
Vector-borne diseases in cattle / Yalçın et al. _______________________________________________________________________________________
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INTRODUCTION
Vector–borne diseases have been increasing in recent years, and
global climate change and animal population movements are particularly
effective in spreading such diseases. These infections occur especially
in tropical and subtropical areas and are also seen in our country [1].
Microorganisms in the Order Rickettsiales, which cause infections
transmitted through arthropods, are small coccobacilli that can multiply
in host cells and show Gram–negative properties [2]. Rickettsia species
belonging to the Rickettsiaceae family and Anaplasma and Ehrlichia
species belonging to the Anaplasmataceae family, within the Order
Rickettsiales [3], are important pathogens for farm animals. They are
important for animal and public health because they contain some
species that can cause human infection [4]. They are bacteria that
settle in endothelial cells, immune system cells or erythrocytes, have
obligate intracellular properties and are transmitted through blood [5].
Anaplasmosis is a bacterial infection that causes serious economic
losses in animal husbandry and is also important for public health.
Transmission is caused by the genera Ixodes, Dermacentor, Rhipicephalus
and Amblyomma ticks. The infection is caused by Anaplasma spp. and
infects the red blood cells of vertebrates [6]. The agent is transmitted
biologically by ticks, mechanically by ies and contaminated materials.
The pathogenic species in cattle is A. marginale [7]. In addition to
this species, A. centrale, A. bovis, A. ovis, A. phagocytophilum and A.
platys cause infection dened as Anaplasmosis in cattle [8, 9, 10].
Ehrlichiosis is a disease caused by species of the Ehrlichia genus.
Ehrlichia species settle intracytoplasmically in the leukocytes of their
host [11]. Ehrlichiosis in cattle can be accompanied by fever, protruding
tongue, oppy ears, turning around, excessive chewing, decreased feed
consumption, conjunctival congestion and lymphadenitis symptoms
[12, 13]. Bovine Ehrlichiosis is mainly caused by E. ruminantium.
Transmission occurs from ticks of the genus Amblyomma, especially
A. variegatum and A. habraeum [12]. Forms of the disease that progress
with high mortality within a few hours in the peracute form and within
36–48 hours in the acute form have been reported [13].
Rickettsia genus bacteria has two main groups: the spotted fever
group and the typhus group. In humans, infections from the spotted
fever group can cause symptoms ranging from mild, like fever and
rash, to life–threatening, depending on the specic agent. In ruminant
animals, the infection tends to be self–limiting; therefore, Rickettsia
infection has not received much attention in these animals [14, 15].
The primary vector of diseases in the spotted fever group is infected
ticks. While various tick species of the Dermacentor, Rhipicephalus,
and Amblyomma genera serve as vectors for R. rickettsii in America,
Rhipicephalus sanguineus has been associated with R. conorii in
Europe and the Mediterranean coasts, and Amblyomma ticks have
been associated with R. africae in Africa. In Asia, R. japonica has
been frequently isolated from various tick species belonging to the
Haemaphysalis, Ixodes and Dermacentor genera [16].
The most common protozoan diseases transmitted by vectors in
cattle are caused by Babesia and Theileria species [17, 18]. Babesiosis
is an important parasitic disease for both animal and public health
[19]. Babesia agents reproduce asexually within the erythrocytes of
mammals, and the erythrocytic forms are called piroplasm. Sexual
reproduction of the agent occurs in ticks in the Ixodidae family [20].
Bovine babesiosis is also commonly called Texas Fever or Blood
Urination Disease. Babesia bovis, B. bigemina and B. divergens species
cause clinical babesiosis in cattle [21, 22]. Babesia infections are
inuenced by the host’s age, immune system, co–infection status, and
genetic factors. Symptoms of acute infection include fever, anemia,
hemoglobinuria, jaundice, weakness, lethargy, and anorexia, while
chronic infection may be asymptomatic [20, 23].
Theileriosis is caused by Theileria agents, which are obligate
intracellular protozoa mostly affecting ruminants and transmitted by
ticks. The infection process involves entering the agents into the host’s
lymphocyte or macrophage cells, followed by asexual proliferation and
development into piroplasmic forms found in erythrocytes at a later
stage [24]. Theileria species are transmitted by ticks of the genus
Hyalomma, Rhipicephalus, Dermacentor, Haemaphysalis, Amblyomma
in the family Ixodidae, and ticks of the genus Ornithodorus in the family
Argasidae. Especially T. parva (East Coast Fever) and T. annulata (Tropical
theileriosis) are highly pathogenic species and can cause clinical disease
in cattle. Symptoms may vary depending on factors such as infection with
the pathogenic Theileria agent, tick infestation severity, other pathogen
infections, host’s immune system, age, race, and vaccination status [25].
The rst symptom after a tick starts sucking blood is fever, followed by an
enlargement of the nearest lymph node. Later symptoms include loss of
appetite, increased heart rate, weakness, petechial bleeding, edema in
the lymph and eyelids, decreased milk yield, and jaundice [26]. Although
various studies have been conducted on the molecular epidemiology
of diseases caused by vector–borne Rickettsial pathogens in cattle in
Türkiye, there is still a lack of information regarding these factors [1]. In
the Mugla region, which is included in the scope of the study, no studies
on these diseases in cattle were found. This study aimed to investigate
Anaplasma, Ehrlichia and Rickettsia, Babesia and Theileria species using
molecular methods in blood samples taken from cattle raised in Mugla
province, located in the west of Türkiye.
MATERIALS AND METHODS
Sampling
Blood samples were collected from apparently healthy dairy cattle
between June and September 2023, when vector ticks were also
active. A total of 100 cattle (Bos taurus) blood samples taken from
11 different farms were used as material. Blood samples were taken
from the jugular veins of the animals into 10 mL tubes with di–sodium
ethylenediamine tetra–acetate (EDTA) under aseptic conditions.
Then, each blood sample collected was divided into sterile 1.5 mL
eppendorf tubes and stored at -20°C (Grundig, GRNE 4302, Türkiye)
until genomic DNA isolations were performed.
DNA extraction and molecular analysis
200 μL of blood was used to isolate genomic DNA (gDNA) from the blood
sample taken from each cattle. At this stage, analyses were performed
using a commercial kit (GeneJET Genomic DNA Purication Kit, Thermo
Scientic, Waltham, MA, USA) in accordance with the manufacturer’s
instructions. The gDNAs obtained were stored at -20°C until PCR analysis.
Three different multiplex–PCR reactions (Rxn) were performed
for bacteria A. centrale and A. marginale (Rxn1) [27], Ehrlichia spp.
and Rickettsia spp. (Rxn2) [28], A. capra, A. bovis, A. ovis and A.
phagocytophilum (Rxn3) (TABLE I) [29]. Ready–made PCR mix was used
for multiplex PCR processes (DreamTaq Hot Start Green PCR Master
Mix, Thermo Scientic, Waltham, MA, USA). Rxn1 was programmed as
follows: 3 min at 95°C, 10 s at 98°C, 30 s at 55 °C, 30 s at 72°C (35cycles)
and a nal extension at 72°C for 5 min. Rxn2 was programmed as
follows: 95°C for 1 min, 95°C for 30 s, 56 °C for 30 s, 72°C for 30s
(40 cycles), and a nal extension of 72°C for 7 min. Amplication