross-reactive. If the primary tests are negative, perform the secondary tests, which may help with diagnosis. If the illness has been present for a month or less submit a convalescent specimen, which may have become positive with progressive infection. If the antigen and antibody tests are negative diagnosis may be established by cytopathology or histopathology of lesions or body fluids.

Recommendations Based on Suspected Diagnosis

Mycosis Primary (most sensitive) Secondary (may be positive if primary tests negative)
Blastomycosis [1Mourning AC, Patterson EE, Kirsch EJ, et al. Evaluation of an enzyme immunoassay for antibodies to a recombinant Blastomyces adhesin-1 repeat antigen as an aid in the diagnosis of blastomycosis in dogs. J Am Vet Med Assoc 2015 Nov 15; 247(10):1133-8., 2Foy DS, Trepanier LA, Kirsch EJ, Wheat LJ. Serum and urine blastomyces antigen concentrations as markers of clinical remission in dogs treated for systemic blastomycosis. J Vet Intern Med 2014 Mar; 28(2):305-10.] Blastomyces urine antigen (code 316) Blastomyces IgG antibody-canine (code 330)
Blastomyces FID antibody-feline (code 328)
[feline IgG not available]
Histoplasmosis [3Hanzlicek AS, Meinkoth JH, Renschler JS, Goad C, Wheat LJ. Antigen Concentrations as an Indicator of Clinical Remission and Disease Relapse in Cats with Histoplasmosis. J Vet Intern Med 2016 Jul; 30(4):1065-73., 4Cunningham L, Cook A, Hanzlicek A, et al. Sensitivity and Specificity of Histoplasma Antigen Detection by Enzyme Immunoassay. J Am Anim Hosp Assoc 2015 Sep; 51(5):306-10.] Histoplasma urine antigen (code 310) Histoplasma IgG antibody (code 327)
Histoplasma FID antibody (code 321)
Histoplasma serum antigen (test code 310)
Coccidioidomycosis [5Kirsch EJ, Greene RT, Prahl A, et al. Evaluation of coccidioides antigen detection in dogs with coccidioidomycosis. Clin Vaccine Immunol 2012 Mar; 19(3):343-5., 6Holbrook ED, Greene RT, Rubin SI, et al. Novel canine anti-Coccidioides immunoglobulin G enzyme immunoassay aids in diagnosis of coccidioidomycosis in dogs. Med Mycol 2019 Jan 12.] Coccidioides IgG antibody (code 329)
Coccidioides FID antibody (code 320)		 Coccidioides serum and urine antigen (code 315)
Cryptococcosis [7Vorathavorn VI, Sykes JE, Feldman DG. Cryptococcosis as an emerging systemic mycosis in dogs. J Vet Emerg Crit Care (San Antonio ) 2013 Sep; 23(5):489-97.] Cryptococcal serum antigen (code 319) None
Aspergillosis-sinonasal [8Billen F, Peeters D, Peters IR, et al. Comparison of the value of measurement of serum galactomannan and Aspergillus-specific antibodies in the diagnosis of canine sino-nasal aspergillosis. Vet Microbiol 2009 Feb 2; 133(4):358-65., 9Barrs VR, Ujvari B, Dhand NK, et al. Detection of Aspergillus-specific antibodies by agar gel double immunodiffusion and IgG ELISA in feline upper respiratory tract aspergillosis. Vet J 2015 Mar; 203(3):285-9.] Aspergillus FID antibody (code 324) Aspergillus serum and urine antigen (code 309)
Aspergillosis-systemic [10Garcia RS, Wheat LJ, Cook AK, Kirsch EJ, Sykes JE. Sensitivity and specificity of a blood and urine galactomannan antigen assay for diagnosis of systemic aspergillosis in dogs. J Vet Intern Med 2012 Jul; 26(4):911-9.] Aspergillus serum and urine antigen (code 309) Aspergillus urine antigen (code 309)
Aspergillus FID antibody (code 324)

Sensitivity and Specificity MiraVista Tests

Test Specimen Sensitivity Specificity Reference
Blastomyces antigen Urine (dog)
Serum (cat)		 100%
100%		 95%
100%		 [1Mourning AC, Patterson EE, Kirsch EJ, et al. Evaluation of an enzyme immunoassay for antibodies to a recombinant Blastomyces adhesin-1 repeat antigen as an aid in the diagnosis of blastomycosis in dogs. J Am Vet Med Assoc 2015 Nov 15; 247(10):1133-8.]
Blastomyces IgG antibody EIA
Blastomyces ID antibody
Blastomyces ID antibody		 Serum (dog)
Serum (dog)
Serum (dog)		 95%
65%
18%		 95%
100%
Not done		 [1Mourning AC, Patterson EE, Kirsch EJ, et al. Evaluation of an enzyme immunoassay for antibodies to a recombinant Blastomyces adhesin-1 repeat antigen as an aid in the diagnosis of blastomycosis in dogs. J Am Vet Med Assoc 2015 Nov 15; 247(10):1133-8.]
[1Mourning AC, Patterson EE, Kirsch EJ, et al. Evaluation of an enzyme immunoassay for antibodies to a recombinant Blastomyces adhesin-1 repeat antigen as an aid in the diagnosis of blastomycosis in dogs. J Am Vet Med Assoc 2015 Nov 15; 247(10):1133-8.]
[11Spector D, Legendre AM, Wheat J, et al. Antigen and antibody testing for the diagnosis of blastomycosis in dogs. J Vet Intern Med 2008 Jul; 22(4):839-43.]
Histoplasma antigen
Histoplasma antigen		 Urine (dog)
Urine (cat)		 89%
94%		 100%
100%		 [4Cunningham L, Cook A, Hanzlicek A, et al. Sensitivity and Specificity of Histoplasma Antigen Detection by Enzyme Immunoassay. J Am Anim Hosp Assoc 2015 Sep; 51(5):306-10.]
[12Cook AK, Cunningham LY, Cowell AK, Wheat LJ. Clinical evaluation of urine Histoplasma capsulatum antigen measurement in cats with suspected disseminated histoplasmosis. J Feline Med Surg 2012 Aug; 14(8):512-5.]
Histoplasma IgG antibody EIA
Histoplasma IgG antibody EIA
Histoplasma ID antibody		 Serum (dog)
Serum (cat)
Serum (human)		 78%
78%
88%		 97%
84%
99%		 MVD
MVD
[13Wheat J, French ML, Kohler RB, et al. The diagnostic laboratory tests for histoplasmosis: analysis of experience in a large urban outbreak. Ann Intern Med 1982 Nov; 97(5):680-5.]
Coccidioides antigen
Coccidioides antigen		 Serum (dog)
Urine (dog)		 30%
12%		 97% [6Holbrook ED, Greene RT, Rubin SI, et al. Novel canine anti-Coccidioides immunoglobulin G enzyme immunoassay aids in diagnosis of coccidioidomycosis in dogs. Med Mycol 2019 Jan 12.]
Coccidioides IgG antibody EIA
Coccidioides ID antibody
Combined IgG and ID antibodies		 Serum (dog)
Serum (dog)
Serum (dog)		 89%
92%
99%		 94%
92%		 [6Holbrook ED, Greene RT, Rubin SI, et al. Novel canine anti-Coccidioides immunoglobulin G enzyme immunoassay aids in diagnosis of coccidioidomycosis in dogs. Med Mycol 2019 Jan 12.]
Cryptococcal antigen (CALAS) Serum (cat)
Serum (dog)		 96%
83%		 97-100% [14Sykes JE. Canine and Feline Infectious Diseases, St. Louis, MO, Elsevier Saunders. 2019., 15Trivedi SR, Sykes JE, Cannon MS, et al. Clinical features and epidemiology of cryptococcosis in cats and dogs in California: 93 cases (1988-2010). J Am Vet Med Assoc 2011 Aug 1; 239(3):357-69., 16Trivedi SR, Malik R, Meyer W, Sykes JE. Feline cryptococcosis: impact of current research on clinical management. J Feline Med Surg 2011 Mar; 13(3):163-72.]
Aspergillus antigen-systemic aspergillosis Serum (dog)
Urine (dog)		 92%
88%		 86%
92%		 [10Garcia RS, Wheat LJ, Cook AK, Kirsch EJ, Sykes JE. Sensitivity and specificity of a blood and urine galactomannan antigen assay for diagnosis of systemic aspergillosis in dogs. J Vet Intern Med 2012 Jul; 26(4):911-9.]
Aspergillus antigen-sinonasal aspergillosis Serum (dog) 24% 76% [8Billen F, Peeters D, Peters IR, et al. Comparison of the value of measurement of serum galactomannan and Aspergillus-specific antibodies in the diagnosis of canine sino-nasal aspergillosis. Vet Microbiol 2009 Feb 2; 133(4):358-65.]
Aspergillus antibody
ID-sinonasal aspergillosis		 Serum (dog) 76% 100% [8Billen F, Peeters D, Peters IR, et al. Comparison of the value of measurement of serum galactomannan and Aspergillus-specific antibodies in the diagnosis of canine sino-nasal aspergillosis. Vet Microbiol 2009 Feb 2; 133(4):358-65.]
Fungitell beta-D-glucan Serum (dog) 100% 72% MVD

REFERENCES:

  1. Mourning AC, Patterson EE, Kirsch EJ, et al. Evaluation of an enzyme immunoassay for antibodies to a recombinant Blastomyces adhesin-1 repeat antigen as an aid in the diagnosis of blastomycosis in dogs. J Am Vet Med Assoc 2015 Nov 15; 247(10):1133-8.
  2. Foy DS, Trepanier LA, Kirsch EJ, Wheat LJ. Serum and urine blastomyces antigen concentrations as markers of clinical remission in dogs treated for systemic blastomycosis. J Vet Intern Med 2014 Mar; 28(2):305-10.
  3. Hanzlicek AS, Meinkoth JH, Renschler JS, Goad C, Wheat LJ. Antigen Concentrations as an Indicator of Clinical Remission and Disease Relapse in Cats with Histoplasmosis. J Vet Intern Med 2016 Jul; 30(4):1065-73.
  4. Cunningham L, Cook A, Hanzlicek A, et al. Sensitivity and Specificity of Histoplasma Antigen Detection by Enzyme Immunoassay. J Am Anim Hosp Assoc 2015 Sep; 51(5):306-10.
  5. Kirsch EJ, Greene RT, Prahl A, et al. Evaluation of coccidioides antigen detection in dogs with coccidioidomycosis. Clin Vaccine Immunol 2012 Mar; 19(3):343-5.
  6. Holbrook ED, Greene RT, Rubin SI, et al. Novel canine anti-Coccidioides immunoglobulin G enzyme immunoassay aids in diagnosis of coccidioidomycosis in dogs. Med Mycol 2019 Jan 12.
  7. Vorathavorn VI, Sykes JE, Feldman DG. Cryptococcosis as an emerging systemic mycosis in dogs. J Vet Emerg Crit Care (San Antonio ) 2013 Sep; 23(5):489-97.
  8. Billen F, Peeters D, Peters IR, et al. Comparison of the value of measurement of serum galactomannan and Aspergillus-specific antibodies in the diagnosis of canine sino-nasal aspergillosis. Vet Microbiol 2009 Feb 2; 133(4):358-65.
  9. Barrs VR, Ujvari B, Dhand NK, et al. Detection of Aspergillus-specific antibodies by agar gel double immunodiffusion and IgG ELISA in feline upper respiratory tract aspergillosis. Vet J 2015 Mar; 203(3):285-9.
  10. Garcia RS, Wheat LJ, Cook AK, Kirsch EJ, Sykes JE. Sensitivity and specificity of a blood and urine galactomannan antigen assay for diagnosis of systemic aspergillosis in dogs. J Vet Intern Med 2012 Jul; 26(4):911-9.
  11. Spector D, Legendre AM, Wheat J, et al. Antigen and antibody testing for the diagnosis of blastomycosis in dogs. J Vet Intern Med 2008 Jul; 22(4):839-43.
  12. Cook AK, Cunningham LY, Cowell AK, Wheat LJ. Clinical evaluation of urine Histoplasma capsulatum antigen measurement in cats with suspected disseminated histoplasmosis. J Feline Med Surg 2012 Aug; 14(8):512-5.
  13. Wheat J, French ML, Kohler RB, et al. The diagnostic laboratory tests for histoplasmosis: analysis of experience in a large urban outbreak. Ann Intern Med 1982 Nov; 97(5):680-5.
  14. Sykes JE. Canine and Feline Infectious Diseases, St. Louis, MO, Elsevier Saunders. 2019.
  15. Trivedi SR, Sykes JE, Cannon MS, et al. Clinical features and epidemiology of cryptococcosis in cats and dogs in California: 93 cases (1988-2010). J Am Vet Med Assoc 2011 Aug 1; 239(3):357-69.
  16. Trivedi SR, Malik R, Meyer W, Sykes JE. Feline cryptococcosis: impact of current research on clinical management. J Feline Med Surg 2011 Mar; 13(3):163-72.