genetic Resistance against gastrointestinal nematodes in sheep: Association with genomic regions

Sheep resistance to Gastrointestinal nematodes:
Association with Genomic regions
Presented by
ISHFAQ MAQBOOL
PhD Scholar (Parasitology) Roll No. : L-2016-V-10-D

Background
Identification of Genomic regions
Candidates genes for genetic resistance
Mechanisms for host response
Conclusion
Genomic regions for 1, 2 and all GINs

Background
 Gastrointestinal nematodes are one of the most
serious cause of disease in small ruminants
 The cost of parasite treatment alone has been
estimated tens of billions of dollars worldwide
(Waller and Chandrawathani, 2005)
Estimated treatment cost alone for H. contortus per
year in India was US $103 million
(McLeod, 2004)

Common GI nematodes
Stomach-: Haemonchus contortus, Trichostrongylus axei
Small intestine: T. colubriformes, Chabertia ovina, Bunostomum
trigonocephalum, Gaigeria pachyscelis.
Large intestine: Oesophagostomum columbianum, Trichuris ovis
Bunostomum spp. Chabertia ovina Oesophagostomum spp. Haemonchus contortus

Anthelminthic resistance
(Burke and Miller, 2006; Chandra et al., 2014, 2015)
Demand for “clean and green” animal
products free of residual chemicals and
growth promoters
(Waller, 2003)
Increasing toxicity and persistence of
chemical residues in the environment
(Wolstenholme et al., 2004)
Lack of commercially available
vaccines
Need for genetically resistant livestock

Viable method of nematode control
Absolute trait nor it is confined to breeds (Eady et al., 1996)
Moderately heritable trait. (Koudande et al., 2005; Yazdi et al., 2010)

Quantitative trait controlled by
polygenes. (Bishop, 2012)
QTL: Location or region of
individual locus or multiple loci in
the genome that affects a quantitative
trait.

Red Masai
Florida Native St. Croix
.
Garole
Barbados blackbelly
(Baker and Gray, 2004, Nimbkar et al., 2003)

Traditional
Breeding
Molecular
Breeding

Parasitological Immunological Pathological
Faecal egg count
Worm count and
Worm length
Serum antibodies (IgG1, IgA
and IgE)
Peripheral eosinophilia
Histology (mast cells,
eosinophils, goblet cells)
Plasma pepsinogen
Plasma albumin
Fructosamine
Packed cell volume
Dag score
Faecal consistency

Limitations of Traditional Breeding
Reduced efficiency in low-heritability-characters
Sex linked traits
Characters expressed later in life
Characters with unfavorable genetic correlation
Characters to be measured in large number of animals
Alternative: Complementation of traditional breeding strategies with
available molecular genetics & biotechnological techniques

Genetic Marker
Any stable & inherited
variation in the DNA,
that can be identified by
a suitable method to
detect the presence of a
specific genotype which
is otherwise very difficult
to detect
Genetic markers are “landmarks’ at the genome that can
be chosen for their proximity to QTL.

Major histocompatibility complex (MHC) region on Ovis aries chromosome 20
(OAR20) most frequently studied
(Janssen, et al. 2002 , Davies, et al. 2006, Riggio, et al. 2013)
Genes belonging to MHC: highly polymorphic, play important roles in
presenting processed antigens to host T lymphocytes and immunological
cascade of events

The interferon γ (IFN-γ) gene on OAR3:
Second to the MHC region, Th1 pro-
inflammatory cytokine involved in
recruiting cellular responses
(Coltman et al., 2001, Crawford et al., 2006 and Silva et al., 2011)
Evidence from murine models shows that
high levels of IFN-γ may negatively
interfere with the development of a Th2
protective immune response against
gastrointestinal parasites. .
(MacKinnon et al., 2015)

Several candidate gene studies demonstrated the role of IFN-γ in
sheep resistance to T. circumcincta.
(Beraldi et al., 2007)
Significant interval markers were also found close to the IFNG region
of sheep resistant to T. circumcincta, suggesting that IFNG
neighbouring genes may also be of interest in host resistance
(Silva et al., 2011)
Down regulating IFN-γ, leading to a stronger Th2 response
(Coltman et al., 2001)

Gene/ gene
family
Functions
Chemokine receptors
 Transduction of signals that stimulate migration of leukocytes
Interleukin receptors
 Antibody-dependent cytotoxicity
 Release of inflammatory mediators and
 Stimulation of phagocytosis
Leukocyte
immunoglobulin-like
receptor superfamily
 Expressed predominantly on monocytes, B-cells and, to a
lesser extent, on dendritic cells, natural killer (NK) cells
 Stimulation or inhibition of NK cell activation

Gene/ gene
family
Functions
Natural resistance
associated
macrophage protein
 Regulation of chemokines
 Inducible nitric oxide release
 MHC class II molecules and
 macrophage activation
Toll-like receptor  Recognition of conserved molecules unique
to pathogens
Tumor necrosis
factor
Multifunctional pro-inflammatory cytokine
(Acevedo and Cunningham, 2006)
Nramp

 QTL studies in livestock have been
conducted in pedigreed populations
for a trait that is quantitative in
measure
 Modest number (100–200) of genetic
markers employed to unveil genomic
regions associated with the trait of
interest
GWAS
 GWAS can fit pedigreed populations
but it can also be used in case-control
studies with no pedigree information
 It employs thousands of
polymorphism (SNP markers) to
unveil genomic regions associated
with the trait of interest
*Identification of genomic regions

Chromosome Regions Parasites
OAR 1 A, B, C, D and E H. contortus, T. circumcincta and T. colubriformis
OAR 3 F, G, H, I, J, K and L H. contortus, T. circumcincta and T. colubriformis
OAR 6 M and N H. contortus, T. circumcincta and T. colubriformis
OAR 20 O and P Teladorsagia circumcincta and Haemonchus contortus
126 significantly associated markers and marker intervals spread across the genome
(Benavides, et al. 2015)

1. Innate and
acquired
immune cell
responses:
Th2 cytokine
production
Delivery of
effector cells and
 IgE to the site of
infection.
2. Mucin
biosynthesis:
Role in gastric
mucosal protection
As a barrier
Triggering swift
parasite expulsion.
3. Hemostasis
pathways :
Required for blood
clotting
Obstruction of
parasite feeding
Maintenance of
host PCV levels.

Parasite infections caused by the pathogenic H. contortus:
characterized by a type 2 helper T cell (Th2) response
Increased number of mucopeptic cells, eosinophilia, mastocytosis,
and immunoglobulin E (IgE) production triggered by secretory
factors produced by parasites
(Balic et al., 2000 and MacKinnon et al., 2015 )
Parasite Chromosome Region and functions
Haemonchus
Contortus
OAR3 a) F region where markers are either closely located to or
overlapped immune response genes for biosynthesis of
interleukins for Th2 response (Balic et al., 2000)
b) H region containing several interleukin receptors and
receptor-like genes (MacKinnon et al., 2015)
Teladorsagia
circumcincta
OAR1 and
OAR3
a) C, J and K: Regions containing genes for IFN-γ, a pro
inflammatory cytokine
(Sayers, G. et al., 2005 and Riggio et al., 2013)
Trichostrongylus
colubriformis
OAR1 a) E: Several genes involved with platelet activation &
aggregation, and with wound healing
(Beh et al., 2002)

Parasite Chromoso
me
Region and functions
H. contortus &
T. circumcincta
OAR1, 3, 6
and 20
OAR1
OAR3
a) B, D, L, M, O and P: Wound healing, platelet
adhesion, and blood clotting, Production of IL-3 and
IL-6, key cytokines for IgE production, and
mounting the Th2 immune response against GI
parasite infection (Finkelman et al., 1997)
b) D: Mucin biosynthesis and glycosylation
L: Mucin biosynthesis and glycosylation
H. contortus &
T. colubriformis
OAR1 and
OAR3
a) I: overlaps the SOCS2 gene which is responsible for IL-3
regulation
(Tannahill, et al. 2005)

Parasite Chromoso
me
Region and functions
H. contortus &
T. circumcincta
OAR20 O: Genes implicated in MHC-mediated antigen
processing and presentation and lymphocyte signalling
pathways
In OAR20, the positions of IL17A and IL17F to be
relatively close to DRB1. These interleukin genes have
not been yet described in sheep resistance studies
(Korn et al., 2009 and Mihi et al., 2014)

IL-17 is the leading inflammatory cytokine in
Th17 cell populations and IL-17 transcripts have
been shown to be strongly upregulated in cattle
after a single challenge with the bovine abomasal
nematode Ostertagia ostertagi
(Korn et al., 2009 and Mihi et al., 2014)

IL-3 gene:
Differentially expressed in the abomasal lymph nodes of lambs
resistant to T. circumcincta infection versus control individuals
(Gossner, et al. 2013)
SOCS2 thus could be important for improving sheep immune
response
(Madden, et al. 1991)

H. contortus,
T. circumcincta
and T.
colubriformis
OAR1 a) A and N: Important genes related to hemostasis
TAL1 that is important for platelet formation and
regulation of mast cell differentiation
PPAP2B involved in regulating wound healing
LRP8 related to the platelet hemostasis pathway

H. contortus,
T.
circumcincta,
and
T.
colubriformis
OAR6 a) N: Several toll-like receptor genes (TLR1, 10, and 6),
members of the TLR signalling pathway, NF-κB family,
and Th17 differentiation pathways
(Moncada et al., 2003)
b) Several other genes involved in the immune
response:
 PDGFRA and KIT which encode proteins
implicated in MHC class I mediated antigen
processing and presentation, and genes encoding the
IFN-γ-induced chemokines CXCL9, 10, 11, and 13
 MUC7: involved in mucin biosynthesis, is also

Sixty other genes or chromosomal regions were found to be scattered
across the ovine genome outside chromosomes 1, 3, 6 and 20
Chromosome Gene function Reference
OAR15 MUC15 Encoding a cell surface-
associated protein belonging to
the O-linked glycosylation
pathway
Salle et al 2012
OAR21 PGA5 Pepsinogen synthesis Benavides et al 2015
OAR23 CD226 Encoding a protein related to
class I MHC-mediated antigen
processing and presentation
Marshall et al 2012
GALNT1 Encoding a mucin-type O-
glycosylation enzyme involved
in mucin biosynthesis
Marshall et al 2012

Segment A B C D E F G H I J K L M N O P
From
(Mbp)
15.2 108.5 171.4 179.5 216.3 12.4 58.8 83.7 125.9 151.5 171.2 193.2 16.8 55.9 7.19 12.32
To (Mbp) 44.7 168.9 175.1 196.9 273 50.8 99.6 134.7 153.8 188 212.3 36.6 112.8 7.28 40.7
Genes
(from
proximal
to distal
inthe
chromoso
me)
TAL1
LRPS
PPAP
2B
(h)
FCER
1A
FCER
1G
(i)
PHLD
B2
MYL
K (h)
MUC
13
MUC
20
MUC
4 (m)
GP5
(h)
RAP2
B
P2RY
1
P2RY
12
CPA3
(i)
PIK3
CB
(h)
DAB2
IP (i)
ADA
M17
TRIB
2
GAL
NT14
(m)
ANX
A4
PLEK
(i)
IL18R
AP
IL18R
1
IL1R
L1
IL1R
L2
IL1R1
IL1R2
(i)
ATP2
B1 (h)
SOCS
2 (i)
ZNF3
85A
(h)
GAL
NT6
(m)
IFNG
(i)
MYH
9 (h)
IL2R
B (i)
VWF
(h)
GALNT
8 (m)
NFKB
1 (i)
SPP1
(h)
TLR1
TLR1
TLR6
(i)
RHO
H (h)
PDGF
RA
KIT
MUC
7
IL8
PPBP
(h)
CXC
L9-13
DYA
TAP1
(i)
TRE
M2
IL17A
IL17F
DRBI
TNFA
MUC
21
HIST
1H2B
F
HIST
1H2B
I
HIST
1H2B
E
UBE2
N (i)
Chro
moso
me
1 3 6 20

Known and proposed
mechanisms for host response
to infection

Increase in GI mucus
Reduction of larval motility and
feeding
Increase in peristalsis
Parasite expulsion from GI tract
GALNT- 1, 3, 5, 6, 7, 8, 13, and 14
MUC- 4, 7, 13, 15, and 20

Blood clotting at
the host–parasite
attachment site
Parasite feeding
restraint
Reorganization
of mucosal tissue
Wound healing
process
Steady host PCV
level
ATP2B1, LRP8, PDGFRA, and
SPP1

Parasite
antigens
Antigen
presentation to
MHC, T cell
activation
IgE production
Eosinophil and
mast cell
migration
IgE armed
eosinophil and
mast cells
degranulation
Mount strong
Th2 response
MHC complex (Class II, DRB1 and DQA)

Th2 response in helminths
(Balic et al., 2000 and MacKinnon et al., 2015 )

Cellular and Molecular Immunology 7th edition

.
Some of the molecules released from eosinophils that cause damage to parasitic helminths.
Text book of veterinary immunology 9th edition Ian R Tizzard
EPO: oxidants and NO
ECP,ENT: Ribonucleases

.
Mast cells
Histamine 5HT Proteinases Prostaglandins Leucotrienes
Increased mucosal permeability/SM contraction

Breeding using molecular markers is viable method for control of GINs
Host resistance is of a quantitative nature.
Numerous molecular markers have been reported to be associated with FEC
and its related traits
Some genomic regions have been shown to be exclusive to particular
parasite species
Not only that host acquired immune system mechanisms are important to
build host resistance but genes involved in gastrointestinal mucus production,
parasite expulsion, and hemostasis regulation are also relevant

genetic Resistance against gastrointestinal nematodes in sheep: Association with genomic regions

genetic Resistance against gastrointestinal nematodes in sheep: Association with genomic regions

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