Choleoeimeria bunopusi sp. n. is described from the gall bladder of the tuberculated gecko Bunopus tuberculatus in Saudi Arabia. The prevalence of infection was 13.3% (2/15). Oocysts were ellipsoidal and measured 31 (30–33) × 21 (20–22) μm. Sporocysts were dizoic, elliptical in shape and measured 12 (11–13) × 7 (6–8) μm. The endogenous development was confined to the gall bladder epithelium. The hypertrophic parasitized biliary epithelium either remained in one layer or became stratified. Meronts, gamonts, and young oocysts were detected

ABDEL-BAKI, Abdel-Azeem S.

AL OTAIBI, Mamdooh S. A.

AL-QURAISHY, Saleh

Portal Czasopism Naukowych (E-Journals)

Acta Protozool. (2013) 52: 267–272 
http://www.eko.uj.edu.pl/ap
doi:10.4467/16890027AP.13.023.1315ACTA
PROTOZOOLOGICA
Choleoeimeria bunopusi sp. n. (Apicomplexa: Eimeriidae) Infecting the 
Gall Bladder of the Tuberculated Gecko Bunopus tuberculatus (Reptilia: 
Gekkonidae) from Saudi Arabia
Saleh AL-QURAISHY1, Abdel-Azeem S. ABDEL-BAKI1,2 and Mamdooh S. A. AL OTAIBI1
1 Zoology Department, College of Science, King Saud University, Riyadh, Saudi Arabia; 2 Zoology Department, Faculty of Science, 
Beni-Suef University, Egypt
Abstract. Choleoeimeria bunopusi sp. n. is described from the gall bladder of the tuberculated gecko Bunopus tuberculatus in Saudi Arabia. 
The prevalence of infection was 13.3% (2/15). Oocysts were ellipsoidal and measured 31 (30–33) × 21 (20–22) µm. Sporocysts were dizoic, 
elliptical in shape and measured 12 (11–13) × 7 (6–8) µm. The endogenous development was confined to the gall bladder epithelium. The 
hypertrophic parasitized biliary epithelium either remained in one layer or became stratified. Meronts, gamonts, and young oocysts were 
detected.
Key words: Coccidia, oocyst, Choleoeimeria, endogenous stages, Gekkonidae, Sauria.
Address for correspondence: Abdel-Azeem S. Abdel-Baki, Zo-
ology Department, College of Science, King Saud University, 
Saudi Arabia, P.O. Box 2455, Riyadh 11451, Saudi Arabia; Fax: 
00966014678514; E-mail: azema1@yahoo.com 
INTRODUCTION
Coccidians of the family Eimeriidae have been re-
corded in all classes of vertebrates, including reptiles 
(Lainson et al. 2008). In general, the taxonomy of the 
reptilian eimeriids coccidia is controversial (Jirků et 
al. 2002). Traditionally, their classification into genera 
and families is based on the morphology of the exog-
enous stages (oocysts) and on the endogenous stages 
of the life cycle (Jirků et al. 2002, Abdel-Baki et al. 
2008). Many eimeriids coccidian species parasitizing 
reptiles have been identified according to the morphol-
ogy of their oocysts only, however, regardless of their 
endogenous development (Abdel-Baki et al. 2009). 
Modrý and Jirků (2006) and Abdel-Baki et al. (2009) 
have stressed the necessity of studying the endogenous 
stages of eimeriid coccidia as a fundamental tool in al-
locating them to a particular genus.
Paperna and Landsberg (1989), meanwhile, gave 
a new generic name, Choleoeimeria, to those Eimerii-
dae in reptiles which underwent a peculiar development 
in the biliary epithelium of the host. Species of the ge-
nus Choleoeimeria are biliary Eimeria-like coccidia of 
reptiles. Morphologically they are similar to the Eime-
ria to which they are closely related. Oocysts of all the 
described species of Choleoeimeria do share similar 
basic characteristics in their endogenous development 
S. Al-Quraishy et al. 268
in the biliary epithelium, sporulation in the gall bladder, 
their elongated oocysts, and the presence of four dizoic 
sporocysts which lack Stieda and sub-Stieda bodies 
and possess a bi-valved structure (Lainson 2003). Ac-
cordingly, herein we describe the present biliary coc-
cidium as a new species of the genus Choleoeimeria 
from the gall bladder of Bunopus tuberculatus, based 
on the morphology of its oocysts and its endogenous 
development.
MATERIALS AND METHODS
Fifteen adult tuberculated gecko Bunopus tuberculatus Blan-
ford, 1874 were collected during a parasitological survey in Al-
Thumamah (24°41′N, 46°42′E), in Riyadh, in the central region of 
Saudi Arabia during July and August, 2012. These animals were 
caged separately and infection was detected by demonstration of 
oocysts in the faeces. Positive geckos were then killed by chloro-
form and the infection in the gall bladder was verified by micro-
scopic examination of bile collected by puncturing the gall bladder 
with a finely pointed glass pipette. To determine the site of infec-
tion, the entire gall bladder of each animal was then fixed in 10% 
neutral buffered formalin, together with portions of liver, spleen, 
kidney, lung, and intestine. Fixed tissues were processed for his-
tological examination; they were sectioned and stained with he-
matoxylin and eosin (H & E). All the developmental stages were 
observed and photographed using an Olympus BX51 microscope 
with an Olympus DP71 camera. Measurements were made using 
an eyepiece micrometre with an oil-immersion lens; they are given 
here in micrometres (µm; mean ± standard deviation), followed by 
the range in parentheses and the length/width ratio.
RESULTS
The prevalence of infection was 13.3% (2/15). The 
infected gall bladder was a deep blue colour, and the 
bile was turbid due to the presence of large numbers of 
free-floating oocysts at different stages of maturation 
with associated cell debris. 
Description
Choleoeimeria bunopusi sp. n. (Figs 1–9)
Mature oocysts were transparent and appeared ellip-
soidal in shape (Figs 1–5, 9). They measured 31 ± 0.7 
(30–33) × 21 ± 0.6 (20–22) µm with length/width ra-
tio1.5 (1.4–1.6) µm. The oocyst wall was smooth, about 
1.0 µm thick, colourless and with no micropyle or stria-
tions (Figs 2–4). This wall was composed of two lay-
ers: an outer, very fine, membrane and a thicker inner 
one (Figs 3, 4). There was no oocyst residuum or po-
lar body. Each oocyst contained four dizoic sporocysts 
(Figs 1–4). They were elliptical in shape with no Stieda 
body. These sprorocysts measured 12 ± 0.6 (11–13) × 7 
± 0.5 (6–8) µm length/width ratio 1.4 (1.3–1.6) µm. The 
sporozoites were banana-shaped (Figs 4, 5) and meas-
ured 14 × 4 μm. Histological examination revealed all 
endogenous stages, including meronts, macrogamonts, 
and microgamonts in the gall bladder epithelium of in-
fected lizards (Figs 6–8). No endogenous stages were 
observed in other organs. Infected epithelial cells were 
displaced from the mucosa toward the lumen, usually 
maintaining contact with the basal membrane only by 
a thin pedicle (Figs 7, 8). The hypertrophic parasitized 
biliary epithelium either remained in one layer (Figs 
6, 7) or became stratified (Fig. 8). Mature meronts were 
nearly rounded to elliptical in shape, measured 16 (15–
17) × 12 (11–13) µm and were estimated to produce 
9–15 merozoites (Figs 6, 8). Mature microgamonts 
measured approximately 18 (17–19) × 15 (14–16) µm 
and were estimated to produce over 50 microgametes 
(Fig. 7). Macrogamonts were mostly spherical and 
measured 16 (15–18) µm in diameter (Fig. 8).
Taxonomic summary
Type host: Bunopus tuberculatus Blanford, 1874.
Type locality: Al-Thumamah (24°41′N, 46°42′E), 
in Riyadh, in the central region of Saudi Arabia.
Prevalence: Two of 15 (13.3%).
Site of infection: Gall bladder.
Sporulation: Sporulation endogenous; both sporu-
lated and unsporulated oocysts were found in the gall 
bladder lumen and intestinal contents prior to being 
voided in the faeces.
Type material: Photosyntypes of the sporulated 
oocysts and syntypes of oocysts of Choleoeimeria bu-
nopusi in gall bladder content of Bunopus tuberculatus 
– sample in 80% ethanol are deposited at Protistologi-
cal Collection of the Inst. of Parasitology ASCR, Insti-
tute of Parasitology, Biology Centre, Academy of Sci-
ences of the Czech Republic with associated collection 
number: IP ProtColl P22.
As well as Photosyntypes and one slide of H & E 
stained endogenous stages in the gall bladder epithe-
lia are deposited in Hungarian Natural History Mu-
seum with associated collection number: Par. Lab. 
HNHM-70393.
Etymology: The specific epithet is derived from the 
host name Bunopus tuberculatus.
New Choleoeimeria from Gecko 269
Figs 1–5. Photomicrographs of freshly collected oocysts of Choleoeimeria bunopusi sp. n. in different stages of development obtained from 
the gall bladder of Bunopus tuberculatus. Mature oocysts surrounded with outer layer (OL) and inner layer (IL) membrane and containing 
four sporocysts (S). Each sporocyst have two sporozoites (Sp) with sporocyst residuum (SR). Scale bars: 10 µm. 
Remarks
According to Duszynski and Wilber (1997), a new 
coccidian species should be compared in detail with 
other coccidian species that are feature-similar and be-
long to the same host family. So far, six Choleoeimeria 
species have been described from the family Gekko-
nidae hosts (Table 1). These species include Choleoe-
imeria faviviridis Setna and Bana 1935 (Paperna and 
Landsberg 1989); Choleoeimeria turcicus Upton et al. 
1988 (Paperna and Landsberg 1989), Choleoeimeria 
pachydactyli Paperna and Landsberg 1989; Choleoe-
imeria rochalimai Carini and Pinto 1926 (Lainson and 
Paperna 1999); Choleoeimeria xiangmaii Paperna 2007 
and Choleoeimeria heteronotis Paperna 2007. Compar-
ing these species, C. faviviridis have thinner oocysts 
(11–14 vs. 17–20) with a larger length/width ratio 
(1.8–2.4 vs. 1.4–1.6) and smaller sporocysts (7–9 vs. 
11–13), while C. turcicus have relatively longer oocysts 
(35–41 vs. 30–33) and a relatively larger oocyst length/
S. Al-Quraishy et al. 270
Figs 6–8. Endogenous stages of Choleoeimeria bunopusi sp. n. at different stages of development in the epithelium of the gall bladder of 
Bunopus tuberculatus. 6 – mature meront (M); 7 – microgamont (Mi); 8 – macrogamont (Ma). Arrows refer to the infected epithelial cells 
that remain in contact with the basal membrane by a thin pedicle. Scale bars: 10 μm. 
width ratio (1.9–2.3 vs. 1.4–1.6). Similarly, C. pachy-
dactyli have thinner oocysts (11–17 vs. 20–22) with a 
relatively larger length/width ratio (2.05 vs. 1.5). The 
oocysts of C. pachydactyli have a small knob occur-
ring at one pole. Although, C. rochalimai have a very 
similar oocyst size to that observed here, they differ 
in that their oocysts frequently possess a conspicuous 
polar body and have shorter pear-shaped sporocysts 
(7.5–10.0 vs. 11–13). Meanwhile, C. xiangmaii and C. 
heteronotis can be differentiated by their thinner oo-
cysts (12.5–16.2, 16.2–17.5 respectively vs. 20–22), 
larger oocyst length/width ratios (1.77–2.60, 1.85–2.10 
respectively vs. 1.4–1.6) and shorter sporocysts (7.5–
10.0, 8.7–10.0 respectively vs. 11–14). During the pre-
sent study, endogenous stages were contained either in 
a single layer bile epithelium or a stratified one. In some 
geckoes Choleoeimeria were found to exhibit a single 
layer of infection, as in the case of C. xiangmaii and C. 
heteronotis (Paperna 2007). In other geckoes, however, 
infection induces stratified rather than single layered 
New Choleoeimeria from Gecko 271
Ta
bl
e 
1.
 C
om
pa
ra
ti
ve
 d
es
cr
ip
ti
ve
 m
ea
su
re
m
en
ts
 (
μm
) 
of
 C
ho
le
oe
im
er
ia
 b
un
op
us
i s
p.
 n
. w
ith
 m
or
ph
ol
og
ic
al
ly
 si
m
ila
r s
pe
ci
es
 fr
om
 h
os
ts
 o
f t
he
 fa
m
ily
 G
ek
ko
ni
da
e.
Sp
ec
ie
s
H
os
t
O
oc
ys
t s
ha
pe
 [L
/W
]
O
oc
ys
t s
iz
e
Sp
or
oc
ys
t
sh
ap
e 
[L
/W
]
Sp
or
oc
ys
t s
iz
e
C
ho
le
oe
im
er
ia
 fa
vi
vi
ri
di
s (
Se
tn
a 
an
d 
B
an
a 
19
35
) 
Pa
pe
rn
a 
an
d 
La
nd
sb
er
g 
19
89
 
H
em
id
ac
ty
lu
s f
av
iv
ir
id
is
(?
) 1
.8
–2
.4
25
–3
4 
× 
11
–1
4
(?
/?
)
7–
9 
× 
5–
7
C
ho
le
oe
im
er
ia
 tu
rc
ic
us
 (U
pt
on
 e
t a
l. 
19
88
) P
ap
er
na
  
an
d 
La
nd
sb
er
g 
19
89
H
em
id
ac
ty
lu
s t
ur
ci
cu
s
C
yl
in
dr
oi
d 
2.
1 
(1
.9
–2
.3
) 
38
.2
 (3
5–
41
) ×
 1
7.
9 
(1
7–
20
) 
Sp
he
ric
al
 to
 
el
lip
so
id
al
 (?
)
11
 (1
0–
12
) ×
 8
.8
 (8
.4
–9
.4
) 
C
ho
le
oe
im
er
ia
 p
ac
hy
da
ct
yl
i P
ap
er
na
 a
nd
 L
an
ds
be
rg
 
19
89
 
Pa
ch
yd
ac
ty
lu
s c
ap
en
si
s
C
yl
in
dr
ic
al
 2
.0
5 
28
.3
 (2
5–
31
) ×
 1
3.
9 
(1
1–
17
)
(?
/?
)
11
.4
 (1
0.
14
–1
2.
6)
 ×
 6
.9
 (6
.5
–7
.2
)
C
ho
le
oe
im
er
ia
 ro
ch
al
im
ai
 (C
ar
in
i a
nd
 P
in
to
 1
92
6)
 
La
in
so
n 
an
d 
Pa
pe
rn
a 
19
99
H
em
id
ac
ty
lu
s m
ab
ou
ia
C
yl
in
dr
ic
al
 
1.
6 
(1
.3
–1
.7
)
29
.1
 (2
6.
2–
31
.2
) ×
 1
7.
8 
(1
6.
2–
18
.7
)
Pe
ar
-s
ha
pe
d 
(?
)
9.
0 
(8
.7
–1
0.
0)
 ×
 7
.5
 (6
.2
–7
.5
)
C
ho
le
oe
im
er
ia
 x
ia
ng
m
ai
i P
ap
er
na
 2
00
7
H
em
id
ac
ty
lu
s f
er
na
tu
s
O
bl
on
g 
1.
97
 (1
.7
7–
2.
60
)
29
.7
 (2
8.
7–
32
.5
) ×
 1
5.
1 
(1
2.
5–
16
.2
)
El
lip
so
id
al
 to
 
su
bs
ph
er
ic
al
 (?
)
9.
3 
(7
.5
–1
0.
0)
 ×
 5
.7
 (5
.0
–7
.5
)
C
ho
le
oe
im
er
ia
 h
et
er
on
ot
is
 P
ap
er
na
 2
00
7
H
et
er
on
ot
ia
 b
in
oe
i
O
bl
on
g 
1.
94
 (1
.8
5–
2.
10
)
32
.8
 (3
2.
5–
33
.7
) ×
 1
6.
9 
(1
6.
2–
17
.5
)
El
lip
so
id
al
1.
3 
(1
.1
–1
.6
)
9.
9 
(8
.7
–1
0.
0)
 ×
 7
.6
 (6
.2
–7
.7
)
C
ho
le
oe
im
er
ia
 b
un
op
us
i s
p.
 n
. (
th
e 
pr
es
en
t s
tu
dy
)
Bu
no
pu
s t
ub
er
cu
la
tu
s
El
lip
so
id
al
1.
5 
(1
.4
–1
.6
)
31
 (3
0–
33
) ×
 2
1 
(2
0–
22
)
El
lip
so
id
al
1.
3 
(1
.2
–1
.5
)
12
 (1
1–
13
) ×
 7
 (6
–8
)
? 
N
o 
da
ta
, [
L
/W
] 
le
ng
th
/w
id
th
 r
at
io
bile epithelium, as in the case of C. faviviridis (Paperna 
and Landsberg 1989) and C. rochalimai (Lainson and 
Paperna 1999).
DISCUSSION
Bovee and Telford (1965) observed two distinct 
groups of Eimeria among lizards based on differences 
in the shape of the oocyst as expressed by its length/
width ratio. The first group has ellipsoidal or cylin-
drical oocysts with a length/width ratio (> 1.4). This 
group also exhibit endogenous sporulation and undergo 
endogenous development in the gall bladder or biliary 
ducts. The second group, on the other hand, has spheri-
cal or rounded oocysts with a length/width ratio (< 1.4). 
This group exhibits exogenous sporulation with the en-
dogenous stages inhabiting the intestinal epithelium.
In 1989, Paperna and Landsberg proposed genus 
Choleoeimeria as a new genus to comprise some tet-
rasporocystic Eimeria-like coccidia infecting the gall 
bladders of reptiles. Use of this new genus has not 
gained wide acceptance because we know very little 
about the endogenous location and development of the 
Fig. 9. Composite line drawing of sporulated oocyst. Scale bar: 
10 µm.
S. Al-Quraishy et al. 272
vast majority of named Eimeria species (Assmundsson 
et al. 2006). Recently, however, molecular studies have 
increased the support for the new genus. For example, 
Morrison (2009) has shown that Eimeria are para-
phyletic and that the single species currently grouped 
within genus Eimeria should in fact be placed in several 
different genera. Furthermore, phylogenetic analysis, 
based on the nucleotide sequence of small sub-unit ri-
bosomal RNA genes, has confirmed the separate status 
of the genus Choleoeimeria (Jirku et al. 2002).
In the present work, we have demonstrated that 
the gall bladder was the only site for the endogenous 
development of C. bunopusi sp. n. and that no endog-
enous stages were detected in intestine. Therefore, we 
have followed Paperna and Landsberg (1989) and Mo-
dry and Jirku (2006) in classifying the biliary coccid-
ium described in this study as a member of the genus 
Choleoeimeria.
Acknowledgement. The authors extend their appreciation to 
the Deanship of Scientific Research at King Saud University 
for funding the work through the research group project number 
RGP–VPP-002.
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Received on 8th January, 2013; revised on 11th March, 2013; ac-
cepted on 11 April, 2013


Choleoeimeria bunopusi sp. n. (Apicomplexa: Eimeriidae) Infecting the Gall Bladder of the Tuberculated Gecko Bunopus tuberculatus (Reptilia: Gekkonidae) from Saudi Arabia

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Choleoeimeria bunopusi sp. n. (Apicomplexa: Eimeriidae) Infecting the Gall Bladder of the Tuberculated Gecko Bunopus tuberculatus (Reptilia: Gekkonidae) from Saudi Arabia

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