Table 1.

Summary of differentiating characteristics of tick susceptible vs. tick resistant cattle (bold font indicates correlation between studies of certain transcripts and or other markers—genetic, cellular, immunohistochemistry) in response to Rhipicephalus microplus ticks.

Methodology (cattle breeds compared)	Susceptible	Resistant	Main conclusion and reference
Cellular IgG isotyping; Cattle naturally infested on pasture; susceptible cattle treated with acaricide when tick counts were >500. Cattle vaccinated with GAVAC. (Holstein and Aberdeen B. t. taurus vs. Nelore B. t. indicus)	IgG1 and IgG2 decrease (anti-salivary gland antibodies only)	IgG1 and IgG2 no change	IgG suppression by tick infestation on susceptible cattle. Kashino et al., 2005
Genetics study, cattle naturally infested in pasture and at 10–14 months infested with 10,000 tick larvae, PCR to determine allele frequency (F2 population of composites B. t. indicus Gyr 1/2 x B. t. taurus Holstein 1/2)		BoLA Class II DRB3.2	The MHC Class II DRB3.2 allele is associated with tick resistance. Martinez et al., 2006
Microarray gene expression, natural tick infestation for ~6 months (“naïve” sample taken post field infest), followed by 5 artificial tick challenges over 17 months with 20,000–25,000 larvae (Hereford Shorthorn cattle- tropically adapted B. t. taurus)	B-cell CLL/lymphoma 10 Collagen, pro α Apolipoprotein Keratin Immunoglobulins IL13 receptor α 1 precursor	Cathepsin B Collagens (type I α2; type III pro; type VI, α 3) Complement component 1, q subcomponent, α	Susceptible cattle upregulated immune responsive genes and those involved in protein synthesis. BDA20, Odorant Binding Protein (OBP) and the dendritic cell protein-B5 over expressed by both groups. Wang Y. H. et al., 2007
Genetic microsatellite and cytokine qPCR of 4 breeds, 17 months natural tick infestation (B. t. indicus Nelore, Composite Canchim 5/8 B.taurus 3/8 B. t. indicus × Nelore, B. t. taurus Aberdeen Angus × Nelore, and B. t. taurus Simmental x Nelore)	IL4	IL8 and IL2 down-regulated	Tick counts were associated with IL4, IL2 is down-regulated in 3 genetically different groups of infested bovines compared to pre-infestation, and IL8 was down-regulated in resistant bovines compared to susceptible animals Regitano et al., 2008
RT-PCR gene expression of peripheral white blood cells, previous natural exposure to ticks, 7 weekly artificial infestations of 10,000 larvae while held in tick infested pasture. (Holstein Friesian B.taurus vs. Brahman cattle B. t. indicus)	Toll-like receptor pathway (TLR5, TLR7, TLR9, NFKBp50, MyD88, Traf-6, CD14, IL-1β) Chemokine ligands and receptors (CCL2, CCL26; CCR-1) IL-10 bovine dander allergen 20 (BDA20)		15 transcripts increased significantly in the susceptible breed to suggest innate inflammatory processes Piper et al., 2008
Cellular, RT-PCR and microarray gene expression (microarray) of leukocytes– infestation as described above (Piper et al., 2008) (Holstein Friesian B. t. taurus vs. Brahman cattle B. t. indicus)	Higher WBC counts Low hemoglobin CD14+ and MHC Class II (macrophage cells) High tick specific immunoglobulins (larvae, adult female salivary gland and gut)—high IgG1 CXCL10	CD4+, CD25+ and γδ T cells IL2, IL2Rα, IL4Rα, TNFα, CCR-1, CCR-7, CXCL4, CD28, CD3E CD40 ligand	Susceptible cattle produce an innate inflammatory type response with high IgG1 titers suggesting also a T cell response. Resistant cattle produce a stable T cell mediated response. Piper et al., 2009
RT-PCR gene expression of calcium signaling genes in skin biopsies. Natural field exposure to ticks, followed by treatment and at 8 months artificial infestation of 10,000 larvae. Skin biopsies at 0, 3, and 24h post infestation (Belmont Red—composite breed ~ 1/2 B. t. taurus and 1/2 B. t. indicus, tropically adapted)		Calcium signaling genes (significant at 24 h post-exposure): AHNAK nucleoprotein (desmoyokin) CASQ (Calsequestrin) IL2 (Interleukin-2) NFAT2CIP (nuclear factor of activated T-cells, calcineurin-dependent) PLCG1 (phospholipase C, γ1)	The significant elevation of some calcium dependent genes following tick exposure suggests that the calcium pathway might be responsive to parasite exposure and could contribute to host immune response. Bagnall et al., 2009
Protein and mRNA expression- infestation as described above (Bagnall et al., 2009) of skin biopsies at 0, 3 and 24 h post-infestation. (Belmont Red – composite breed ~ 1/2 B. t. taurus and 1/2 B. t. indicus, tropically adapted)		Keratins lipocalin 9 epidermal barrier catalyzing enzyme transglutaminase 1 transcriptional regulator B lymphocyte-induced maturation protein 1 lipid processing proteins	Resistant cattle have physically stronger epidermal layers of the skin Kongsuwan et al., 2010
Host gene expression microarrays (skin); infestation as described above Piper et al., 2008 (Holstein Friesian B. t. taurus vs. Brahman B. t. indicus cattle)	Chemokine ligands (C-X-C motif) 2 and 5; (C-C motif) 2 and 8 – CXCL2, CXCL5, CCL2, CCL8 Complement component 1 and 3; Complement factor D (adipsin) IL8 Major histocompatibility complex Class II DRB3 Chemokine Regakine 1 (chemotactic activity for neutrophils) Spleen trypsin inhibitor; Plasminogen activator, tissue; Serpin peptidase inhibitor, clade F; Proapoptotic caspase adapter protein (inflammatory role, recruitment of immune cells to infected tissue); Peptidoglycan recognition protein 1 (inducer of TNFα and IFN γ); Apolipoprotein D Lysozyme Prostaglandin D2 synthase (inhibitor of platelet aggregation)	Keratocan, osteoglycin, lumican, Collagen, (types I and III); Lysyl oxidase-like 4 (formation of collagen) Down-regulated Cytochrome P450 (CYP4F3, CYP11A1) Serpin peptidase inhibitor, clade A (alpha-1 antiproteinase, antitrypsin); Phospholipase A2, group VII (platelet-activating factor acetylhydrolase, plasma); Secreted phosphoprotein 2 (cystatin); Procollagen C-endopeptidase enhancer Lipid metabolism Amino acid metabolism Oxidoreductases	Genes involved with inflammatory processes and immune responsiveness upregulated in susceptible cattle. Genes encoding consistuents of the extracellular matrix were up-regulated in resistant cattle. Piper et al., 2010
qPCR analysis after artificial tick infestation, skin biopsies from tick lesions. (Gyr -B. t. indicus × Holstein - B. t. taurus)	Calcium-dependent signal transduction (S100A7), histamine releasing protein (TPT1), epithelial calcium channel 2 (TRPV6) cysteine proteinase inhibitor (CST6)		Susceptible cattle develop hypersensitive reaction which is not protective Nascimento et al., 2010
EST analysis, natural tick infestation followed by acaricide treatment and artificial infestation of 10,000 larvae, biopsies collected at Days 5 and 12 from base of tail (F2 population from Gyr -B. t. indicus × Holstein - B. t. taurus)	CD44 antigen (lymphocyte activation) CD63 antigen (marker for activated basophils and IgE-mediated allergy) ADAM metallopeptidase odorant binding protein poly A binding protein ribosomal proteins	MHC Class antigen 1 Cathepsin L2 precursor Collagen (type I alpha; type III alpha) Keratins Ribosomal proteins	Structural proteins and MHC Class I in resistant cattle; Immune response transcripts in susceptible cattle Nascimento et al., 2011
Immunohistochemistry, infestation of Holstein Friesian and Brahmans as described by (Piper et al., 2008) above. Infestation of tick naïve Santa Gertrudis cattle−10,000 larvae artificially weekly for 13 weeks followed by natural infestation on pasture (Holstein Friesian B. t. taurus vs. Brahman B. t. indicus) (Composite Santa Gertrudis breed B. t. taurus 5/8 × B. t. indicus 3/8)	CD45, CD45RO (Holstein-Friesian) Mixed CD45 and CD45RO reactions (Santa Gertrudis)	CD4+, CD8+, CD25+ and γδ T cells MHC Class II cells Mixed CD45 and CD45RO reactions (Santa Gertrudis)	Higher number of γδ T cells present in the skin of tick naïve resistant cattle. Epitopes recognized by some antibodies might not be present on the cell populations from all breeds of cattle or might be expressed in different levels. Santa Gertrudis resistance associated with lower cellular reaction at the larval attachment sites. Constantinoiu et al., 2010, 2013; Jonsson et al., 2014
Microarray gene expression (skin), natural infestation on pasture (10 months), acaricide treatment, followed by artificial infestation 10,000 larvae, skin biopsies 0, 24 and 48 h post-artificial infestation. (F2 of composite Gyr -B. t. indicus × Holstein - B. t. taurus)	Calcium ion control genes Cytochrome P450 (CYP4F3)—leukotriene (allergy inducing chemical mediator) degradation	Complement factors 1, 2 and 3 complexes (C1, C2, C3); complement factor 4 binding protein; complement factor B properdin, galectin-1 Coagulation Factor XIIIa Lipid metabolism	Lipid metabolism in inflammation control in resistant cattle. Acute phase response impaired in susceptible cattle. Carvalho et al., 2014
RT-PCR gene expression of peripheral white blood cells, natural tick infestation followed by acaricide treatment, subsequent artificial infestation with 20,000 larvae, blood samples taken at 0, 24, and 48 h post-infestation. (F2 of composite Gyr -B. t. indicus × Holstein - B. t. taurus)	CXCL10 and CXCL8 were down-regulated	CD4+, CD25+ and γδ T cells IL10, FoxP3, CXCL10, CD25	A correlation between T γδ cell activity and immunological mechanisms in resistant cattle Domingues et al., 2014
Cellular and humoral responses (blood), 10,000 larvae artificially weekly for 13 weeks followed by natural infestation on pasture, blood collected 21 days post 1st infestation and weekly to measure changes in measure parameters (Composite Santa Gertrudis breed B. t. taurus 5/8 x B. t. indicus 3/8)	High tick-specific (anti-salivary and gut, anti-larval) IgG1 titers; variable IgG2 responses Decreased hemoglobin, decreased RBC count Increased CD25+, CD4+, WC1+	No IgG2 Increased CD4+ cells (prior to infestation) Decreased CD3+, CD4+, CD8+ Increased MHCII+, WC3+ and CD14+ cells	A non-protective high level of IgG1 in tick susceptible cattle. Piper et al., 2017
Microarray gene expression (skin) and histology Acaricide treatment of calves prior to artificial tick infestation with 10,000, skin biopsies with and without feeding ticks were taken prior to infestation (day 0) followed by 2 days (larvae) and 9 days (nymphs). Biopsies without feeding ticks to emulate “stressed” phenotype. Selected immune response genes differentially expressed (>2-fold) by larvae in comparison to baseline and stress responses (no ticks) in susceptible and resistant cattle (Table S5). (Holstein B. t. taurus vs. Nelore B. t. indicus)	CD209 antigen (leads to IL10 production) Fibroleukin (fibrinogen type 2) Secretogranin-2 Secretoneurin (chemotaxis of monocytes and eosinophils) CXC6 (downregulated); CCL3, CCL4 high affinity immunoglobulin epsilon receptor subunit alpha precursor (NF-kB, MAPK activation) Complement factor D Complement C1q tumor necrosis factor-related protein 7 precursor T-cell surface glycoprotein CD3 delta chain platelet-derived growth factor receptor-like protein precursor (wound healing) Serine/threonine-protein kinase 1 Secreted phosphoprotein 24 (endopeptidase, platelet degranulation) Keratin Protein S100-G (calcium binding) Lysozyme	CD3+ and γδ T cells WC1+ Activation of basophils interferon-induced GTP-binding protein Mx2 Interferon alpha-inducible proteins 6 and 27 IL3 cathepsin D precursor	Susceptible cattle produce more tick attracting volatiles. Resistant hosts expose ticks to an earlier inflammatory response. Franzin et al., 2017