A lifelong study of a pack Rhodesian ridgeback dogs reveals subclinical and clinical tick-borne Anaplasma phagocytophilum infections with possible reinfection or persistence

Table 1 Results of serological and molecular diagnostic tests for anaplasmosis on the six Rhodesian ridgeback dogs performed at multiple time points throughout their lives

Clinic			PCR					Serology
Dog (ID)	Birth- Death	Lethality	VMDC	RIVM 2001	RIVM 2003	RIVM 2014	Total	IFT	Immunoblot	Snap test	Total
			16S rRNA	16S rRNA	16S rRNA	msp2 & groEL		Whole cell	P44	P44
M12	1994–2001	Splenitis, fever and pain		1/1		1/3, spleen neg	2/3	1/2	2/2	0/4	2/5
L11	1998–2009	Malignant tumor in spleen, lung	1/4	2/3	2/9	3/4, spleen pos	5/16	2/3	3/12	3/18	7/21
L12	1998–2005	Splenitis, fever and pain		0/1	0/1	0/1, spleen neg	0/4	nd	0/3	3/5	3/7
L13	1998–2009	Haemangiosarcoma		0/1	0/1	0/2, spleen neg	0/5	nd	0/2	1/6	1/6
L21	1999–2005	Lymphoma		0/1	0/1	0/1, spleen neg	0/3	nd	1/2	0/6	1/7
N00	1995–2009	Metastasis mammae tumor		0/1	0/1	0/1, spleen nd	0/3	nd	1/1	2/6	3/6

Notes: Different laboratories were involved during the years, employing varying serological and molecular tests. Results are presented separately and overall: (i) The Veterinary Microbiological Diagnostic Center, Veterinary Faculty, Utrecht University, The Netherlands (VMDC) performed serological and molecular tests for ehrlichiae and Babesia as of 2001. Immunofluorescent tests for E. phagocytophilum and E. canis were performed on the 2001 samples; (ii) The National Institute of Public Health and Environment (RIVM) employed an immunoblot with the p44 protein of A. phagocytophilum and amplified the 16S rRNA gene and confirmed the result with RLB (see Methods); (iii) The RIVM followed the same methods in 2003 with another team of technicians; (iv) The molecular detection by the RIVM in 2014 targeted a different set of genes (than in 2 and 3) in blood and spleen tissues (see Methods). The in-house SNAP® 4 Dx test (IDEXX) as serological assay was also employed.
Abbreviations: neg, negative, nd not determined, pos positive

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