Tick-Borne Disease Case Challenges

The Saga of Leia

8 year old, female, intact German Shepherd dogPresented to the referring veterinarian for an acute onset of lethargy, anorexia and panting

Medical History

Diagnosed bilateral pannus two weeks prior to this presentation

Topical prednisolone acetate 1%, q8hOptimmune ointment (0.2% cyclosporine) q12h

Recent estrus cycle

Physical Exam FindingsRectal Temp. 103.50FMild discomfort in caudal abdomenHR normalCapillary refill time was normalMild petechiation was noted on gumsNo vaginal discharge noted

Work-Up

CBCBiochemical profileAbdominal radiographs

CBCHCT 39.5% (37.0 – 54.0)RBC 5.93 x 106 (5.4 – 7.8)RBC indices WNLPLT 32.0 x 103 (150 – 430)MPV 21.8 fl (8.0 – 16.0)WBC 15.69 x 103 (6.0 – 17.0)Neutrophils 12.54 x 103 (3.9 – 8.0)Band 0.100 x 103 (< 0.3)Lymphs 1.1 x 103 (1.3 – 4.1)Eosin 0.0 x 103

Monos 1.98 x 103 (0.2 – 1.1)

Blood Film Evaluation

Moderate rouleauxNo other erythrocyte abnormalities noted

Most lymphocytes and monocytes reactivePlatelets reduced in number (no clumping)

Some large

Interpretation

Stress leukogramEvidence of nonspecific antigenic stimulationThrombocytopenia

Biochemical Profile

Elevated blood glucose (129 mg/dl; N= 76-119)

Attributed to stress

Hyperglobulinemia (5.3 g/L; N= 2.7 – 4.4)No other significant abnormalities

Abdominal Radiographs

Mild to moderate hepatosplenomegalyGas-filled loops of SI (enteritis?)No radiographic evidence of pyometraLumbar spondylosis

DDx for ThrombocytopeniaBone marrow production

Unlikely, with no other cytopenias(neutropenia or nonregenerative anemia)

Peripheral consumption / utilizationMicrovascular disease (DIC, Vasculitis, thromboembolic disease etc.)Infectious disease (TBD)Immune-mediated / idiopathic (?)

Drugs (topical cyclosporine and prednisolone)

RDVM Plan

Rule out infectious diseasesSNAP 3Dx assay for Lyme, E. canis and HWD were all negativeTiters submitted for RMSF and Babesia spp

Coagulation profilePT, PTT, D-dimers and FDPs all within normal limits

RDVM Dx and Tx

Suspected IMTDog placed on prednisone at 1 mg/lb sidDoxycycline 5 mg/kg sidMonitor for 2 days

Platelet count begin to climb (65,000 cells / l)

Discharge with instructions to continue on oral pred and doxy, ophthalmic treatment for pannus

Return in two weeks

Return Visit to RDVM

Leia doing wellTiters for RMSF and Babesia were negativePlatelet count 212,000 cells / lPrednisone decreased to 0.5 mg/lb sidDoxycycline discontinuedReturn 2 weeks

4 Weeks Later, Leia Returns

Leia is sickLethargic, anorexic, pantingRectal Temp. 1040FCapillary refill time normal

Moderate petechiation present again

Refer to UF VMCStill on ophthalmic medicationsOff of oral prednisone for 2 weeks

Physical Exam Finding

Rectal Temp. 1040FCapillary refill time normal

Moderate petechiation present

Mild generalized lymphadenopathyMinimum Data Base

CBCBiochemical profileUrinalysis

Biochemical Profile

NSF

Urinalysis / Cystocentesis

Light yellowSG 1.018pH 8.0Protein negativeHemoprotein traceSediment

3-5 RBCs / hpf

CBC FindingsHCT 29.0% (37.0 – 54.0)RBC 4.70 x 106 (5.4 – 7.8)MCV 62.5 fl (66 – 75)MCHC 32.0 g/dl (34.0 – 36.0)PLT 28.0 x 103 (150 – 430)MPV 24.5 fl (8.0 – 16.0)WBC 15.80 x 103 (6.0 – 17.0)Neutrophils 11.21 x 103 (3.9 – 8.0)Band 0.63 x 103 (< 0.3)Lymphs 1.2 x 103 (1.3 – 4.1)Monos 3.00 x 103 (0.2 – 1.1)

Reticulocytes (1.3%) 61,000 (>80,000)

Interpretation of Hemogram

Microcytic, hypochromic anemiaPoorly regenerativeChronic blood loss

Fecal exam - Stool dark and tarry (fecal exam for parasites negative)Hemoccult test – positive for blood

Thrombocytopenia with increased MPV More pronounced left shiftLymphopenia and monocytosis

Infectious Cyclic Thrombocytopenia

Anaplasma platys(formerly known asEhrlichia platys)Only intracellular organism known to specifically infect platelets

Infectious Cyclic Thrombocytopenia

Tick vectorSuspected to be Rhipicephalus sanguineus, Brown Dog Tick

Eastern and Southeastern USWestern Indian Reservations (Unpublished data)

Clinical Signs

Mild clinical disease or subclinical infection in most dogs

German Shepherd dogs?Immunosuppressive therapy?

Fever, anorexia, sometimes petechiation or epistaxis1 to 2 weeks post-infection

Laboratory Findings

Cyclic thrombocytopenia occurring at 1 to 2 week intervals

Platelets below 20,000 cells / l initially, but rapidly increaseMild thrombocytopenia coinciding with parasitemic episodes

DiagnosisVisualization of organisms in peripheral bloodConfirmation

PCR analysisIFA test (L.S.U.)

Cross reactivity Anaplasma phagocytophilumon SNAP 4Dx assayCross-reactivity with Ehrlichia spp. (E. canis) minimal if any

Not an EhrlichiaLeia was SNAP 3Dx negative

Treatment and PrognosisTreatment same as other Ehrlichias and Anaplasmas (doxycycline)Prognosis excellent

Leia’s Serology

SNAP 3DxLyme, HWD and E. canis

all negative

IFA for A. platys1:160

Treatment Plan for Leia

Doxycycline 5 mg/ kg BID for 30 daysFerrous sulfate (10 mg/kg po bid with meal)

continued until PCV and red cell indices return to normal.

Continue on ophthalmic therapy for pannus

Ocular lesions resolving

Follow-upPlatelet count 2 days later

91,000 cells / l

Discharged to RDVM to recheck in two weeksMaintain on Doxycycline therapy for entire 30 days

The Saga Continues

7 weeks after discharge from UF VMCReturns to referring DVM

Panting, anorexic, rectal temp 1040FBilateral epistaxisMild peripheral lymphadenopathy

Refer to UF VMC for re-evaluation

Return to UF VMC

Physical Exam FindingsPanting, anorexic, rectal temp 103.50FBilateral epistaxisPeripheral lymphadenopathy, most prominent in submandibular nodesNo obvious petechiation

CBC FindingsHCT 34.0% (37.0 – 54.0)RBC 5.10 x 106 (5.4 – 7.8)MCV 67.5 fl (66 – 75)MCHC 34.0 g/dl (34.0 – 36.0)PLT 105.0 x 103 (150 – 430)MPV 18.0 fl (8.0 – 16.0)WBC 17.50 x 103 (6.0 – 17.0)Neutrophils 14.18 x 103 (3.9 – 8.0)Band 0.87 x 103 (< 0.3)Lymphs 1.05 x 103 (1.3 – 4.1)Monos 1.4 x 103 (0.2 – 1.1)

Reticulocytes (1.7%) 86,700 (>80,000)

Blood Film Evaluation

Moderate anisocytosis and polychromasia

Occasional NRBC

Most lymphocytes and monocytes reactivePlatelets mildly reduced in number (no parasites seen)

Problem List

FeverMild thrombocytopenia

Bilateral epistaxis, more in left nostril??

Generalized lymphadenopathy (mild) Very prominent submandibular lymph nodes especially on left side

Plan?

PlanAspirate left submandibular LN

PlanAspirate left submandibular LN

New Plan

Remember what you learned from E.B.B.Sick dog with a history of TBD (A. platys) that responded to doxy only to relapse when discontinuedEpistaxis without significant thrombocytopeniaGranulomatous/pyogranulomatous lymphadenitis

New PlanSend serum to NCSU Vector Borne Disease Diagnostic LaboratoryBartonella vinsonii subsp. berkhoffii and Bartonella henselaeAzithromycin

5-10mg/kg PO Q24 for 5 to 7 daysSame dose every other day for 5 more weeks

Follow-up

72 hours laterTemperature normalEpistaxis resolvedPlatelet count normal

SerologyPositive for B. henselae (1:256)

Leia had uneventful recovery

Co-InfectionsAP and Lyme

Same vector and co-infections common in upper Midwest and Northeastern USCo-infected animals more likely to have severe diseaseTreatment same

AP and Bartonella spp. ( Diroff et al. JVIM 2006, 20:762)

Dogs in Northeastern US that were Bartonella positive on serology

25% also AP positive; no significant association with LymeBartonella but not AP associated with peripheral lymphadenopathyDoxy ineffective in treatment of Bartonellosis

Co-InfectionsLiterature full of data regarding co-infections with tick-transmitted organismsE. canis and epistaxis

Epistaxis has not been reported in experimental E. canis infectionsEpistaxis associated with Bartonella infection

Thrombocytopenia or not (endothelial cell invasion)Vasculitis, vascular weakness

Warrants testing for co-infection

Non-responders or relapses after appropriate therapy

L.S.U. Butterfly

Dog Case - Coco6 year oldfemale spayedmixed breedVomited 2 days agoPresented to rDVM

CBC marked lymphocytosisSuspected lymphoid leukemia

Referred to UFVMC

Dog Case - CBC PCV % 33 L (37.0-54.0) RBC Indicies WNLP.P. g/dL >14 H(6.0-7.8)Fibrinogen mg/dL N/A (100-400)WBC/µL 13,400 (6.0-17.0103)Segs/µL 3,200 LN (3.0-11.5103)Bands/µL 130 (0.0-0.3103)Lymphs/µL 9,700 H (1.0-4.8103)Monos/µL 400 (0.2-1.4103)Platelets/µL 106,000 L (160-430103)

ChemistryTotal protein g/dL 13.8 H (5.6-7.4)

Albumin g/dL 1.4 L (2.8-3.8)

Globulins g/dL 12.4 H (2.3-4.2)

A:G 0.1 L (.7-1.4)

ALT 326 H (10 – 109)

AST 275 H (13 – 15)

Total Bil. 0.2 (0.1 – 0.3)

Calcium 8.1 L (8.7-11.5)

Rest WNL

CBC

Lymphocytosis

Bicytopenia (anemia, thrombocytopenia)with low normal neutrophils

Hyperproteinemia

Serum chemistry

Hyperproteinemia due to hyperglobulinemiaHypoalbuminemiaElevated ALT and ASTNormal total bilirubin

Problem List

Reasons for Additional Tests

Serum protein electrophoresisHyperproteinemia due to hyperglobulinemia

Liver aspirate / bile acidsBone marrow aspirate

Monomorphic lymphoidpopulation Bicytopenia w/ low normal neutrophils

AnemiaThrombocytopenia

Dog Case - SPE

Normal Dog This Dog

α γβ

albumin

albumin

α β

γ

Dog Case - SPE

Normal Dog This Dog

α γβ

albumin

albumin

α β

γ

Monoclonal gammopathy !!!with a polyclonal base

Serum protein electrophoresisMonoclonal gammopathy

Bone marrow aspirateLymphoplasmacytic infiltrate

Lymphocytes: 56% (<1%)Plasma cells: 8% (< 2%)Adequate megakaryocytes

Bile acids –normal pre and post

Ionized calciumNot performed

Additional TestsSerum protein electrophoresis

Monoclonal gammopathy

Bone marrow aspirateLymphoplasmacytic infiltrate

Lymphocytes: 56% (<10%)Plasma cells: 8% (< 2%)Adequate megakaryocytes

Is it a tumor?Is it infection?Immune-mediated disease?

Additional Tests

Serum protein electrophoresisMonoclonal gammopathy

Bone marrow aspirateLymphoplasmacytic infiltrate

Lymphocytes: 56% (<10%)Plasma cells: 8% (< 2%)

Tick-borne disease titerSNAP 3Dx positive

Additional Tests

Serum protein electrophoresisMonoclonal gammopathy

Bone marrow aspirateLymphoplasmacytic infiltrate

Lymphocytes: 56% (<10%)Plasma cells: 8% (< 2%)

Tick-borne disease titerSNAP 3Dx positiveEhrlichia canis positive IFA tier (1:10,240)

Additional Tests Outcome

Treatment with Doxycycline10 mg/kg BID for 30 days

Dog recovered uneventfullyStill positive??

Lost to follow-up

Canine Monocytic Ehrlichioses

Intracellular agents that reside in the monocytes and lymphocytes of infected hostsCan cause clonal proliferation of lymphocytsesMost often E. canis and E. chaffeensis (HME)Diagnosis problematic due to scarcity of circulating organisms

Monocytic Ehrlichiosis vs. Multiple Myeloma

EhrlichiaPlasmacytosishyperglobulinemiaMonoclonal GammopathySerology positive

Multiple MyelomaPlasmacytosishyperglobulinemiaMonoclonal GammopathyHypercalcemia, lytic bone lesions, Bence-Jones proteins

Acute phase

1 - 3 weeks after infectionClinical signs mild and nonspecific (fever, lethargy, weight loss, anorexia)Thrombocytopenia +/- anemia, leukopeniaTiters may be negative

Subclinical PhaseFew if any clinical signsThrombocytopenia (usually mild)Hyperglobulinemia with positive titerMay last months to years?

Chronic PhaseReappearance of clinical disease

breed, stress, concurrent disease

Mild to severeage, breed, strain of organism

Weakness, anorexia, weight loss, fever, pallorLymphadenopathy, hepatomegaly, splenomegaly, nephropathies, retinal lesions, edema, nonseptic polyarthritis, CNS disease, and mortality

DiagnosisPresumptive diagnosis based on clinical and routine laboratory findings

+/- history of tick exposureConfirmed with serology or PCR analysisPCR used to speciate infectious agents

Not as sensitive as serology

PCR

False negative, particularly in chronically infected dogs

Must do PCR analysis on splenic aspirates or bone marrow aspirates to evaluate therapy

Snap3Dx (IDEXX Laboratories Inc.)Uses P30 and P30-1 antigens of E. canisHigh specificity (100%), but low sensitivity (79.2%)(Bélanger et al., J. Clin Microbiol, 2001

High positive and negative predictive values in low prevalence populationsCannot differentiate between E. canis and E. chaffeensis infections

Treatment and PrognosisDoxycycline (5 mg/kg PO bid for 21 -30 days) or tetracycline (20 mg/kg PO tid)Eliminates clinical signs (titers may persist)Prognosis excellent in acute cases and mild chronic casesPrognosis guarded in dogs with severe pancytopenia or aplastic bone marrow