Improving the reference standard for the diagnosis of canine visceral leishmaniasis: a challenge for current and future tests

Teixeira, Ana Izabel Passarella; Silva, Debora Marcolino; Vital, Tamires; Nitz, Nadjar; de Carvalho, Bruna Caroline; Hecht, Mariana; Oliveira, Diana; Oliveira, Edward; Rabello, Ana; Romero, Gustavo Adolfo Sierra

doi:10.1590/0074-02760180452

Brasil

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ORIGINAL ARTICLES • Mem. Inst. Oswaldo Cruz 114 • 2019 • https://doi.org/10.1590/0074-02760180452 linkcopy

Improving the reference standard for the diagnosis of canine visceral leishmaniasis: a challenge for current and future tests

Authorship SCIMAGO INSTITUTIONS RANKINGS

BACKGROUND Studies aimed at validating canine visceral leishmaniasis diagnostic tests present heterogeneous results regarding test accuracy, partly due to divergences in reference standards used and different infection evolution periods in animals.

OBJECTIVE This study aimed to evaluate the accuracy of the rapid test-dual path platform (TR-DPP) (Biomanguinhos®), EIE-Leishmaniose-Visceral-Canina-Biomanguinhos (EIE-LVC) (Biomanguinhos®), enzyme-linked immunosorbent assay (ELISA) rK39 (in-house), and the direct agglutination test (DAT-Canis) against a reference standard comprising parasitological and molecular techniques.

METHODS A phase II/III validation study was carried out in sample sera from 123 predominantly asymptomatic dogs living in an area endemic for visceral leishmaniasis.

FINDINGS Sixty-nine (56.1%) animals were considered infected according to the reference standard. For each test, the sensitivity and specificity, respectively, were as follows: TR-DPP, 21.74% [confidence interval (CI)95% 13.64% to 32.82%] and 92.59% (CI95% 82.45% to 97.08%); EIE-LVC, 11.59% (CI95% 5.9% to 21.25%) and 90.74% (CI95% 80.09% to 95.98%); ELISA rK39, 37.68% (CI95% 27.18% to 49.48%) and 83.33% (CI95% 71.26% to 90.98%); and DAT-Canis, 18.84% (CI95% 11.35% to 29.61%) and 96.30% (CI95% 87.46% to 98.98%).

CONCLUSION We concluded that improving the sensitivity of serum testing for diagnosing asymptomatic dogs must constitute a priority in the process of developing new diagnostic tests to be used in the visceral leishmaniasis control program in Brazil.

Key words: canine visceral leishmaniasis; diagnostic tests; serology

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Instituto Oswaldo Cruz, Ministério da Saúde Av. Brasil, 4365 - Pavilhão Mourisco, Manguinhos, 21040-900 Rio de Janeiro RJ Brazil, Tel.: (55 21) 2562-1222, Fax: (55 21) 2562 1220 - Rio de Janeiro - RJ - Brazil
E-mail: memorias@fiocruz.br

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[TFN1] a: direct agluttination test; b: immuno-enzymatic assay for CVL; c: rapid test dual path platform; d: enzyme linked immunosorbent assay rk39.w

Findings	Score
Findings	0	1	2	3
Bodily condition	Obese/Normal	Thin	Cachectic	--
Mucosae	Normal	Pale	Jaundiced	--
Dehydration	Absent	Light	Moderate to intense	--
Muscle atrophy on limbs	Absent	Light	Moderate to intense/widespread	--
Skin lesions	Absent	Present	--	--
Hepatosplenomegaly	Absent	Present	--	--
Conjunctivitis and / or Keratitis	Absent	Unilateral and light	Severe unilateral / bilateral	--
Uveitis and / or Blepharitis	Absent	Unilateral and light	Severe unilateral / bilateral	--
Lymph adenomegaly	Absent	1 to 2 lymph nodes	--	3 or more / widespread
Epistaxis	Absent	Present	--	--
Mouth ulcers or nodules	Absent	1 to 2	3 or more	--
Vomit	Absent	Present	--	Frequent, with vomit
Diarrhea	Absent	Present	--	--
Claudication	Absent	Present	--	--
Erythema	Absent	1 to 25% of the body	25 to 40% of the body	40% or more of the body
Dry exfoliative dermatitis	Absent	1 to 25% of the body	25 to 40% of the body	40% or more of the body
Ulcerative dermatitis	Absent	1 to 25% of the body	25 to 40% of the body	40% or more of the body
Nodular dermatitis	Absent	1 to 25% of the body	25 to 40% of the body	40% or more of the body
Pustular dermatitis	Absent	1 to 25% of the body	25 to 40% of the body	40% or more of the body
Alopecia	Absent	1 to 25% of the body	25 to 40% of the body	40% or more of the body
Altered pigmentation	Absent	Present	--	--
Hyperkeratosis of truffles and cushions	Absent	Present	--	--
Onychogryphosis	Absent	Present	--	--

Index tests		Reference standard		Test accuracy (%)		Likelihood ratio
Index tests		Positive (+)	Negative (-)	Sensitivity (CI95%)	Specificity (CI95%)	Positive (CI95%)	Negative (CI95%)
DAT-Canis ^a	(+)	13	2	18.84	96.30	5.08	0.84
DAT-Canis ^a	(-)	56	52	(11.35 to 29.61)	(87.46 to 98.98)	(1.19 to 21.58)	(0.74 to 0.95)
EIE -LVC^b	(+)	8	5	11.59	90.74	1.25	0.97
EIE -LVC^b	(-)	61	49	(5.99 to 21.25)	(80.09 to 95.98)	(0.43 to 3.61)	(0.86 to 1.09)
R-DPP^c	(+)	15	4	21.74	92.59	2.93	0.84
R-DPP^c	(-)	54	50	(13.64 to 32.82)	(82.45 to 97.08)	(1.03 to 8.33)	(0.73 to 0.97)
ELISA rK39^d	(+)	26	9	37.68	83.33	2.26	0.74
ELISA rK39^d	(-)	43	45	(27.18 to 49.48)	(71.26 to 90.98)	(1.15 to 4.41)	(0.60 to 0.93)