Cutaneous leishmaniasis in northwestern Saudi Arabia: identification of sand fly fauna and parasites
Haouas et al. Parasites & Vectors
Cutaneous leishmaniasis in northwestern Saudi Arabia: identification of sand fly fauna and parasites
Najoua Haouas 0 3
Omar Amer 2
Fawwaz Freih Alshammri 1
Shorooq Al-Shammari 2
Latifa Remadi 0 3
Ibrahim Ashankyty 2
0 Laboratoire de Parasitologie-Mycologie Médicale et Moléculaire (LR12ES08), Département de Biologie Clinique B, Faculté de Pharmacie, Université de Monastir , Monastir , Tunisia
1 Dermatology service, King Khaled Hospital , Hail, Kingdom of Saudi Arabia
2 College of Applied Medical Sciences, Clinical Laboratory Sciences Department, University of Hail , Hail, Kingdom of Saudi Arabia
3 Laboratoire de Parasitologie-Mycologie Médicale et Moléculaire (LR12ES08), Département de Biologie Clinique B, Faculté de Pharmacie, Université de Monastir , Monastir , Tunisia
Background: Cutaneous leishmaniasis (CL) is a vector-borne disease transmitted by the bite of an infected sand fly. This disease is highly prevalent in Saudi Arabia where Leishmania major and L. tropica are the etiological agents. In the region of Hail, northwestern of Saudi Arabia, the incidence is about 183 cases/year. However, the epidemiology of the disease in this area is not well understood. Thus, an epidemiological survey was conducted in 2015-2016 to identify the circulating parasite and the sand fly fauna in the region of Hail. Skin lesion scrapings were collected from suspected patients with CL. Methods: The diagnosis was made by microscopic examination of Giemsa-stained smear and PCR. The parasite was identified by PCR and sequencing of the single copy putative translation initiation factor alpha subunit gene. Sand fly specimens were collected and identified morphologically. Total DNA was extracted from the abdomen of female specimens and Leishmania DNA was detected by PCR. Results: Among the 57 examined patients, 37 were positive for CL. The identification of the parasite has revealed the single species Leishmania major. The 384 sand flies were collected belonged to two genera (Phlebotomus and Sergentomyia), six sub-genera and six species. Phlebotomus papatasi, Ph. kazeruni and Sergentomyia clydei were the dominant species. Leishmania DNA was detected in two females of Ph. papatasi two of Ph. kazeruni and one specimen of Sergentomyia clydei. Conclusions: Leishmania major is confirmed to be the etiological agent of cutaneous leishmaniasis in northwestern Saudi Arabia. The molecular detection of Leishmania DNA in Ph. papatasi and Ph. kazeruni supports the potential role of these two species in the transmission of Leishmania. Further epidemiological studies are needed to prove their role and to evaluate the burden of CL in the study region.
Cutaneous leishmaniasis; Leishmania major; Sand fly; Hail; Northwestern Saudi Arabia
Cutaneous leishmaniasis is a parasitic disease caused by
a flagellated protozoan belonging to the genus
Leishmania. It is known to be the ninth largest disease
burden among the 13 parasitic and bacterial neglected
tropical diseases worldwide [
]. Although it is
selfhealing, CL causes skin ulcers and disfiguring scars that
can result in serious social and psychological stigma [
Because of the devastating consequences to the patient,
CL is recognized as a special public health problem. In the
Middle East and across to central Asia, CL is endemic
with an average estimated annual incidence of 321,300
cases . Among countries of this region, Saudi Arabia
was reported as the fourth most endemic focus of
zoonotic CL after Afghanistan, Iran and Pakistan with an
estimated incidence ranging from 9600 to 15,800 cases/year
]. In this region, the first real documented cases date
from 1973 [
]. Many Saudi provinces are endemic for CL
including the Al-Hassa oasis, Al-Madinah Al-Munawarah
and Al Qassim provinces where desert rodents
(Psammomys obesus and Meriones libycus) are the main reservoir
hosts and the vector Ph. papatasi is prevalent [
most endemic areas in Saudi Arabia, the causative
organism was identified as L. major. Cutaneous leishmaniasis
due to L. tropica is less prevalent compared to zoonotic
CL caused by L. major. It occurs within small endemic
foci in the west (Al Madina Al-Munawarah and Al
Qassim) and southwest (high plateau of Aseer) provinces
9, 10, 12–15
Despite the large distribution of this parasitic disease,
studies focusing on the identification of the Leishmania
species and the sand fly fauna are rare. Indeed, the
characterization of the parasite circulating in this
geographical area only started at the beginning of the 1990s.
Since then and up to now only about 224 Leishmania
isolates have been identified to the species level either by
isoenzymatic methods (n = 38 isolates) or using a
molecular approach (from culture and human clinical
samples n = 102 and sand flies n = 84) [
9, 10, 13, 16–18
Several sand fly investigations have described
phlebotomine species composition, their geographical
distribution and their role in disease dynamics in Saudi Arabia.
These surveys have revealed the presence of 25 species
with a predominance of Ph. papatasi in all investigated
areas of the kingdom [
]. Moreover, it was
demonstrated that the geographical distribution of CL coincides
with the distribution of the sand fly vectors Ph. papatasi
and Ph. sergenti [
According to the Saudi Ministry of Heath seven-year
reports (2006–2012), Hail Province, Northwestern Saudi
Arabia, is reported as the fifth most infected region
among the 20 Saudi provinces after El Qassim,
AlMadinah Al-Munawarah, El Hassa and Riyadh [
Indeed, the average reported incidence of CL in Hail was
183 cases/year. Despite the endemic state of Hail
province to CL, no published report is available describing
the epidemiological profile of this disease including the
identification of the parasite and sand fly species and
their geographical distribution.
Here, we aimed to identify both the Leishmania
species and the sand fly species composition in the Hail
region in northwestern Saudi Arabia including the
monthly abundance of circulating phlebotomine species.
Such data are used to understand the structure of this
CL focus and to clarify the role of the sand fly species in
the transmission of the disease and thereby to establish
effective control and preventive measures against this
Description of the study area
Hail is located in the northwestern region of the
Kingdom of Saudi Arabia (between 25°35′–29°00′N, 39°01′–
44°45′E). It is bordered by Al Jouf to the North,
Northern borders to the northeast, Al Qassim and Riyadh to
the south and Tabuk and Al Madina Al Munawara to
the west. Hail covers an area of almost 118,322 km2 and
has a population of 527,000 (Ministry of the Interior,
2013 estimate) (Fig. 1). It is located at 914 m above
mean sea level and has an annual rainfall of 100.6 mm.
The principal part of Hail is composed of the Nafud
Desert, covering about 64,000 km2. The weather system
of Hail Region is arid to extra arid. Summer
temperatures typically rise as high as 50 °C during daytime with
a diurnal variation of about 25 °C. Winter temperatures
hover around freezing at night especially at higher
altitudes although the ground occasionally freezes and
daytime temperatures nearly always reach 25 °C in the sun.
Human CL cases were collected from dermatology
services in different hospitals of the Hail region. For each
patient, socio-demographic (gender, age, geographical
residency of patients and nationality) and clinical data
(number of lesions, precise location, size, clinical aspect
and evolution duration of each lesion) were recorded on
a pre-printed sheet. For each patient, two samples were
collected with the first used to prepare two smears for
Giemsa staining and microscopic examination. The
second sample was stored in phosphate buffered saline
(PBS) at -20 °C for further molecular studies.
Sand flies were captured twice a week from September
2015 to October 2016 in nine geographical locations
within Hail Province (Fig. 1) from where the regional
health authorities reported human CL cases. On each
occasion, thirty sticky traps (A4 white paper sheets
coated with crude Castor oil) were placed in different
biotopes. Indoor (animal shelters) traps were placed at
1.5 m above ground sheltered from air streams. Outdoor
(at the entrance of rodent burrows and in sandy areas
and valleys) traps were put on the ground with a stick as
shown in Fig. 2. Traps were placed at sunset and
collected the next day before sunrise.
Molecular detection and identification of Leishmania parasites
DNA extraction was performed on clinical samples and
female sand flies using the ReliaPrep™ gDNA Tissue
Miniprep System Kit (Promega, Madison, United States),
following the manufacturer’s instructions.
PCR amplification was carried out using the
genusspecific primers of the conserved sequences of the small
subunit ribosomal DNA (SSU-rDNA) gene (PCR-Lei) for
the detection of Leishmania spp. infection according to
the protocol of Spanakos et al. [
]. This PCR is
genusspecific and does not allow the identification of the
species. The forward primer Lei70L was 5′-CGC AAC CTC
GGT TCG GTG TG-3′ and the reverse primer Lei70R
was 5′-CGC GGT GCT GGA CAC AGG GTA-3′. PCR
was performed in a final volume of 50 μl containing 1×
PCR buffer, 1.5 mM MgCl2, 0.2 mM of each
deoxynucleotide, 3 units of Taq DNA Polymerase (all obtained
from Promega) and 100 pM of each primer. PCR was
performed in a PTC 100 Thermal Cycler, with the
following conditions: initial denaturation at 94 °C for
5 min, 40 cycles at 94 °C for 30 s, 65 °C for 30 s and 72 °
C for 30 s and a final elongation step at 72 °C for
10 min. For each experiment, positive (DNA extracted
from Leishmania culture) and negative (Human DNA
extracted from a healthy donor) controls were used.
Products were separated by 2% agarose gel
electrophoresis containing 0.5 mg/ml ethidium bromide and
visualized on a UV transilluminator.
Thereafter, DNA from the positive sample was
reanalyzed using a second set of primers targeting a
coding DNA sequence (CDS) of the putative translation
initiation factor alpha subunit gene according to the
protocol of El Baidouri et al. [
]. Genomic DNA was
amplified by real-time PCR using SYBR Green method
(Light Cycler 480 II, Roche, Rotkreuz, Switzerland). The
amplified products were sequenced on both strands
(Eurofins MWG Operon, Ebersberg, Germany) and the
obtained sequences were blasted using Blastn algorithm
against the “non-redundant” GenBank sequence
database for the identification of Leishmania species.
Phlebotomine sand flies identification
Sand flies were firstly washed with 90% ethanol to
remove excess oil and then kept in labelled vials
containing 70% ethanol. The head and posterior part of the
abdomen of both male and female sand flies were
dissected, cleared in Marc-Andrée solution [
] and then
mounted between slide and coverslip in chloral gum
]. Their abdomens and thorax were
preserved in ethanol for further molecular analysis. Female
and male specimens were morphologically identified by
observing head and genital structures under the
microscope using the morphological keys and characters of
identification proposed by Croset et al. [
] and Leger et
]. For each female specimen, the presence of eggs
(gravid), blood (total or partial engorged) or unfed (no
visible blood in the abdomen) status was recorded.
Leishmania DNA detection and identification from female sand flies
After removal of the head and posterior extremity from
each female specimen, the remainder of the body was
transferred to individual sterilized 1.5 ml vials, from
which genomic DNA was isolated using ReliaPrep™
gDNA Tissue Miniprep System kit (Promega, Madison,
United States), following the manufacturer’s instructions.
The maceration of the insect’s tissues was carried out
with a piston pellet, and the final elution volume was
100 μl. DNA extracts were stored at -20 °C until use.
Five μl of the extracted DNA was used as a template
for the PCR reaction for the detection of a Leishmania
SSU rDNA sequence as described by Spanakos et al.
] and detailed above. Identification of Leishmania
species detected from female sand flies was performed
by the amplification and sequencing of the putative
translation initiation factor alpha subunit locus
according to the protocol of El Baidouri et al. [
Number of recruited human cases
From September 2015 to October 2016, 57 patients with
suspected human CL were included in our study. The
patients ranged from one to 65 years of age, with a
median of 30 years. The sex ratio of men to women was 2.8
(men: n = 42; women: n = 15). Among them, 29
(50.87%) were Saudi and 28 (49.12%) were non Saudi
(Egyptian: n = 10; Indian: n = 8; Pakistani: n = 4;
Sudanese: n = 4; Bangladeshi: n = 1; Filipino: n = 1).
Diagnosis and epidemiological characteristics of cutaneous leishmaniasis
Thirty-seven patients (64.91% of the diagnosed cases)
were diagnosed as cases of CL. Among the 57 diagnosed
patients, 18 (31.57%) had a positive direct examination.
The diagnosis of the 39 remaining negative cases was
confirmed by the detection of Leishmania DNA using
PCR-Lei. Thus, among these 39 negative cases, 19 cases
(48.71%) had shown a positive PCR result confirming
their infection with the Leishmania parasite.
Among the 37 confirmed CL cases, 29 were males
(78.37%), and 8 were females (21.62%). Their age ranged
from three to 65 years old with a median age of 34 years.
The majority (72.97%) of cases occurred in the age
group 15–45 years. CL cases declined for the two other
age groups (i.e. 8.10% in patients < 15 and 18.91% in
patients > 45 years of age).
By comparing the infection rate between Saudi and
non-Saudi patients, 17 (45.94%) were indigenous, and 20
(54.05%) were expatriate workers (Egyptian: n = 6;
Indian: n = 6; Pakistani: n = 4; Sudanese: n = 3; and
Bangladeshi: n = 1). All patients (Saudi and non-Saudi)
confirmed that they had not left the Hail region in the
previous five months before the onset of lesions. The
positive cases were unequally distributed in the different
regions of the Province of Hail. In fact, the majority of
CL cases were from Hail city (n = 22, 59.45%), followed
by Chinan (n = 7, 18.91%). Sporadic cases were from Al
Rodha (n = 2, 5.4%), Baqaa (n = 2, 5.4%), Qina (n = 2,
5.4%), Samirah (n = 1, 2.7%) and Guefar (n = 1, 2.7%).
Clinical features of cutaneous leishmaniasis
Among the positive CL cases, 94.6% had lesions over
exposed parts of the body. The most commonly affected
sites were the lower limbs (n = 21, 56.75%) (leg: n = 13;
foot: n = 8) followed by the upper limbs (n = 10,
27.02%) and face (n = 4, 10.81%). Nevertheless, two CL
lesions (5.40%) were found on the back (n = 1, 2.7%) and
the groin (n = 1, 2.7%).
Among positive CL cases, the size of the lesion varied
between 0.3 and 9 cm; about 40% of positive lesions
were 3–5 cm (n = 15), whereas 27.02% were 1–3 cm.
The lesions of 1 cm and > 5 cm represented 16.21%
each. Among the same group, 22 patients (59.45%) had a
single lesion, 10 patients (27.02%) with 2–5 lesions and
5 patients (13.5%) had 5–20.
The duration of the disease varied between three
weeks to four months. Most CL cases (75.67%)
presented for diagnosis one to two months after the onset
of their skin lesions. However, 5.40% of the patients
diagnosed their disease within three weeks and 18.91%
after more than two months. The detailed data are
summarized in Table 1.
An important clinical polymorphism of the CL lesions
was noticed among the studied series. Among the 37
positive cases of CL, 6 different clinical forms were
noted. The ulcero-crusted form was the most common
(n = 19, 51.35%) followed by the ulcerated form (n = 8,
21.62%), the nodular (n = 4, 10.81%), the pseudotumoral
(n = 3, 8.10%), the vegetant (n = 2, 5.40%) and the
nodulo-papular (n = 1, 2.70%).
Leishmania species identification in clinical samples
Among the 37 positive CL cases, 25 (including 18
positive cases by direct examination and 7 positive cases by
PCR-Lei) were positive for the amplification of the single
copy gene. These PCR products were sequenced and
identified by comparison with the nucleotide-nucleotide
Basic Local Alignment Search Tool (BLAST) (GenBank
DNA sequence database, National Centre for
Biotechnology Information) (www.ncbi.nlm.nih.gov/blast/). All
of the 25 CL cases were identified as L. major. No case
of L. tropica was identified.
Sand fly species composition and abundance
During the entomological survey, a total of 46 weekly
collections were performed from September 2015 to
October 2016. In total, 384 wild-caught phlebotomine sand
flies were captured including 212 males and 172 females.
Among these females, four specimens were blood fed,
and the abdomen of 18 others were filled with eggs.
Regarding sand fly individuals, the highest number of flies
were recorded in Guefar (n = 187), followed by Mrifag
(n = 119) and Al Swifla (n = 27). However, only eight
specimens were collected in Al Hmayria, seven in
Mogag and a single fly in Baqaa.
The collected sand fly specimens were distributed among
two genera [Phlebotomus (n = 292, 76.04%) and
Sergentomyia (n = 92, 23.95%)], six sub-genera [Phlebotomus
(n = 242, 63.02%), Paraphlebotomus (n = 49, 12.76%),
Larroussius (n = 1, 0.26%), Sintonius (n = 81, 21.09%),
Sergentomyia (n = 9, 2.34%), and Grassomyia (n = 2, 0.52%)]
and six species (Ph. papatasi, Ph. kazeruni, Ph. syriacus, Se.
clydei, Se. antennata and Se. dreyfussi) (Tables 2 and 3).
Sergentomyia dreyfussi was exclusively found at the
station of Guefar and Qina (Table 3). Among the
Phlebotomus species caught, Ph. papatasi was the most
predominant (82.87%) species, followed by Ph. kazeruni
(16.78%) and Ph. syriacus (0.34%). For the genus
Sergentomyia, Se. clydei represents 88.04% of the collected
specimens, followed by Se. antennata (9.78%). Se.
dreyfussi was rare and represented just 2.17% of the collected
Sergentomyia flies. All blood-fed female sand flies were
identified as Ph. papatasi.
Overall, the male to female sand fly ratio was 6:5 (212/
172). This sex ratio (M/F) varied according to the sand
fly species and the station (Tables 2 and 3). For all of the
three identified Phlebotomus species, the males
outnumbered the females. However, in case of Se. antennata the
ratio of male/female is 0.5, whereas Ph. syriacus and Se.
dreyfussi were only represented by males.
The monthly abundance (fly/month) was examined for
the three common sand fly species Ph. papatasi, Ph.
kazeruni and Se. clydei (Fig. 3). In general Ph. papatasi
and Se. clydei sand fly species were active from April to
November with increased activity between July and
August indicated by prominent peaks during these two
months. However, Ph. kazeruni species had shown a
peak of activity in May which markedly decreased in
June and July, and the species disappeared from August
when the temperature exceeded 40 °C.
Leishmania DNA detection in female sand flies
Molecular detection of Leishmania was carried out using
individual DNA extract prepared from 75 sandflies
female specimens (44 Ph. papatasi, 15 Ph. kazeruni, 14 Se.
clydei and 2 Se. antennata). Leishmania DNA was
detected using SSU rDNA sequence as described by
Spanakos et al. [
]. The infection rate was calculated as
6.66% (5/75). Among these five infected specimens, two
were Ph. papatasi, two Ph. kazeruni and one Se. clydei.
All of them were non-blood-fed females. The presence
of Leishmania DNA was further confirmed by
sequencing of the PCR products using the same primers as the
The DNA extract of the five infected specimens was
used for a second PCR amplifying a single copy
housekeeping gene to identify the Leishmania species.
Although the PCR was repeated on five separate occasions,
no specimen was found positive for Leishmania DNA.
This is the consequence of the low sensitivity of this
second PCR compared to the first one which amplifies
multi-copy DNA sequence.
Cutaneous leishmaniasis is endemic in Hail. According
to the Ministry of Health of the Kingdom of Saudi
Arabia (2006–2012) reports, about 183 CL cases are
reported each year in this region [
]. Despite its endemic
state, this disease is rarely studied in the Hail region. A
single retrospective study was carried out in 2014
emphasising the extension of this disease in different
parts of Hail [
]. No data are available concerning the
causative species of this disease or the vector. For this
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reason, the purpose of our study was to identify the
Leishmania species causing the disease and the sand flies
species circulating in the Hail region.
To our knowledge, this is the first time that
Leishmania species are identified in this study area. All
positive human CL cases were caused by L. major indicating
the presence of a homogeneous focus of zoonotic
cutaneous leishmaniasis (ZCL) in this study area.
Nevertheless, the absence of human CL cases caused by L.
tropica in our study did not exclude the presence of
small micro-foci of anthroponotic CL (ACL) in Hail
region. Further studies are needed to investigate all Hail
regions to affirm or deny the presence of ACL since this
nosogeographical CL form is less prevalent than ZCL.
This last form is endemic in many regions of Saudi
Arabia including Al-Hassa Oasis, Al-Madinah
AlMunawarah and Al Qassim provinces where desert
rodents (Psammomys obesus and Meriones libycus) are the
main reservoir hosts [
]. However, ACL caused by
L. tropica was reported only in small foci such Aseer in
the southwestern region [
9, 10, 13, 14
Variation in the clinical presentation of CL was
noticed among the positive cases. Six different clinical
forms were noticed. The ulcero-crusted form was the
most common clinical form followed by the ulcerated
form then the nodular form. The same result was
reported in Tunisia by Masmoudi et al. [
] who noticed
54.9% of the diagnosed patients with this clinical form
]. Also, the same study had reported the presence of
nine clinical forms among a studied population of 102
cases of CL. Douba et al. [
] have reported five types of
CL in the region of Aleppo, Syria. Among them, the
two most common types were the papulonodular and
It is important to highlight that the size of the lesion is
directly linked to the duration of infection. Here, small
nodular lesions were noted among patients who have
diagnosed their disease within one month or less since
their onset. It is known that a single Leishmania species
can elicit a range of clinical patterns [
specific factors that influence the clinical outcome of these
infections remain to be completely elucidated but likely
is influenced by host genetics or immune responses [
potentially different sand fly vector species or
] and the presence of Leishmania spp.
]. Clinical polymorphism could also be
explained by the degree of virulence of the Leishmania
strain causing the lesion. Indeed, recent studies have
reported that the degree of virulence of Leishmania
decreases in the presence of a mutation in the gene
encoding a protein BBSome complex or the gene
encoding the enzyme Sphingosine kinase [
Zangger et al. [
] showed that the virulence of L.
guyanensis is due to its co-infection by a virus LRV
(Leishmania RNA virus). Major efforts are still needed
to understand the real causes of the observed clinical
The second part of our study was an entomological
survey of sand flies in the Hail Province of Saudi Arabia.
The goal of this investigation was to identify the species
of sand fly and assess the infection rate in female
specimens. Thus, six species were identified including three
of the genus Phlebotomus (Ph. papatasi, Ph. kazeruni
and Ph. syriacus) and three of the genus Sergentomyia
(Se. antennata, Se. clydei and Se. dreyfussi). Phlebotomus
papatasi was the most abundant species identified in the
current study. This result is consistent with the
epidemiological state of leishmaniasis in Hail where ZCL is
the extended noso-geographical form in this study. It is
known that in Saudi Arabia, ZCL is transmitted to
humans from infected rodent reservoir hosts
(Psammomys obesus and Meriones libycus) through the bites of
the sand fly vector Ph. papatasi [
]. The same species
was also reported as the predominant sand fly species in
many parts of the Kingdom of Saudi Arabia, such as
AlMadinah Al-Munawarah [
], Jeddah , Riyadh
] and Al-Qassim [
]. Since 1985, Killicki-Kendrick
et al. [
] have isolated L. major MON-26 from a
specimen of Ph. papatasi is proving its role as a vector of
ZCL. A study on the seasonal abundance of Ph. papatasi
in Riyadh had demonstrated that the greatest number of
sand flies occurred most commonly in the summer with
two peaks in June and September [
It is important to highlight that a complete survey
carried in Saudi Arabia to study its phlebotomine sand fly
fauna was that of Lewis & Buttiker [
] in 1982. During
this survey, 25 phlebotomine species were identified in
the whole of Saudi Arabia [
]. Inside this study area,
sand fly fauna was identified in the Hail region. Between
1975 and 1980 only 14 specimens were collected in Hail,
and only two species were identified: Se. clydei (79%)
and Se. antennata (21%). During the same investigation,
the second survey in Hail was carried in 1981, which
identified Ph. kazeruni, Ph. syriacus, Ph. naqbenius, Se.
christophersi, Se. clydei, Se. fallax and Se. tiberiadis.
Since 1981, no study has been carried out in this region.
Here, we identify for the first time Se. dreyfussi from two
specimens. More entomological investigations are
crucial to study the abundance of this species in the
different parts of Hail.
Assessment of the Leishmania infection rate in sand
flies is of paramount importance, and PCR-based
approaches have been successfully used for detection of
parasite DNA in sand flies [
]. The detection of
Leishmania DNA in two unfed females of Ph. papatasi
caught in a rural area of Hail provides additional
evidence in favour of its role as a potential vector for L.
major in this study area. Also, the detection of
Leishmania DNA in two unfed Ph. kazeruni specimens (a
close species to Ph. sergenti) suggests a potential role of
Paraphlebotomus spp. in the transmission of L. tropica.
It should be pointed out that the molecular detection of
Leishmania DNA in sand flies is not enough to
incriminate these insects as vector conclusively. Such finding
could be explained by recent feedings on reservoirs
resulting in parasite DNA remnants after blood meal
digestion. Thus, isolation of the parasite from the
midgut of these sand flies and its isoenzymatic identification
remain mandatory to confirm this hypothesis. Finally,
one specimen of Se. clydei was also found to be positive
for the detection of Leishmania. In the Old World,
phlebotomine species of the genus Sergentomyia are known
as reptile-biting sand flies transmitting Sauroleishmania
or the “lizard Leishmania” parasite [
some recent investigations in Tunisia and Portugal have
reported the detection of L. major and L. infantum DNA
in Se. minuta [
48, 52, 53
] suggesting its potential role in
the transmission of mammal-infecting Leishmania. Also,
Senghor et al. and Berdjane-Brouk et al. suggested that
Sergentomyia species could be involved in the
transmission of L. major and L. infantum in Africa [
confirm this hypothesis, it will be crucial to demonstrate
that Sergentomyia species feed on humans and can
support the complete development of the parasite in natural
conditions after the digestion of an infectious blood meal.
In conclusion, L. major is the parasitic agent responsible
for CL in Hail, Saudi Arabia. It exhibits a large clinical
polymorphism and consequently should be included in
the differential diagnosis of many common and
uncommon dermatological diseases. The life-cycle of L. major
is still not elucidated in this area but the detection of
Leishmania DNA in two specimens of Ph. papatasi is
evidence for the role of this sand fly species in the
transmission of L. major. Further studies including the
isolation and identification of the parasite from sand fly are
needed to prove this hypothesis. Entomological data
showed that Ph. papatasi, Se. clydei and Ph. kazeruni are
the most dominant species in our collection. The
detection for the first time of Leishmania DNA in two
specimens of Ph. kazeruni supports the vector status of this
species. Our investigation has provided a valuable set of
data about the epidemiological features of CL in
Northwestern Saudi Arabia. Nevertheless, further studies in
humans, vectors and potential reservoirs are needed to
determine the burden of cutaneous leishmaniasis in the
whole of Hail.
CL: cutaneous leishmaniasis; ZCL: zoonotic cutaneous leishmaniasis
We thank all the staff of the Control of Communicable Diseases and Vectors
Administration in Hail region (Mr Hichem Al Tamimi, Marzouk Al-Shammary,
Abdul Rahman Al-Salah and Salem Al-Nozha) for their contribution in field
work and sand fly collection. Also, we are grateful to Dr Christophe Ravel for
identifying Leishmania species and Dr Jérôme Depaquit for helping in sand
fly morphological identification. Finally, we sincerely acknowledge Mr
Slaheddine Amor for his technical help.
Ethical approval and consent to participate
This study was embedded in activities of the KACST research project, which
has received ethical approval from the CAMS Medical Research Ethics
Committee in University of Hail, Hail, KSA (reference no. 10/Apr/015).
This study received financial support from the King Abdulaziz City for
Science and Technology, Kingdom of Saudi Arabia, ID number SG-35-181.
The funders had no role in study design, data collection and analysis,
interpretation of the data, or writing the manuscript.
Availability of data and materials
The data sets supporting the conclusions of this article are included within
the article. The sequences are submitted in the GenBank database under the
accession numbers MG018949–MG018957.
This work was accomplished by the contribution of all authors. NH has been
involved in the study design, sand fly identification, results analysis and the
manuscript drafting. OA has provided local contacts, contributed to the
study design and manuscript reviewing. FFA has contributed to the
collection of human samples and the annotation of the human clinical cases.
SA was involved in the collection and identification of sand flies. LR has
performed the molecular study. IA has contributed in data analysis. All
authors read and approved the final manuscript.
Consent for publication
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