Acta Zoologica (Stockholm) doi: 10.1111/azo.

12173

Microanatomy of the digestive system of Supachai’s

caecilian, Ichthyophis supachaii Taylor, 1960 (Amphibia:
Gymnophiona)
Itsares Muikham,1 Nopparat Srakaew,1 Kannika Chatchavalvanich1 and Pramote Chumnanpuen1,2

1
Department of Zoology, Faculty of
Science, Kasetsart University, Bangkok
10900, Thailand; 2Computational
Biomodelling Laboratory for Agricultural
Science and Technology (CBLAST),
Kasetsart University, Bangkok 10900,
Thailand
Keywords:
microanatomy, histochemistry, digestive
system, caecilian, amphibian
Accepted for publication:
22 July 2016
Abstract
Muikham, I., Srakaew, N., Chatchavalvanich, K. and Chumnanpuen, P. 2016.
Microanatomy of the digestive system of Supachai’s caecilian, Ichthyophis supachaii
Taylor, 1960 (Amphibia: Gymnophiona). — Acta Zoologica (Stockholm) 00: 1–19.
The gross anatomy and microanatomy of the digestive system of Ichthyophis
supachaii were investigated. The microscopic structures of the digestive system
are similar to those in other caecilians. Functional and developing teeth are pre-
sent in adults. The tongue contains the genioglossus muscle. The digestive tract
is elongated and consists of the mucosa, submucosa, muscularis and adventitia/
serosa. The oesophagus contains longitudinal folds and lacks oesophageal
glands. We report for the first time the caecilian gastric rugae and specific local-
ization of oxynticopeptic cells in the anterior gastric region. The intestinal
folds are exclusively present in the anterior intestinal region. The liver comprises
30–40 incomplete hepatic lobes, lying in an imbricate manner. Each lobe is
enveloped by haematopoietic tissue that produces and delivers blood cells into
sinusoids. Hepatic parenchyma is organized into anastomosing, two-cell-thick
plates, having sinusoids at the basal domain and bile canaliculi at the apical
domain of hepatocytes. Pigment cells are scattered inside sinusoids. The
pancreas contains pancreatic acini interspersed with islets of Langerhans. The
gallbladder proper is thin and continuous with the cystic duct wall. Neutral and
carboxylated acid mucosubstances are secreted along the digestive tract, while
sulphated mucosubstances are not produced by the stomach and anterior
intestinal regions.
Nopparat Srakaew and Pramote Chumnanpuen, Department of Zoology,
Faculty of Science, Kasetsart University, Bangkok 10900, Thailand. E-mails:
fscinrsr@ku.ac.th (NS) and pramote.c@ku.ac.th (PC)

The caecilian digestive system comprises the digestive tract

Introduction
Caecilians (order Gymnophiona) are the least studied of the
three lineages of extant amphibians primarily because almost
all caecilians have a subterranean lifestyle and they are geo-
graphically restricted to the tropics and subtropics (Kamei
et al. 2012; San Mauro et al. 2014). Consequently, many
aspects of the caecilian biology are poorly understood, includ-
ing the digestive system that contributes to body homoeostasis
with major functions of transporting food along the digestive
tract, mechanically and enzymatically digesting food, absorb-
ing water and nutrients derived from digested food and elimi-
nating indigestible residues as faeces.
and organs associated with the digestive tract (Exbrayat and
Estabel 2006). Basic knowledge on the microscopic structures
of the caecilian digestive system has been gathered from sev-
eral studies of various species (Wiedersheim 1879; Sarasin
and Sarasin 1887; Chatterjee 1936; Junqueira et al. 1999;
Exbrayat and Estabel 2006). However, histological descrip-
tions of the whole digestive system from a single species are
still not available.
The goal of this study was to describe the microanatomy
of the digestive system of Ichthyophis supachaii (Supachai’s
caecilian), which is distributed in southern Peninsular Thai-
land and probably Malaysia (van Dijk et al. 2004; Kupfer and

© 2016 The Royal Swedish Academy of Sciences 1

Digestive system of Supachai’s caecilian  Muikham et al.
M€ uller 2004). Improved fundamental knowledge on the
structures of the caecilian digestive system will contribute to a
better understanding of the digestive system of caecilians in
comparison with other amphibians.
Materials and Methods
Wild I. supachaii (three males and two females) were captured
on Phra Thaew mountain, Phuket Province, southern
Thailand. Species identification was based on Kupfer and
M€ uller (2004). The animal use protocol for small, non-
laboratory animals was not required by Kasetsart University
at the time of the study. Specimens were immediately trans-
ported to the laboratory without feeding and euthanized using
0.1% tricaine methanesulfonate (MS222) in sodium bicar-
bonate buffer (pH 7.0) according to Summers and O’Reilly
(1997). Animals were weighed, and their body lengths were
measured. They were decapitated and the abdomen incised
midventrally from the cloacal orifice to the anterior aspect of
the body. The digestive organs were examined for their gross
morphology. The digestive tract (mouth, pharynx, oesopha-
gus, stomach and intestine) and organs associated with the
digestive tract (liver, pancreas and gallbladder) were removed
and placed into Bouin’s solution for 24 h. Tissues were pro-
cessed according to the standard histological protocol for
paraffin embedding (Bancroft and Gamble 2008) and cut into
5-lm-thick slices. Paraffin sections were stained with haema-
toxylin–eosin (H&E), Masson’s trichrome for collagen fibres,
periodic acid–Schiff–haematoxylin (PAS-H) for neutral
mucosubstances/glycoproteins, alcian blue (AB) pH 1.0 for
sulphated acid mucosubstances/glycoproteins and AB pH 2.5
for carboxylated acid mucosubstances/glycoproteins (Bancroft
and Gamble 2008). In a separate study of the carbohydrate
histochemistry, paraffin sections were pretreated with diastase
in phosphate buffer saline for glycogen digestion prior to
PAS-H histochemistry (D-PAS; Luna 1968). Histological
sections were viewed under a light microscope, and photomi-
crographs were taken.
Results
Gross anatomy of the digestive system
Animals designated as C1–C5 have mean body length (SD)
of 21.32  3.20 cm and mean body weight of 7.28  2.52 g
(Table 1). The digestive system of I. supachaii is composed of
the digestive tract (oral cavity, pharynx, oesophagus, stomach,
intestine and cloaca) and accessory digestive organs (liver,
pancreas and gallbladder; Fig. 1). Almost all parts of the
digestive tract appear as a straight tube except the anterior
region of the intestine, which is slightly convoluted. The
mouth of I. supachaii is anteroventral. The oral cavity contains
the oral floor, palate, teeth and tongue. The tongue is rela-
tively flat and appears as a small appendage attached to the
posterior region of the oral floor and continuous anteriorly
2 © 2016 The Royal Swedish Academy of Sciences
Acta Zoologica (Stockholm) 0: 1–19 (August 2016)
with the oral mucosa. Teeth are organized into four U-shaped
dental rows: two rows (premaxillary–maxillary and vomeropa-
latine tooth rows) in the upper jaw and two rows (dentary and
splenial tooth rows) in the lower jaw. Teeth within the same
dental rows and among the dental rows are of similar size.
Each tooth has a pointed crown that curves posteriorly. It is
difficult to determine the dental cusps via macroscopic obser-
vation due to the limited magnification and resolution. The
pharynx communicates with the oral cavity anteriorly and
with the oesophagus posteriorly. There is no sharp demarca-
tion between the oesophagus and the stomach. The stomach
is dilated and lies parallel to the liver (Fig. 1). The gastroin-
testinal junction is the approximate area posterior to the stom-
ach where the wider gastric calibre becomes relatively
narrower. There is no distinct constriction of the digestive
tract at the junction area. The pancreas is bilobed. The ante-
rior pancreatic lobe is adjacent to the spleen, while the poste-
rior lobe is close to the anterior region of the intestine
(Fig. 1). The liver is elongated at the dorsal region, while it
has incomplete 30–40 hepatic lobes (leaflets) at the ventral
region (Fig. 1). These hepatic lobes are arranged in an imbri-
cate manner in the anteroposterior direction (Fig. 1). The
most anterior hepatic lobe is at the level of the heart, while the
most posterior lobe is close to the gallbladder (Fig. 1).
Numerous black melanized spots are scattered throughout the
liver (Fig. 1). The gallbladder is ellipsoid, dark green and
located just anterior to the most posterior hepatic lobe
(Fig. 1). The anterior region of the intestine appears relatively
convoluted compared to the straight posterior intestinal
region, the latter opening into the cloaca.
Microanatomy of the digestive tract
Oral cavity. The anterior regions of the oral floor and of the
palate are covered by stratified squamous non-keratinized
epithelium (Figs 2A, 3A and 5A,B), while the posterior
regions of the two structures are lined by stratified cuboidal
epithelium with unicellular goblet cells (Figs 2, 3 and 5A,B).
In addition, simple tubular mucous glands open into the pos-
terior oral and palatine regions (Figs 2 and 3). These glands
are composed of basal cells and mucous cells with spherical or
Table 1 Body size of Ichthyophis supachaii
Body size
Animal Sex Weight (g) Length (cm)
C1 M 8.78 23.50
C2 M 8.25 21.80
C3 M 8.32 23.10
C41 F 8.26 22.50
C51 F 2.78 15.70
Mean  SD 7.28  2.52 21.32  3.20
1
Animals C4 and C5 are animals F1 and F2, respectively (see table 1 in
Pewhom et al. 2015).
Acta Zoologica (Stockholm) 0: 1–19 (August 2016)
Fig. 1—Gross morphology of the Bouin-fixed digestive system of
Ichthyophis supachaii.
ellipsoid shapes and the basally located nucleus (Figs 2B and
3B). The supranuclear cytoplasm of the mucous cells is dis-
tended due to congregated mucinogen granules that cause the
slightly acidophilic cytoplasm (Figs 2B and 3B). These cells
produce neutral, carboxylated acid and sulphated acid muco-
substances, as demonstrated by carbohydrate histochemistry
(Figs 2C–F and 3C–F; Table 2). Pretreatment of the oral tis-
sues with diastase does not change PAS reactivity, compared
with the untreated tissues (Table 2). The lamina propria of
the oral floor and of the palate contains loose connective tis-
sue, extensive blood sinuses and developing teeth (Figs 2A–
C,E,F and 3A–E). In this study, several histological sections
of the oral, lingual and pharyngeal mucosa from all specimens
were carefully investigated for the presence of taste buds. The
taste buds were not observable.
The muscular tongue pad is flat and short in the floor of
the mouth (Fig. 4A). The anterior and lateral margins of the
tongue are free and continuous with the oral mucosa overlying
the lower jaw. The tongue is lined by stratified columnar
© 2016 The Royal Swedish Academy of Sciences
Muikham et al.  Digestive system of Supachai’s caecilian
epithelium, consisting of prismatic cells, goblet cells and basal
cells (Fig. 4B). Lingual glands are scatted throughout the lin-
gual mucosa (Fig. 4A,C–F). These glands contain mucous
cells and basal cells (Fig. 4B). Goblet cells and glandular
mucous cells are reactive to PAS, AB pH 2.5 and, to a lesser
extent, AB pH 1.0 (Fig. 4C–F; Table 2), suggesting that
these cells produce a mixture of neutral, carboxylated acid
and sulphated acid mucosubstances. PAS reactivity of the lin-
gual tissues treated with diastase is similar to that of the
untreated tissues (Table 2). In addition, melanocytes are pre-
sent in the lamina propria of the tongue (Fig. 4B–F). Embed-
ded in the lamina propria is an extrinsic musculature, the
genioglossus, which is the main structure of the tongue pad
(Fig. 4A). This muscle originates from the inner side of the
lower jaw and fans out to insert between adjacent lingual
glands, thus forming periodically disposed bundles of skeletal
muscle fibres, which are separated by collagenous connective
tissues with venous sinuses (Fig. 4A–C,E).
The terminology for the microscopic structures of devel-
oping and functional teeth of I. supachaii follows that previ-
ously described for Hypogeophis rostratus and Grandisonia
sechellensis (synonym of Grandisonia diminutiva; Lawson
1965a,b; Casey and Lawson 1981). Early tooth development
involves extension of the oral epithelium into the underlying
connective tissue to become a dental lamina (Fig. 5A). An
enamel organ, a tooth primordium, is formed as cellular thick-
ening at the base of the dental lamina. Cells in the enamel
organ are arranged into a bilaminar structure, consisting of
the inner and outer dental laminae (Fig. 5B). Subsequently,
the base of the enamel organ becomes concave in conjunction
with protrusion of the mesenchyme into the enamel organ to
become a dental papilla (Figs 2C and 5C). At this stage, the
enamel organ appears as a cup-like structure enclosing the
mesenchymal dental papilla (Figs 2C and 5C). The outer-
most mesenchymal cells of the dental papilla differentiate into
odontoblasts, which are organized into a single-cell layer
immediately subjacent to the inner dental lamina (Figs 2C
and 5C). Cells in the inner dental lamina become ameloblasts,
which produce an enamel (Fig. 5C). Deposited between the
odontoblastic and ameloblastic layers is a conical uncalcified
predentine, which is a collagenous matrix and positive to ani-
line blue upon Masson’s trichrome staining (Fig. 5C). The
predentine is further mineralized to become the dentine
(Fig. 5C). In the course of tooth development, a tooth germ
elongates bidirectionally towards the oral cavity and dentiger-
ous bones (Fig. 5D). The mineralized enamel is formed on
top of the dentine (Fig. 5C,F).
Functional teeth of I. supachaii are composed of two dis-
crete regions (crown and pedicel) that are separated by an
indented area, called the annular dividing zone (Fig. 5D–F).
The crown protrudes and curves posteriorly into the oral cav-
ity, while the pedicel is embedded in the oral floor and the
palate where it is ankylosed to dentigerous bones (Fig. 5D–
F). Dental substances contain the acellular dentine and
enamel (Fig. 5F). In the crown, the bilaminar dentine consists
3
Digestive system of Supachai’s caecilian  Muikham et al.
A B
C D
E F
of an inner circumpulpar layer, which is adjacent to the pulp
cavity, and an outer pallial layer, which is closely apposed to
the enamel (Fig. 5F). The circumpulpar layer is penetrated
by numerous dentinal tubules that lie perpendicularly to the
dental surface (Fig. 5F). These tubules are fewer in the pallial
layer than in the circumpulpar layer (Fig. 5F). The dentine of
the pedicel is similar to that of the crown except for the
absence of the dentinal tubules (Fig. 5F). The dividing zone
at the anterior aspect of the tooth consists of a single bundle of
fibrous tissue, while two fibrous bundles are observed at the
posterior aspect of each tooth (Fig. 5E,F). The pedicel is cov-
ered by a layer of cementocytes (cellular cementum; Fig. 5F).
The pulp cavity contains loose connective tissue, which is
overlain by an odontoblastic layer (Fig. 5F).
Pharynx. The pharyngeal region close to the tongue is lined
by stratified ciliated columnar epithelium, while the region
near the oesophagus is covered by pseudostratified ciliated
columnar epithelium (Fig. 6). The pharyngeal epithelium
possesses ciliated cells, goblet cells and basal cells (Fig. 6A).
Goblet cells secrete a mixture of neutral, carboxylated acid
and sulphated acid mucosubstances, all of which are liberated
into the pharyngeal cavity (Fig. 6C–F; Table 2). These
4 © 2016 The Royal Swedish Academy of Sciences
Acta Zoologica (Stockholm) 0: 1–19 (August 2016)
Fig. 2—Microanatomical structure of the oral
floor. —A. Oral mucosa showing two distinct
epithelia: stratified squamous epithelium
(SSE) and stratified cuboidal epithelium
(SCE) at the anterior region and the posterior
region of the oral floor, respectively. —B. The
posterior region of the oral mucosa showing
goblet cells (GC) and oral mucous glands
(OG) composed of mucous cells (MC) and
basal cells (BC). —C–F. Glycoconjugate his-
tochemistry showing neutral (C), carboxy-
lated acid (D) mixed (neutral and
carboxylated acid) (E) and sulphated acid (F)
mucosubstances in goblet cells and oral
mucous glands. Ab, ameloblasts; BV, blood
vessel; De, dentine; DP, dental papilla; DT,
developing teeth; LP, lamina propria; Ob,
odontoblasts. —A and B, H&E staining; C,
PAS-H staining; D, AB pH 2.5 staining; E,
PAS-AB pH 2.5 staining; F, AB pH 1.0
staining.
neutral mucosubstances are not sensitive to diastase treat-
ment, as determined by PAS staining (Table 2). The lamina
propria comprises vascularized loose connective tissue with
scattered melanocytes (Fig. 6). The muscularis houses longi-
tudinal bundles of skeletal muscle fibres (Fig. 6A–C). The
adventitia is not distinctive due to its continuation with dense
irregular connective tissue of the dermis.
Oesophagus. The oesophagus is constituted by three tissue lay-
ers from the inside to the outside: mucosa, muscularis and
adventitia (Fig. 7). The oesophageal mucosa consists of pseu-
dostratified ciliated columnar epithelium and lamina propria
that contains highly vascularized loose connective tissue
(Fig. 7). Three cell types are identified in the oesophageal
epithelium, namely ciliated cells, goblet cells and basal cells
(Fig. 7B). The oesophageal goblet cells exhibit similar histo-
chemical reactions to those in the oral and pharyngeal mucosa
(Fig. 7C,D; Table 2). Pretreatment of the oesophageal tis-
sues with diastase has no effects on PAS reactivity, compared
with the untreated tissues (Table 2). It is noted that numer-
ous blood vessels in the lamina propria reveal intimate associa-
tion with the oesophageal epithelium (Fig. 7B). There are no
oesophageal glands in the mucosa. The oesophageal lumen is
Acta Zoologica (Stockholm) 0: 1–19 (August 2016) Muikham et al.  Digestive system of Supachai’s caecilian

A B

C D

Fig. 3—Microscopic structure of the palatine

region. —A. Palatine mucosa showing strati-
fied squamous epithelium and stratified
cuboidal epithelium at the anterior and poste-
rior palatine regions, respectively. —B. Pala- E F
tine mucous glands (PG) at the posterior
palatine region. —C–F. Carbohydrate histo-
chemistry showing neutral (C), carboxylated
acid (D), mixed (neutral and carboxylated
acid) (E) and sulphated acid (F) mucosub-
stances. Bo, bone; L, glandular lumen; OE,
olfactory epithelium. —A and B, H&E stain-
ing; C, PAS-H staining; D, AB pH 2.5 stain-
ing; E, PAS-AB pH 2.5 staining; F, AB pH
1.0 staining.

corrugated as a result of the projection of several mucosal lon-
gitudinal folds (Fig. 7). Diffuse lymphatic tissues are found in
the lamina propria (Fig. 7C,D). The submucosa is not pre-
sent due to the absence of the muscularis mucosae. The mus-
cularis consists of two muscular tunics: the inner circular and
outer longitudinal smooth muscles (Fig. 7B). The outermost
adventitia contains thin layers of loose collagenous connective
tissue (Fig. 7A).
Stomach. The stomach has a larger calibre than any other
parts of the tubular digestive tract. It is histologically divided
into the anterior and posterior regions (Figs 8 and 9). The
gastric wall consists of four tissue layers: mucosa, submucosa,
muscularis and serosa. We report for the first time the caecil-
ian gastric rugae, which are formed by elevation of the mucosa
and submucosa into the gastric lumen (Figs 8A and 9A–C).
The posterior gastric region contains more prominent rugae
than the anterior gastric region (Figs 9A–C versus 8A). In
addition, the mucosal surface invaginates to become several
furrows, called gastric pits (Figs 8B–F and 9B–D). The gas-
tric mucosa is composed of simple columnar epithelium, lam-
ina propria and muscularis mucosae (Figs 8B,C and 9A,B).
The surface epithelium is formed by surface mucous cells,
while the gastric pits are lined by foveolar mucous cells
(Figs 8B–F and 9B–D). The surface and foveolar mucous
cells have the basal, oval nucleus and supranuclear mucinogen
granules, thus creating a mucous sheet of cells (Figs 8B–F
and 9B–D). Both cell types are intensely stained with PAS,
but weakly stained with AB pH 2.5 (Figs 8D–F and 9C–D;
Table 2). At the anterior region of the stomach, the lamina
propria possesses vascularized loose connective tissue and a
number of simple tubular gastric glands (Fig. 8B–D,F). Few
gastric glands open into a single gastric pit. These glands are
not present in the posterior region of the stomach (Fig. 9).
The gastric glands are divided into the upper neck region and
the lower region. Mucous neck cells are found at the neck por-
tion of the gastric glands (Fig. 8B–F). These cells are spheri-
cal with a basal, flat nucleus (Fig. 8B,C). The mucous neck
cells are reactive to both PAS and AB pH 2.5, but not reactive
to AB pH 1.0 (Fig. 8D–F; Table 2). There are no changes in
PAS reactivity of the gastric tissues following diastase treat-
ment, compared with the untreated tissues (Table 2). The
lower part of the gastric glands contains oxynticopeptic (chlo-
ridropeptic) cells that are characterized by their polymorphic

© 2016 The Royal Swedish Academy of Sciences 5

Digestive system of Supachai’s caecilian  Muikham et al. Acta Zoologica (Stockholm) 0: 1–19 (August 2016)

Table 2 Glycoconjugate histochemistry of the digestive tract in epithelium (Figs 10 and 11). Two main cell types constitute
Ichthyophis supachaii the intestinal epithelium: columnar absorptive enterocytes
and goblet cells (Figs 10B,F and 11). Absorptive enterocytes
Histochemical method
have basal, ellipsoid nuclei, and the apical striated border is
AB AB PAS-AB
reactive to PAS, but negative to AB staining (Figs 10D,F and
Digestive tract/cell PAS D-PAS pH 1.0 pH 2.5 pH 2.5 11). In the anterior intestinal region, goblet cells are stained
magenta with PAS and blue with AB pH 2.5, and costaining
Oral cavity (oral floor, palate and tongue) the intestinal tissues with PAS-AB pH 2.5 results in purplish
Goblet cells ++++ ++++ ++ +++ ++++ blue colouration (Fig. 10C–F; Table 2). However, these cells
Glandular mucous cells ++++ ++++ ++ +++ ++++ are not reactive to AB pH 1.0 (Table 2). The lamina propria
Pharynx
of the intestinal folds contains loose connective tissue with
Goblet cells ++++ ++++ ++ +++ ++++
Oesophagus
blood vessels. There are no lymphatic central lacteals in the
Goblet cells ++++ ++++ + +++ ++++ folds. The posterior intestinal region has more goblet cells
Anterior region of the stomach than the anterior intestinal region (Figs 11 versus 10). These
Surface mucous cells ++++ ++++ – ++ ++++ cells reveal histochemical reactions with PAS, AB pH 1.0 and
Foveolar mucous cells ++++ ++++ – ++ ++++ AB pH 2.5 (Fig. 11B–D; Table 2). Pretreatment of the
Mucous neck cells +++ +++ – ++ +++
intestinal tissues with diastase does not interfere with PAS
Posterior region of the stomach
reactivity (Table 2). The intestinal wall lacks the muscularis
Surface mucous cells ++++ ++++ – ++ ++++
Foveolar mucous cells ++++ ++++ – ++ ++++ mucosae and plicae circulares. The inner circular and outer
Anterior region of the intestine longitudinal smooth muscles constitute the intestinal muscu-
Goblet cells ++++ ++++ – +++ ++++ laris (Figs 10 and 11). The serosa consists of thin connective
Posterior region of the intestine tissue covered externally by the mesothelium (Figs 10A and
Goblet cells ++++ ++++ + +++ ++++ 11A,B,D).
–, no reaction; +, weak reaction; ++, moderate reaction; +++, strong reaction;
++++, very strong reaction. Microanatomy of accessory digestive organs
Liver. Almost the entire hepatic lobe is encapsulated by loose
shape, cytoplasmic eosinophilic granules and concentric connective tissue, which is further invested by the mesothe-
spherical nucleus (Fig. 8D–F). The oxynticopeptic cells are lium (Fig. 12A,E). Some areas of the hepatic lobes are in
negative to PAS and AB staining (Fig. 8D–F; Table 2). Nota- direct contact with the mesothelium (Fig. 12A). Embedded
bly, these cells are exclusively localized to the anterior gastric in the capsule is the perihepatic subcapsular haematopoietic
region (Figs 8 versus 9). The muscularis mucosae separate tissue that circumscribes the hepatic parenchyma (Fig. 12A,
the lamina propria from the submucosa (Figs 8B,C and 9A, C,E). The haematopoietic tissue houses numerous developing
B). The submucosa contains loose collagenous connective tis- blood cells. Mitotic activity of developing blood cells is fre-
sue interspersed with blood vessels (Figs 8B,C and 9A,B). quently found in the perihepatic subcapsular haematopoietic
The muscularis comprises an inner circular smooth muscle tissue. Mature blood cells leave the perihepatic subcapsular
and an outer longitudinal smooth muscle (Figs 8B–D,F and haematopoietic tissue, enter sinusoids and are further trans-
9B). The muscularis of the posterior gastric region is thicker ported to the central veins. The hepatic parenchyma consists
than that of the anterior gastric region (Figs 9A–C versus 8B– mainly of hepatocytes that are organized into anastomosing,
D,F). The serosa is thin and covered by the mesothelium two-cell-thick hepatic plates (Fig. 12B,E,F). Hepatocytes
(Figs 8D and 9B). have a polygonal shape and the spherical nucleus at the basal
domain (Fig. 12B,E,F). These cells exhibit polarity in the
Intestine. The transition between the stomach and the intes- way that the basal domain is in close contact with sinusoids,
tine is marked by abrupt histological changes from the gastric while the apical domain of opposing cells in a bilayer hepatic
mucosa with a mucous sheet of cells to the intestinal mucosa plate faces each other with bile canaliculi located in the middle
with numerous intestinal folds. There is no distinct muscular of the opposing cells (Fig. 12B,E,F). The hepatic parench-
sphincter at the transition area. The intestine is divided into yma is not organized into distinct hepatic lobules. The sinu-
the anterior and posterior intestinal regions, both having three soids are irregularly disposed and vary widely in size
tissue layers without submucosa (Figs 10 and 11). There are (Fig. 12A,B,D-F). Hepatic pigment cells are scattered in the
no submucosal (Brunner’s) glands and glandular crypts sinusoids (Fig. 12). These cells contribute to numerous black
(crypts of Lieberk€ uhn). The mucosa of the anterior intestinal spots on the liver surfaces (Fig. 1). The portal area consists of
region extends into the intestinal lumen and becomes several the hepatic vein and the bile duct (Fig. 12C). The bile duct is
intestinal folds (Fig. 10A–C,E,F). Interestingly, the posterior lined by simple cuboidal epithelium (Fig. 12C,D). Several
intestinal region is devoid of the intestinal folds (Fig. 11). The bile ducts drain into a hepatic duct, which is located at the
luminal surface of both regions is lined by columnar peripheral area of the hepatic lobes (Fig. 12D). The lining of

6 © 2016 The Royal Swedish Academy of Sciences

Acta Zoologica (Stockholm) 0: 1–19 (August 2016) Muikham et al.  Digestive system of Supachai’s caecilian

A B

C D

Fig. 4—Microanatomy of the tongue. —A.

Longitudinal section of Ichthyophis supachaii
head showing the muscular tongue pad (TP)
supported by the fan-shaped genioglossus
(Ge). —B. Lingual mucosa composed of
stratified columnar epithelium and lamina E F
propria with lingual glands (LG) and
genioglossus muscle. —C–F. Glycoconjugate
histochemistry showing neutral (C), carboxy-
lated acid (D), mixed (neutral and carboxy-
lated acid) (E) and sulphated acid (F)
mucosubstances. LE, lingual epithelium; Me,
melanocytes; PC, prismatic cells; S, skin. —A
and B, H&E staining; C, PAS-H staining; D,
AB pH 2.5 staining; E, PAS-AB pH 2.5 stain-
ing; F, AB pH 1.0 staining.

the hepatic duct is of simple cuboidal epithelium (Fig. 12D).
Pretreatment of the liver tissues with diastase results in a slight
reduction in PAS reactivity of the hepatocytes (Fig. 12F),
compared with the tissues without enzymatic treatment
(Fig. 12E), suggesting that these hepatocytes accumulate a
small amount of glycogens.
Gallbladder. The gallbladder wall is thin, compared to other
parts of the digestive tract. The lumen is lined by simple
cuboidal epithelium overlying the lamina propria (Fig. 13A).
The muscularis consists of smooth muscle cells (Fig. 13A).
The gallbladder is covered externally by the adventitia, which
is composed of thin layers of loose connective tissue
(Fig. 13A). The cystic duct proper is continuous with the gall-
bladder wall (Fig. 13A). This duct is lined by simple cuboidal
epithelium (Fig. 13A).
Pancreas. The bilobed pancreas consists of exocrine and
endocrine pancreatic tissues (Fig. 13B–D). Exocrine acinar
cells are organized into acini that are enclosed by connective
tissue (Fig. 13B–D). Several acini form a pancreatic lobule.
Pancreatic acinar cells have a pyramidal shape with the
spherical or ellipsoid nucleus situated at the basal region of
the cells (Fig. 13C–D). Numerous eosinophilic zymogen
granules are found in the cytoplasm of acinar cells (Fig. 13C).
The acinar lumen is occupied by centroacinar cells
(Fig. 13C). Pancreatic ducts are lined by simple cuboidal
epithelium and are found at the border between the pancre-
atic tissues and the intestine (Figs 10C and 13B). The apical
border of these ducts exhibits PAS reactivity (Fig. 10C).
Endocrine cells, also called islets of Langerhans, are smaller
than exocrine cells and scattered among the pancreatic par-
enchyma (Fig. 13D). Small blood vessels are found inside the
endocrine tissue.
Discussion
The digestive tract and liver of I. supachaii are elongated as is
the body, similar to other caecilians (e.g. Delsol et al. 1995;
Junqueira et al. 1999; Exbrayat and Estabel 2006). In the oral
cavity, the lingual surface of I. supachaii is non-keratinized
and moistened by mucus secreted from surface goblet cells
and lingual glands, thus preventing the tongue from being
dehydrated. The lingual glands of I. supachaii are distributed

© 2016 The Royal Swedish Academy of Sciences 7

Digestive system of Supachai’s caecilian  Muikham et al.
A B
C D
E F
throughout the lingual mucosa, as in other caecilians, such as
Ichthyophis glutinosus (Teipel 1932), Dermophis mexicanus
(Bemis et al. 1983), Siphonops annulatus (Junqueira et al.
1999) and Boulengerula boulengeri (Budzik et al. 2013), but
different from Hypogeophis rostratus in which the lingual glands
are localized to the median region of the lingual mucosa (Mar-
cus 1932). The presence of surface goblet cells and glandular
mucous cells in the tongue of I. supachaii is similar to several
caecilians, including I. glutinosus (Zylberberg 1972, 1977),
D. mexicanus (Bemis et al. 1983), I. kohtaoensis, Typhlonectes
compressicauda (Zylberberg 1986) and S. annulatus (Junqueira
et al. 1999). These cells reveal similar histochemical reactions
to carbohydrate moieties, as in I. glutinous, some urodeles
(family Salamandridae) and some anurans (family Alytidae,
Bufonidae, Hylidae and Ranidae; Zylberberg 1977). It is sug-
gested that the short lingual glands of caecilians are less devel-
oped than the long tubular lingual glands found in anurans
and urodeles (Zylberberg 1977).
The muscular tongue of I. supachaii consists of a single
type of the voluntary protractor muscle, the genioglossus, as
described in other caecilians, including H. rostratus (Teipel
1932), D. mexicanus (Bemis et al. 1983), T. compressicauda
8 © 2016 The Royal Swedish Academy of Sciences
Acta Zoologica (Stockholm) 0: 1–19 (August 2016)
Fig. 5—Histology of the oral apparatus show-
ing odontogenesis of Ichthyophis supachaii at
the oral floor (A, C and D) and the palate (B,
E and F). —A. Early odontogenesis showing
a dental lamina (DL) derived from ingrowth
of the oral epithelium. —B. Bilaminar struc-
ture of a tooth primordium comprising the
inner dental lamina (IDL) and the outer den-
tal lamina (ODL). —C. Formation of acellu-
lar dentine produced by odontoblasts and
acellular enamel (E) from ameloblasts. Miner-
alization of the tooth primordium to form
dentine. —D. More advanced odontogenesis
showing erupting tooth linked to the dentiger-
ous bone (DB). —E and F. Functional
erupted tooth showing the bipartite dental
structure: crown (Cr) and pedicel (Pe), both
divided by the dividing zone (DZ). CL, cir-
cumpulpar layer; Ce, cementocyte; T, denti-
nal tubules; P, pulp cavity; Pd, predentine;
PL, pallial layer. A, E and F, H&E staining;
B, PAS-H staining; C and D, Masson’s tri-
chrome staining.
(Hraoui-Bloquet and Exbrayat 1997) and S. annulatus (Jun-
queira et al. 1999). While the anuran tongue is extensible to
capture prey partly due to the activity of the genioglossus
(O’Reilly and Nishikawa 1995; Meyers et al. 2004), the cae-
cilian tongue is not protrusible, suggesting that the caecilian
tongue does not directly participate in predation (Bemis et al.
1983; Wake 1992). However, it was suggested that contrac-
tion of the caecilian genioglossus results in (i) constriction of
the basal region of the lingual glands, thus facilitating glandu-
lar secretion; and (ii) compression of the tongue pad, thus
enabling food swallowing (Bemis et al. 1983).
Teeth of I. supachaii are a bipartite structure consisting of
the crown and the pedicel, both of which are separated by the
annular dividing zone, as in other caecilians (Oltmanns 1952;
Parsons and Williams 1962; Wake 1976; Casey and Lawson
1981; Hraoui-Bloquet and Exbrayat 1996). The morphology
of the tooth crown in adult caecilians reveals interspecific vari-
ation (Taylor 1968; Wake 1978; Wake and Wurst 1979; Gre-
ven 1984, 1986; Wilkinson 1991). Although the dental
cuspedness was unable to be determined in the present study,
it appears that all species of the genus Ichthyophis possess
pointed, recurved and bicuspid teeth (Mark Wilkinson,
Acta Zoologica (Stockholm) 0: 1–19 (August 2016) Muikham et al.  Digestive system of Supachai’s caecilian

A B

C D

Fig. 6—Microscopic structure of the pharynx.

—A and B. Pharyngeal wall showing pseu-
dostratified ciliated columnar epithelium E F
(PCE), lamina propria and muscularis (M).
Note the presence of melanocytes in lamina
propria. —C-F. Carbohydrate histochemistry
showing neutral (C), carboxylated acid (D),
mixed (neutral and carboxylated acid) (E)
and sulphated acid (F) mucosubstances. CC,
ciliated cells. A, H&E staining; B, Masson’s
trichrome staining; C, PAS-H staining; D,
AB pH 2.5 staining; E, PAS-AB pH 2.5 stain-
ing; F, AB pH 1.0 staining.

A B

Fig. 7—Microanatomical structure of the

oesophagus. —A. Transverse sectional view
of the oesophagus showing several longitudi-
nal folds, formed by the mucosa. —B. Oeso-
phageal epithelium consisting of ciliated cells, C D
goblet cells and basal cells. Note the presence
of numerous subepithelial blood vessels. —C
and D. Carbohydrate histochemistry showing
neutral (C) and mixed (neutral and carboxy-
lated acid) (D) mucosubstances. Ad, adventi-
tia; CM, circular smooth muscle; LM,
longitudinal smooth muscle; LT, diffuse lym-
phatic tissue; Muc, mucus. —A and B, H&E
staining; C, PAS-H staining; D, PAS-AB pH
2.5 staining.

© 2016 The Royal Swedish Academy of Sciences 9

Digestive system of Supachai’s caecilian  Muikham et al.
A B
C D
E F
A B
C D
10
Acta Zoologica (Stockholm) 0: 1–19 (August 2016)
Fig. 8—Microscopic structure of the anterior
region of the stomach. —A. Cross-sectional
view of the anterior gastric region showing
rugae (R), formed by elevated mucosa and
submucosa. —B and C. The gastric proper
showing gastric pits (GP) and gastric glands,
the latter housing mucous neck cells (MNC)
and oxynticopeptic cells (OC). —D–F. Pres-
ence of neutral (D), carboxylated acid (E)
and mixed (neutral and carboxylated acid)
(F) mucosubstances. FMC, foveolar mucous
cells; MM, muscularis mucosae; Se, serosa;
SMC, surface mucous cells; Sub, submucosa.
—A. and B, H&E staining; C, Masson’s tri-
chrome staining; D, PAS-H staining; E, AB
pH 2.5 staining; F, PAS-AB pH 2.5 staining.
Fig. 9—Microanatomy of the posterior region
of the stomach. —A. Cross-sectional view of
the posterior gastric region showing promi-
nent rugae. —B. Gastric pits without gastric
glands. —C and D. Glycoconjugate histo-
chemistry showing neutral and carboxylated
acid mucosubstances, respectively. —A and
B, H&E staining; C, PAS-H staining; D, AB
pH 2.5 staining.
© 2016 The Royal Swedish Academy of Sciences
Acta Zoologica (Stockholm) 0: 1–19 (August 2016) Muikham et al.  Digestive system of Supachai’s caecilian

A B

C D

Fig. 10—Histology of the anterior region of

the intestine. —A. Transverse sectional view
of the anterior intestinal region showing
intestinal folds (IF). —B. Magnified image of
A, showing goblet cells and absorptive entero-
cytes (Ec) with apical striated border (SB). — E F
C. Carbohydrate histochemistry of the pan-
creato-intestinal junction showing neutral
mucosubstances in goblet cells and apical
domain of pancreatic ducts (PD). —D. Car-
boxylated acid mucosubstances in goblet cells.
—E and F. Coproduction of neutral and car-
boxylated acid mucosubstances in goblet cells.
Pa, pancreas. —A and B, H&E staining; C,
PAS-H staining; D, AB pH 2.5 staining; E
and F, PAS-AB pH 2.5.

personal communication). In addition to erupted functional
teeth, there exist various stages of developing teeth in mature
I. supachaii, as seen in other caecilians, including D. mexi-
canus, H. rostratus, G. larvata, G. multiplicata, G. seraphini
and T. compressicauda (Reuther 1931; Wake 1976; Casey and
Lawson 1981). Dental development and replacement in
amphibians are continually dynamic processes occurring
throughout the life (Gillette 1955; Kerr 1960; Wake 1980).
While the taste buds are present in the oropharyngeal
mucosa of several caecilians (Wake and Schwenk 1986;
Junqueira et al. 1999; Budzik et al. 2013, 2015), these struc-
tures were not detectable in the present study. However, it
remains unclear whether the taste buds are absent in the adult
I. supachaii. Alternatively, more sample size of the animals
may be required for investigation of these structures in adult
I. supachaii and more research on the presence and absence of
taste buds in caecilians would be useful.
The oesophagus of I. supachaii is lined by pseudostratified
ciliated columnar epithelium with different cell types, a typical
characteristic of the oesophagus found in three amphibian
orders (Norris 1959; Suganuma et al. 1981; Setoguti et al.
1987; Gallego-Huidobro et al. 1992; Delsol et al. 1995; Gal-
lego-Huidobro and Pastor 1996; Junqueira et al. 1999; Ferri
et al. 2001; Liquori et al. 2002, 2005, 2007; Exbrayat and
Estabel 2006; Machado-Santos et al. 2014). In addition, high
vascularization at the subepithelial region of I. supachaii
oesophagus may supply oxygen to the epithelial cells for their
cellular respiration, thus providing energy and driving cellular
activities, such as ciliary movement and exocytosis of the
mucus.
Pepsinogen and hydrochloric acid are two major gastric
juice components in vertebrates. In amphibians, three secre-
tion patterns for the two components have been categorized
based on the regional distribution of secretory cells. (i)
Pepsinogen is mainly produced by peptic cells in the oesopha-
geal glands, whereas hydrochloric acid is mainly secreted from
oxyntic cells in the gastric glands, as seen in the frogs Pelophy-
lax kl. esculentus (formerly Rana esculenta; Bani et al. 1992)
and Rana aurora aurora (Ferri et al. 2001). (ii) Pepsinogen is
produced by oesophageal peptic cells and oxynticopeptic cells
in the gastric glands; the latter also secrete hydrochloric acid,
such as in the frog Lithobates pipiens (formerly Rana pipiens;
Gallego-Huidobro et al. 1992; Gallego-Huidobro and Pastor
1996) and the toad Rhinella marina (formerly Bufo marinus;
Ruiz et al. 1993). (iii) Both pepsinogen and hydrochloric acid
are synthesized from the oxynticopeptic cells in the gastric

© 2016 The Royal Swedish Academy of Sciences 11

Digestive system of Supachai’s caecilian  Muikham et al.
glands due to the lack of the oesophageal glands, as observed
in anurans, including Bombina variegata (Bani et al. 1992)
and Bufo (Pseudepidalea) viridis (Liquori et al. 2002); urodele
Triturus carnifex (Liquori et al. 2005); and caecilians, such as
S. annulatus (Junqueira et al. 1999), M. unicolor (Exbrayat
and Estabel 2006) and likely I. supachaii in the present study.
However, further study is needed to determine coproduction
of pepsinogen and hydrochloric acid from the oxynticopeptic
cells in I. supachaii, for example using an immunohistochemi-
cal approach to detect pepsinogen and H,K-ATPase, an inte-
gral protein localized to the oxynticopeptic cells and
responsible for gastric acidification (Liquori et al. 2005).
Specifically, the oxynticopeptic cells are present exclusively in
the anterior region of the stomach of I. supachaii, suggesting
that proteolytic digestion for food materials initially occurs in
this gastric region and the process may continue in the poste-
rior region of the stomach.
There are mucosal and mucosal–submucosal projections
towards the lumen of the oesophagus, stomach and intestine,
thus creating the longitudinal folds, rugae and intestinal folds,
respectively. The gastric rugae have been shown in the stom-
ach of anurans, including L. pipiens (Gallego-Huidobro and
Pastor 1996) and Hyla orientalis (Akat et al. 2014), and are
reported for the first time in a caecilian here. It is possible that
upon relaxation of the circular musculature of the oesophagus
and stomach, the longitudinal folds and rugae become
unfolded, thus providing a potential space for the organs to
accommodate food passage, a similar suggestion by Budzik
et al. (2015) on the pharyngeal expansion in the caecilians
during food swallowing. While the oesophageal longitudinal
folds and gastric rugae do not seem to alter the total surface
areas of the oesophagus and the stomach, respectively, the
A B
C D
12
Acta Zoologica (Stockholm) 0: 1–19 (August 2016)
intestinal folds increase the intestinal surface areas for enzy-
matic digestion and nutrient absorption, like the mammalian
intestinal villi (Ross and Pawlina 2015). However, the intesti-
nal folds of I. supachaii are structurally different from the
avian and mammalian villi of which the lamina propria pos-
sesses the central lacteal and the base leads into the intestinal
tubular glands (crypts of Lieberk€ uhn; Andrew and Hickman
1974; Ishikawa et al. 1985; Chikilian and de Speroni 1996;
Reynolds and Rommel 1996; Zaher et al. 2012). The intesti-
nal folds of I. supachaii are present in the anterior intestinal
region and absent in the posterior intestinal region, while in
T. compressicauda, the folds become shorter in the posterior
region than in the anterior region (Martınez-Leones et al.
2004). The complete absence of the intestinal folds and a
decreased number of absorptive enterocytes in the posterior
intestinal region of I. supachaii suggest that this region has
reduced nutrient absorption and enzymatic digestion in com-
parison with the anterior intestinal region.
Mucosubstances are present throughout the digestive tract
of I. supachaii from the oral cavity to the intestine, as
described in this study, and in the cloaca, as reported in previ-
ous studies (Pewhom et al. 2015, 2016). The distribution pat-
terns of glycoconjugates along the digestive tract of
I. supachaii exhibit regional variation and possibly interspecific
variation, compared to other caecilians, including I. glutinosus
(Zylberberg 1977), Caecilia gracilis, H. rostratus, M. unicolor,
T. compressicauda (Exbrayat 1996, 2003), S. annulatus (Jun-
queira et al. 1999) and I. cf. kohtaoensis (Kuehnel et al.
2012); anurans, such as Duttaphrynus melanostictus (formerly
Bufo melanostictus; Loo and Wong 1975), Bufo viridis and
Rana graeca (Carmignani and Zaccone 1977), Alytes obstetri-
cians, Bufo bufo, Discoglossus pictus, H. arborea, P. ridibundus
Fig. 11—Histological structure of the poste-
rior region of the intestine. —A. Transverse
sectional view of the posterior intestinal region
showing intestinal proper. —B–D. Carbohy-
drate histochemistry showing neutral (B), car-
boxylated acid (C) and mixed (neutral and
carboxylated acid) (D) mucosubstances in
goblet cells. A, H&E staining; B, PAS-H
staining; C, AB pH 2.5 staining; D, PAS-AB
pH 2.5 staining.
© 2016 The Royal Swedish Academy of Sciences
Acta Zoologica (Stockholm) 0: 1–19 (August 2016) Muikham et al.  Digestive system of Supachai’s caecilian

A B

Fig. 12—Microanatomical structure of the

liver. —A. Organization of the liver showing
C D
hepatic parenchyma enveloped by perihepatic
subcapsular haematopoietic tissue (PSH).
Note the presence of pigment cells (Pi) in
sinusoids (Si) and transport of mature blood
cells from perihepatic subcapsular
haematopoietic tissue to sinusoids. —B.
Organization of hepatic parenchyma into two-
cell-thick hepatic plates, having sinusoids at
the basal domain and bile canaliculi (BC) at
the apical domain of opposing hepatocytes. —
C. Portal area showing the bile duct (BD)
and hepatic vein (HV). —D. The peripheral E F
zone of the liver showing the hepatic duct
(HD) lined by simple cuboidal epithelium. —
E and F. PAS histochemistry showing glyco-
gen granules (GG) accumulated in hepato-
cytes (E) as confirmed by a decreased PAS
reactivity following diastase treatment (F). *,
area of hepatic parenchyma without perihep-
atic subcapsular haematopoietic tissue; He,
hepatocytes. —A, C and D, H&E staining; B,
E and F, PAS-H staining.

(Zylberberg 1977), Hyla japonica, Pelophylax nigromaculatus
(formerly Rana nigromaculata), Xenopus laevis (Suganuma
et al. 1981), R. aurora aurora (Ferri et al. 2001), P. viridis and
Rana temporaria (Liquori et al. 2002) and Rhinella icterica
(Machado-Santos et al. 2014); and urodeles, including
Calotriton asper (formerly Euproctus asper), Lissotriton helveticus,
Pleurodeles waltii, Salamandra salamandra, Triturus cristatus,
T. marmoratus (Zylberberg 1977), Ambystoma mexicanum
(Suganuma et al. 1981) and T. carnifex (Liquori et al. 2007).
In general, these mucosubstances serve to lubricate the diges-
tive lumen, thus allowing smooth passage for food and pro-
tecting the mucosal surfaces from mechanical abrasion
(Gallego-Huidobro et al. 1992). A good example of this is an
increase in the mucous cell density in the posterior intestinal
region of I. supachaii compared with that of the anterior
region, suggesting that mucosubstances in the posterior
intestinal region aid transport and coat faecal materials for
their ejection, as in the anuran Hyla orientalis (Akat et al.
2014).
It is stated that the caecilian liver comprises 20–40 hepatic
lobes (Delsol et al. 1995). Intraspecific variation in the num-
ber of the hepatic lobes is observed in five mature I. supachaii,
ranging from 30 to 40 lobes. In addition, the hepatic lobe
number of I. supachaii also reveals interspecific variation,
compared to other caecilians, such as I. glutinosus (38 lobes),
U. narayani (as many as 40 lobes; Ahmad 1943) and S. annu-
latus (approximately 20 lobes; Junqueira et al. 1999). The
structural organization of the amphibian hepatic parenchyma
is categorized into three patterns: one-, two- and multilay-
ered-cell-thick hepatic plates (Akiyoshi and Inoue 2012). It is
suggested that the two-cell-thick and multilayered-cell-thick
plate types are of the primitive forms, while the one-cell-thick
plate type, as seen in the mammalian liver, is a more advanced
characteristic (Elias and Bengelsdorf 1952; Akiyoshi and
Inoue 2004). In the majority of anurans, the liver contains
only the one-cell-thick hepatic plates, but a combination of
the one- and two-cell-thick plates is observed in X. laevis,
X. tropicaris and Microhyla ornata (Akiyoshi and Inoue 2012).
In urodeles, the hepatic parenchymal arrangement exhibits
interspecific variation, in having the one-cell-thick plate type,
combined one- and two-cell-thick plate type and combined
two- and multilayered-cell-thick plate type (Akiyoshi and
Inoue 2012). The liver of I. supachaii is constituted by the
anastomosing, two-cell-thick hepatic plates, consisting of
polygonal hepatocytes, as described in other caecilians
(Exbrayat and Estabel 2006), but different from the aquatic

© 2016 The Royal Swedish Academy of Sciences 13

Digestive system of Supachai’s caecilian  Muikham et al.
A B
C D
Typhonectes sp., which has one-cell-thick hepatic plates
(Akiyoshi and Inoue 2012). In addition, the hepatocytes of
I. supachaii are the major epithelial cell type of the liver and
exhibit cell polarity by forming the apical and basal poles, as
observed in other vertebrates (Hua et al. 2012; M€ usch 2014).
This cell polarity is related to functional compartmentalization
of the hepatocytes in the way that bile is secreted into bile
canaliculi at the apical region, while bile acids, glucose, amino
acids and plasma proteins are transported between the hepa-
tocytes and sinusoids at the basal region of the hepatocytes
(Mescher 2013).
The liver of some amphibians, likely including I. supachaii,
also serves as a functional haematopoietic organ in adulthood
(Akiyoshi and Inoue 2012). However, some adult anurans are
devoid of the haematopoietic tissues in this organ (de Abreu
Manso et al. 2009; Akiyoshi and Inoue 2012). The presence
of haematopoietic cells in the perihepatic subcapsular
haematopoietic tissue of I. supachaii is similar to other caecil-
ians (Zapata et al. 1982; Delsol et al. 1995; Paillot et al.
1997a,b; Bleyzac et al. 2005; Akiyoshi and Inoue 2012);
urodeles (Jordan and Speidel 1924; Campbell 1969; High-
tower and St. Pierre 1971; Hightower and Haar 1975; Henry
and Charlemagne 1977; Xie et al. 2011; Akat and G€ ocßmen
2014; Lopez et al. 2014); and anurans (Spornitz 1975b;
Hadji-Azimi et al. 1987). However, I. supachaii lacks the
haematopoietic tissue at the periportal regions, unlike Typhlo-
nectes sp. in which haematopoiesis also takes place in these
regions (Akiyoshi and Inoue 2012). The liver of I. supachaii
also serves as a storage tissue for glycogen, similar to that of
the caecilian T. compressicauda (Hraoui-Bloquet and Exbrayat
1992) and of anurans and urodeles (Spornitz 1975a, 1978;
Delsol et al. 1995; Barni et al. 1999; Xie et al. 2011; Akat and
G€ocßmen 2014), while the digestive tract of I. supachaii does
14
Acta Zoologica (Stockholm) 0: 1–19 (August 2016)
Fig. 13—Microanatomy of the gallbladder
and pancreas. —A. Histological structure of
the gallbladder and cystic duct (CD) lined by
simple cuboidal epithelium. Inset showing the
gallbladder proper. —B. Pancreato-intestinal
junction showing pancreatic acini (Ac) and
pancreatic ducts. —C. Pancreatic acini show-
ing pyramidal acinar cells and centroacinar
cells (Ca) in acinar lumen. —D. Pancreatic
acini interspersed with islets of Langerhans
(IL). Ad, adventitia; CE, simple cuboidal
epithelium; I, intestine; ZG, zymogen gran-
ules. H&E staining.
not accumulate glycogen. In addition, hepatic pigment cells
are found in hepatic sinusoids of I. supachaii, as in other cae-
cilians including M. unicolor (Delsol et al. 1995) and T. com-
pressicauda (Exbrayat and Estabel 2006); anurans, such as
X. laevis (Spornitz 1975a; Zuasti et al. 1998), P. kl. esculentus
(Guida et al. 1998; Barni et al. 1999), Physalaemus nattereri
(synonym of Eupemphix nattereri; Franco-Belussi et al. 2012)
and Rhinella marina (Silva et al. 2013); and urodeles including
Ichthyosaura alpestris apuanus (formerly Triturus alpestris apua-
nus), T. carnifex (Barni et al. 1999), Necturus maculosus, Pro-
teus anguinus (Prelovsek and Bulog 2003; Prelovsek et al.
2008), Cynops orientalis (Xie et al. 2011), Lyciasalamandra ari-
kani (Akat and G€ ocßmen 2014) and Lissotriton italicus
(Bernab o et al. 2014). These hepatic pigment cells, aka mela-
nomacrophage centres, are also present in the liver of teleosts,
such as gilthead sea bream (Sparus aurata) and sea bass
(Dicentrarchus labrax; Meseguer et al. 1994) and Prochilodus
argenteus (Ribeiro et al. 2011) and the mud turtle (Kinosternon
flavescens; Christiansen et al. 1996). The presence of the hep-
atic pigment cells results in black speckled pigmentation on
the liver surface upon gross examination, similar to the anu-
rans Dendropsophus nanus, Physalaemus cuvieri and Rhinella
schneideri (Moresco and de Oliveira 2009) and urodeles C. ori-
entalis (Xie et al. 2011) and L. italicus (Bernabo et al. 2014).
Accumulated evidence has indicated that these extracuta-
neous pigment cells arise from Kupffer cells that belong to a
mononuclear phagocytic system of the mesodermal origin
(Guida et al. 1998; Corsaro et al. 2000; Gallone et al. 2002;
Sichel et al. 2002). Therefore, the origin of pigment cells is
different from that of integumentary melanocytes; the latter
are derived from an ectodermal neural crest cell lineage (Gal-
lone et al. 2007). Nonetheless, the hepatic pigment cells in
amphibians, such as melanocytes, are capable of synthesizing
© 2016 The Royal Swedish Academy of Sciences
Acta Zoologica (Stockholm) 0: 1–19 (August 2016)
Fig. 14—Schematic microscopic organization
of the tubular digestive tract in Ichthyophis
supachaii.
melanins, which are stored in the melanosomes (Scalia et al.
1988; Sichel 1988; Sichel et al. 1997; Guida et al. 2000; Pur-
rello et al. 2001; Prelovsek and Bulog 2003; Gallone et al.
2007). These pigment cells in the amphibian liver play protec-
tive roles in defying cytotoxic agents that mediate stress condi-
tions (Geremia et al. 1984; Schiel et al. 1987; Scalia et al.
1990; Fenoglio et al. 2005).
In conclusion, this article describes the microanatomy and
histochemical characteristics of the digestive system of I. su-
pachaii. Major tissue proper of the digestive system of I. su-
pachaii is similar to that of other vertebrates and summarized
in Fig. 14. Similar to other caecilians, odontogenesis is a con-
tinually occurring process in adult I. supachaii. Mucosub-
stances are present along the digestive tract and are associated
with digestion and protection of the underlying tissues from
harm. The liver not only functions as an accessory digestive
organ, but acts as a haematopoietic organ in adult I. supachaii
as well. The present study contributes to a better understand-
ing of the microanatomy of the caecilian digestive system, and
we believe it will stimulate similar research on the microscopic
structures and functions of organs in other caecilian taxa such
that the evolution and functions of their organs are also better
understood.
Acknowledgements
The authors thank Mr Suwit Punnadee, Mr Phamon
Sumpanthamitr and Mr Sayun Makong for their assistance
during specimen collection. The authors also gratefully
acknowledge Mr Watcharapong Hongjamrassil and Mrs
Naiyana Trinestsampan for assistance in the manuscript
preparation. This work was financially supported by the
Human Resource Development in Science Project (Science
Achievement Scholarship of Thailand, SAST) and the
Department of Zoology, Faculty of Science, Kasetsart Univer-
sity, Bangkok, Thailand.
© 2016 The Royal Swedish Academy of Sciences
Muikham et al.  Digestive system of Supachai’s caecilian
References
de Abreu Manso, P. P., de Brito-Gitirana, L. and Pelajo-Machado,
M. 2009. Localization of hematopoietic cells in the bullfrog (Litho-
bates catesbeianus). – Cell and Tissue Research 337: 301–312.
Ahmad, B. A. G. 1943. Histology and cytology of the liver of Ichthyo-
phis glutinosus (Linn.) and Uraeotyphlus narayani (Seshachar)-Part I.
– Journal of the Mysore University 4: 41–55.
Akat, E. and G€ ocßmen, B. 2014. A histological study on hepatic struc-
ture of Lyciasalamandra arikani (Urodela: Salamandridae). – Rus-
sian Journal of Herpetology 21: 201–204.
Akat, E., Arıkan, H. and G€ ocßmen, B. 2014. Histochemical and bio-
metric study of the gastrointestinal system of Hyla orientalis (Bed-
riaga, 1890) (Anura, Hylidae). – European Journal of Histochemistry
58: 291–295.
Akiyoshi, H. and Inoue, A. 2004. Comparative histological study of
teleost livers in relation to phylogeny. – Zoological Science 21:
841–850.
Akiyoshi, H. and Inoue, A. M. 2012.Comparative histological study
of hepatic architecture in the three orders amphibian livers. – Com-
parative Hepatology 11: 2.
Andrew, W. and Hickman, C. P. 1974. Histology of the Vertebrates,
pp. 297–315. C. V. Mosby, Saint Louis.
Bancroft, J. D., Gamble, M. 2008. Theory and Practice of Histologi-
cal Techniques, pp. 83–186. Churchill Livingstone Elsevier,
Philadelphia.
Bani, G., Formigli, L. and Cecchi, R. 1992. Morphological observa-
tions on the glands of the oesophagus and stomach of adult Rana
esculenta and Bombina variegata. – Italian Journal of Anatomy and
Embryology 97: 75–87.
Barni, S., Bertone, V., Croce, A. C., Bottiroli, G., Bernini, F. and
Gerzeli, G. 1999. Increase in liver pigmentation during
natural hibernation in some amphibians. – Journal of Anatomy 195:
19–25.
Bemis, W. E., Schwenk, K. and Wake, M. H. 1983. Morphology and
function of the feeding apparatus in Dermophis mexicanus (Amphi-
bia: Gymnophiona). – Zoological Journal of the Linnean Society 77:
75–96.
Bernab o, I., Biasone, P., Macirella, R., Tripepi, S. and Brunelli, E.
2014. Liver histology and ultrastructure of the Italian newt (Lissotri-
ton italicus): Normal structure and modifications after acute
15
Digestive system of Supachai’s caecilian  Muikham et al.
exposure to nonylphenol ethoxylates. – Experimental and Toxicologic
Pathology 66: 455–468.
Bleyzac, P., Cordier, G. and Exbrayat, J.-M. 2005. Morphological
description of embryonic development of immune organs in Typhlo-
nectes compressicauda (Amphibia, Gymnophiona). – Journal of Her-
petology 39: 57–65.
_
Budzik, K. A., Zuwała, K. and Measey, G. J. 2013. The occurrence
of taste buds in adults of the terrestrial caecilian Boulengerula boulen-
geri Tornier, 1898 (Lissamphibia: Gymnophiona: Herpelidae). –
African Zoology 48: 407–411.
_
Budzik, K. A., Zuwała, K., Kupfer, A., Gower, D. J. and Wilkinson,
M. 2015. Diverse anatomy of the tongue and taste organs in five
species of caecilian (Amphibia: Gymnophiona). – Zoologischer
Anzeiger 257: 103–109.
Campbell, F. R. 1969. Electron microscopic studies on granulocy-
topoiesis in the slender salamander. – The Anatomical Record 163:
427–441.
Carmignani, M. P. A. and Zaccone, G. 1977. Histochemical studies
on the tongue of anuran amphibians-II. Comparative morpho-
chemical study of the taste buds and the lingual glands in Bufo viri-
dis Laurenti and Rana graeca Boulenger with particular reference
to the mucosaccharide histochemistry. – Cellular and Molecular Biol-
ogy 22: 203–217.
Casey, J. and Lawson, R. 1981. A histological and scanning electron
microscope study of the teeth of caecilian amphibians. – Archives of
Oral Biology 26: 49–58.
Chatterjee, B. K. 1936. The anatomy of Uraeotyphlus menoni Annan-
dale. Part I. The digestive, circulatory, respiratory, and urino-geni-
tal systems. – Anatomischer Anzeiger 81: 393–414.
Chikilian, M. and de Speroni, N. B. 1996. Comparative study of the
digestive system of three species of tinamou. I. Crypturellus
tataupa, Nothoprocta cinerascens, and Nothura maculosa (Aves:
Tinamidae). – Journal of Morphology 228: 77–88.
Christiansen, J. L., Grzybowski, J. M. and Kodama, R. M. 1996.
Melanomacrophage aggregations and their age relationships in the
yellow mud turtle, Kinosternon flavescens (Kinosternidae). – Pigment
Cell Research 9: 185–190.
Corsaro, C., Scalia, M., Leotta, N., Mondio, F. and Sichel, G. 2000.
Characterisation of Kupffer cells in some Amphibia. – Journal of
Anatomy 196: 249–261.
Delsol, M., Flatin, J. and Exbrayat, J.-M. 1995. Le tube digestif des
Amphibiens adultes. In: Grasse, P. P. and Delsol, M. (Eds): Traite
de Zoologie, Tome XIV, Fasc.I A., pp. 497–508. Masson, Paris.
van Dijk, P. P., Chuaynkern, Y., Kupfer, A. and M€ uller, H. 2004.
Ichthyophis supachaii. IUCN Red List of Threatened Species. Ver-
sion 2004: e.T59636A11974248, http://www.iucnredlist.org.
Elias, H. and Bengelsdorf, H. 1952. The structure of the liver of verte-
brates. – Acta Anatomica 14: 297–337.
Exbrayat, J.-M. 1996. Croissance et cycle du cloaque chez Typhlo-
nectes compressicaudus (Dumeril et Bibron, 1841), Amphibien Gym-
nophione. – Bulletin de la Soci
et
e Zoologique de France 64: 37–50.
Exbrayat, J.-M. 2003. Apport de l’histologie a l’etude de quelques
aspects endocrines de la reproduction d’animaux rares issus de col-
lections. – Revue Francßaise D’Histotechnologie 16: 19–29.
Exbrayat, J.-M. and Estabel, J. 2006. Anatomy with particular refer-
ence to the reproductive system. In: Jamieson, B. G. M. (Ed.):
Reproductive Biology and Phylogeny of Gymnophiona (Caecil-
ians), vol. 5, pp. 79–155. Science Publishers, New Hampshire.
Fenoglio, C., Boncompagni, E., Fasola, M., Gandini, C., Comizzoli,
S., Milanesi, G. and Barni, S. 2005. Effects of environmental pollu-
tion on the liver parenchymal cells and Kupffer-melanomacropha-
gic cells of the frog Rana esculenta. – Ecotoxicology and Environmental
Safety 60: 259–268.
16
Acta Zoologica (Stockholm) 0: 1–19 (August 2016)
Ferri, D., Liquori, G. E., Natale, L., Santarelli, G. and Scillitani, G.
2001. Mucin histochemistry of the digestive tract of the red-legged
frog Rana aurora aurora. – Acta Histochemica 103: 225–237.
Franco-Belussi, L., de Souza Santos, L. R., Zieri, R., Vicentini, C. A.,
Taboga, S. R. and de Oliveira, C. 2012. Liver anatomy, histochem-
istry, and ultrastructure of Eupemphix nattereri (Anura: Leiuperidae)
during the breeding season. – Zoological Science 29: 844–848.
Gallego-Huidobro, J. and Pastor, L. M. 1996. Histology of the
mucosa of the oesophagogastric junction and the stomach in adult
Rana perezi. – Journal of Anatomy 188: 439–444.
Gallego-Huidobro, J., Pastor, L. M. and Calvo, A. 1992. Histology of
the esophagus of the adult frog Rana perezi (Anura: Ranidae). –
Journal of Morphology 212: 191–200.
Gallone, A., Guida, G., Maida, I. and Cicero, R. 2002. Spleen and
liver pigmented macrophages of Rana esculenta L. A new melano-
genic system?. – Pigment Cell Research 15: 32–40.
Gallone, A., Sagliano, A., Guida, G., Ito, S., Wakamatsu, K., Capozzi,
V., Perna, G., Zanna, P. and Cicero, R. 2007. The melanogenic sys-
tem of the liver pigmented macrophages of Rana esculenta L.-tyrosi-
nase activity. – Histology and Histopathology 22: 1065–1075.
Geremia, E., Corsaro, C., Bonomo, R., Giardinelli, R., Pappalardo,
P., Vanella, A. and Sichel, G. 1984. Eumelanins as free radicals
trap and superoxide dismutase activities in Amphibia. – Compara-
tive Biochemistry and Physiology Part B: Comparative Biochemistry 79:
67–69.
Gillette, R. 1955. The dynamics of continuous succession of teeth in
the frog (Rana pipiens). – American Journal of Anatomy 96: 1–36.
Greven, H. 1984. The dentition of Gegenophis ramaswamii Taylor,
1964 (Amphibia, Gymnophiona), with comments on monocuspid
teeth in the Amphibia. – Zeitschrift f€ ur Zoologie Systematisch Evolution
Forschungen 22: 342–348.
Greven, H. 1986. On the diversity of tooth crowns in Gymnophiona.
– M
emoires de la Soci
et
e Zoologique de France 43: 85–86.
Guida, G., Maida, I., Gallone, A., Boffoli, D. and Cicero, R. 1998.
Ultrastructural and functional study of the liver pigment cells from
Rana esculenta L. – In Vitro Cellular & Developmental Biology–Animal
34: 393–400.
Guida, G., Gallone, A., Maida, I., Boffoli, D. and Cicero, R. 2000.
Tyrosinase gene expression in the Kupffer cells of Rana esculenta L.
– Pigment Cell Research 13: 431–435.
Hadji-Azimi, I., Coosemans, V. and Canicatti, C. 1987. Atlas of adult
Xenopus laevis laevis hematology. – Developmental and Comparative
Immunology 11: 807–874.
Henry, M. and Charlemagne, J. 1977. Plasmocytic series in the peri-
hepatic layer of the urodele amphibian Pleurodeles waltlii Michah
(Salamandridae). – Developmental and Comparative Immunology 1:
23–32.
Hightower, J. A. and Haar, J. L. 1975. A light and electron micro-
scopic study of myelopoietic cells in the perihepatic subcapsular
region of the liver in the adult aquatic newt, Notophthalmus viri-
descens. – Cell and Tissue Research 159: 63–71.
Hightower, J. A. and St. Pierre, R. L. 1971. Hemopoietic tissue in the
adult newt, Notopthalmus viridescens. – Journal of Morphology 135:
299–307.
Hraoui-Bloquet, S. and Exbrayat, J.-M. 1992. Developpement
embryonnaire du tube digestif chez Typhlonectes compressicaudus
(Dumeril et Bibron, 1841), Amphibien Gymnophione vivipare.
– Annales de Sciences Naturelles, Zoologie, Paris, 13
eme S
erie 13:
11–23.
Hraoui-Bloquet, S. and Exbrayat, J.-M. 1996. Les dents de Typhlo-
nectes compressicaudus (Amphibia, Gymnophiona) au cours du
developpement. – Annales de Sciences Naturelles, Zoologie, Paris,
13
eme S
erie 17: 11–23.
© 2016 The Royal Swedish Academy of Sciences
Acta Zoologica (Stockholm) 0: 1–19 (August 2016)
Hraoui-Bloquet, S. and Exbrayat, J.-M. 1997. Developpement de la
langue de Typhlonectes compressicaudus (Dumeril et Bibron, 141),
Amphibien Gymnophione vivipare. – Bulletin de la Soci
et
e
Herp
etologique de France 82–83: 39–46.
Hua, M., Zhang, W., Li, W., Li, X., Liu, B., Lu, X. and Zhang, H.
2012. Molecular mechanisms regulating the establishment of hepa-
tocyte polarity during human hepatic progenitor cell differentiation
into a functional hepatocyte-like phenotype. – Journal of Cell Science
125: 5800–5810.
Ishikawa, K., Matoba, M., Tanaka, H. and Ono, K. 1985. Anatomi-
cal study of the intestine of the insect-feeder bat, Myotis frater
kaguae. – Journal of Anatomy 142: 141–150.
Jordan, H. E. and Speidel, C. C. 1924. Studies on lymphocytes. III.
Granulocytopoiesis in the salamander, with special reference to the
monphyletic theory of blood-cell origin. – American Journal of Anat-
omy 33: 485–505.
Junqueira, L. C. U., Jared, C. and Antoniazzi, M. M. 1999.
Structure of the caecilian Siphonops annulatus (Amphibia, Gym-
nophiona): general aspect of the body, disposition of the organs
and structure of the mouth, oesophagus and stomach. – Acta
Zoologica 80: 75–84.
Kamei, R. G., San Mauro, D., Gower, D. J., Van Bocxlaer, I., Sher-
ratt, E., Thomas, A., Babu, S., Bossuyt, F., Wilkinson, M. and
Biju, S. D. 2012. Discovery of a new family of amphibians from
northeast India with ancient links to Africa. – Proceedings of the Royal
Society B 279: 2396–2401.
Kerr, T. 1960. Development and structure of some actinopterygian
and urodele teeth. – Proceedings of the Zoological Society of London
133: 401–422.
Kuehnel, S., Herzen, J., Kleinteich, T., Beckmann, F. and Kupfer, A.
2012. The female cloaca of an oviparous caecilian amphibian
(Gymnophiona): Functional and seasonal aspects. – Acta Zoologica
93: 208–221.
Kupfer, A. and M€ uller, H. 2004. On the taxonomy of ichthyophiid
caecilians from southern Thailand: A reevaluation of the holotype
of Ichthyophis supachaii Taylor 1960 (Amphibia: Gymnophiona:
Ichthyophiidae). – Amphibia-Reptilia 25: 87–97.
Lawson, R. 1965a. The development and replacement of teeth in
Hypogeophis rostratus (Amphibia, Apoda). – Proceedings of the Zoolog-
ical Society of London 147: 352–362.
Lawson, R. 1965b. The teeth of Hypogeophis rostratus (Amphibia,
Apoda) and tooth structure in the Amphibia. – Proceedings of the
Zoological Society of London 145: 321–325.
Liquori, G. E., Scillitani, G., Mastrodonato, M. and Ferri, D. 2002.
Histochemical investigations on the secretory cells in the oesopha-
gogastric tract of the Eurasian green toad, Bufo viridis. – The Histo-
chemical Journal 34: 517–524.
Liquori, G. E., Zizza, S., Mastrodonato, M., Scillitani, G., Calamita,
G. and Ferri, D. 2005. Pepsinogen and H,K-ATPase mediate acid
secretion in gastric glands of Triturus carnifex (Amphibia, Caudata).
– Acta Histochemica 107: 133–141.
Liquori, G. E., Mastrodonato, M., Zizza, S. and Ferri, D. 2007. Gly-
coconjugate histochemistry of the digestive tract of Triturus carnifex
(Amphibia, Caudata). – Journal of Molecular Histology 38: 191–199.
Loo, S. K. and Wong, W. C. 1975. Histochemical observations on
the mucins of the gastrointestinal tract in the toad (Bufo melanostic-
tus). – Acta Anatomica 91: 97–103.
Lopez, D., Lin, L., Monaghan, J. R., Cogle, C. R., Bova, F. J.,
Maden, M. and Scott, E. W. 2014. Mapping hematopoiesis in a
fully regenerative vertebrate: The axolotl. – Blood 124: 1232–1241.
Luna, L. G. 1968. Manual of Histologic Staining Methods of the
Armed Forces Institute of Pathology, pp. 153–173. McGraw-Hill,
New York.
© 2016 The Royal Swedish Academy of Sciences
Muikham et al.  Digestive system of Supachai’s caecilian
Machado-Santos, C., Pelli-Martins, A. A., Abidu-Figueiredo, M. and
de Brito-Gitirana, L. 2014. Histochemical and immunohistochemi-
cal analysis of the stomach of Rhinella icterica (Anura, Bufonidae). –
Journal of Histology. Article ID 872795, doi:10.1155/2014/872795.
Marcus, E. 1932. Weitere Versuche und Beobachtungen u € ber die
Vorderdarmentwicklung bei den Amphibien. – Zoologische Jahr-
buch, Anatomie 55: 581–602.
Martınez-Leones, T., Viloria-Narvaez, M., Camacho-Bracho, J.,
Godoy-Brice~ no, R., Herrera-Marquez, E. and Mu~ noz-Gotera, R.
2004. Aspectos histol ogicos del intestino de Typhlonectes venezuelen-
sis (Amphibia: Gymnophiona, Typhlonectidae). – Revista Cient
ıfica
14: 237–243.
Mescher, A. L. 2013. Junqueira’s Basic Histology: Text and Atlas,
pp. 18–54. McGraw-Hill, New York.
Meseguer, J., L opez-Ruiz, A. and Esteban, M. A. 1994. Melano-
macrophages of the seawater teleosts, sea bass (Dicentrarchus labrax)
and gilthead seabream (Sparus aurata): Morphology, formation and
possible function. – Cell and Tissue Research 277: 1–10.
Meyers, J. J., O’Reilly, J. C., Monroy, J. A. and Nishikawa, K. C.
2004. Mechanism of tongue protraction in microhylid frogs. – Jour-
nal of Experimental Biology 207: 21–31.
Moresco, R. M. and de Oliveira, C. 2009. A comparative study of the
extracutaneous pigmentary system in three anuran amphibian spe-
cies evaluated during the breeding season. – South American Journal
of Herpetology 4: 1–8.
M€usch, A. 2014. The unique polarity phenotype of hepatocytes. –
Experimental Cell Research 328: 276–283.
Norris, J. L. 1959. The normal histology of the esophageal and gastric
mucosae of the frog, Rana pipiens. – Journal of Experimental Zoology
141: 155–173.
Oltmanns, E. 1952. Zur Morphologie der Z€ahne rezenter Amphibien.
– Anatomischer Anzeiger 24: 369–389.
O’Reilly, S. R. and Nishikawa, K. C. 1995. Mechanism of tongue
protraction during prey capture in the spadefoot toad Spea multipli-
cata (Anura: Pelobatidae). – Journal of Experimental Zoology 273:
282–296.
Paillot, R., Estabel, J. and Exbrayat, J.-M. 1997a. Organes
hematopo€ıetiques et cellules sanguines chez Typhlonectes compressi-
caudus et Typhlonectes natans (Amphibien, Gymnophione). – Bul-
letin Mensuel de la Soci
et
e Linn
eenne de Lyon 66: 124–134.
Paillot, R., Estabel, J., Keller, E. and Exbrayat, J.-M. 1997b. Pre-
mieres observations sur le foie hematopoietique chez les Gymno-
phiones. – Bulletin de la Soci
et
e Herp
etologique de France 82–83:
29–40.
Parsons, T. S. and Williams, E. E. 1962. The teeth of amphibia and
their relation to amphibian phylogeny. – Journal of Morphology 110:
375–389.
Pewhom, A., Chumnanpuen, P., Muikham, I., Chatchavalvanich, K.
and Srakaew, N. 2015. Microscopic structures of the ovary and
female genital ducts of Supachai’s caecilian, Ichthyophis supachaii
Taylor, 1960 (Amphibia: Gymnophiona). – Acta Zoologica. doi:
10.1111/azo.12139. (Early view).
Pewhom, A., Chumnanpuen, P., Muikham, I., Chatchavalvanich, K.
and Srakaew, N. 2016. Histomorphological studies of the testis and
male genital ducts of Supachai’s caecilian, Ichthyophis supachaii Tay-
lor, 1960 (Amphibia: Gymnophiona). – Acta Zoologica 97: 76–89.
Prelovsek, P. M. and Bulog, B. 2003. Biogenesis of melanosomes in
Kupffer cells of Proteus anguinus (Urodela, Amphibia). – Pigment
Cell Research 16: 345–350.
Prelovsek, P.-M., Mali, L. B. and Bulog, B. 2008. Hepatic pig-
ment cells of Proteidae (Amphibia, Urodela): A comparative
histochemical and ultrastructural study. – Animal Biology 58:
245–256.
17
Digestive system of Supachai’s caecilian  Muikham et al.
Purrello, M., Scalia, M., Corsaro, C., Di Pietro, C., Piro, S. and
Sichel, G. 2001. Melanosynthesis, differentiation, and apoptosis in
Kupffer cells from Rana esculenta. – Pigment Cell Research 14:
126–131.
Reuther, G. 1931. Die Zahnleiste von Hypogeophis: Beitrag zur Ken-
ntnis der Gymnophionen. XIV. – Morphologisches Jahrbuch 68:
105–112.
Reynolds, J. E. III and Rommel, S. A. 1996. Structure and
function of the gastrointestinal tract of the Florida manatee,
Trichechus manatus latirostris. – The Anatomical Record 245: 539–
558.
Ribeiro, H. J., Proc opio, M. S., Gomes, J. M. M., Vieira, F. O.,
Russo, R. C., Balzuweit, K., Chiarini-Garcia, H., Santana Castro,
A. C., Rizzo, E. and Corr^ea., J. D. Jr 2011. Functional dissimilarity
of melanomacrophage centres in the liver and spleen from females
of the teleost fish Prochilodus argenteus. – Cell and Tissue Research
346: 417–425.
Ross, M. H. and Pawlina, W. 2015. Histology: A Text and Atlas With
Correlated Cell and Molecular Biology, pp. 568–625. Lippincott
Williams & Wilkins, Philadelphia.
Ruiz, M.-C., Acosta, A., Abad, M. J. and Michelangeli, F. 1993.
Nonparallel secretion of pepsinogen and acid by gastric oxyn-
topeptic cells of the toad (Bufo marinus). – American Journal of
Physiology – Gastrointestinal and Liver Physiology 265: G934–
G941.
San Mauro, D., Gower, D. J., M€ uller, H., Loader, S. P., Zardoya, R.,
Nussbaum, R. A. and Wilkinson, M. 2014. Life-history evolution
and mitogenomic phylogeny of caecilian amphibians. – Molecular
Phylogenetics and Evolution 73: 177–189.
Sarasin, P. and Sarasin, F. 1887–1890. Ergebnisse Naturwis-
senschaftlicher Forschungen auf Ceylon in den Jahren 1884–1886
Band II: Zur Entwicklungsgeschichte und Anatomie der Ceylone-
sischen Blindw€ uhle Ichthyophis Glutinosus, pp. 1–263. C. W. Krie-
del’s Verlag, Wiesbaden.
Scalia, M., Geremia, E., Corsaro, C., Santoro, C., Sciuto, S. and
Sichel, G. 1988. The extracutaneous pigmentary system: Evidence
for the melanosynthesis in Amphibia and Reptilia liver. – Compara-
tive Biochemistry and Physiology Part B: Comparative Biochemistry 89:
715–717.
Scalia, M., Geremia, E., Corsaro, C., Santoro, C., Baratta, D. and
Sichel, G. 1990. Lipid peroxidation in pigmented and unpigmented
liver tissues: Protective role of melanin. – Pigment Cell Research 3:
115–119.
Schiel, G., Corsaro, C., Scalia, M., Sciuto, S. and Geremia, E. 1987.
Relationship between melanin content and superoxide dismutase
(SOD) activity in the liver of various species of animals. – Cell Bio-
chemistry and Function 5: 123–128.
Setoguti, T., Matsumura, H. and Chen, H.-S. 1987. Correlated his-
tochemical and electron microscopic studies of the esophageal
epithelium in the salamander, Hynobius nebulosus. – Archivum Histo-
logicum Japonicum 50: 283–297.
Sichel, G. 1988. Biosynthesis and function of melanins in hepatic pig-
mentary system. – Pigment Cell Research 1: 250–258.
Sichel, G., Scalia, M., Mondio, F. and Corsaro, C. 1997. The
amphibian Kupffer cells build and demolish melanosomes: An
ultrastructural point of view. – Pigment Cell Research 10:
271–287.
Sichel, G., Scalia, M. and Corsaro, C. 2002. Amphibia Kupffer cells.
– Microscopy Research and Technique 57: 477–490.
Silva, J. P. E., da Silva, D. C. B., Melo, F. T. V., Giese, E. G., Fur-
tado, A. P. and Santos, J. N. 2013. Liver histopathology in the cane
18
Acta Zoologica (Stockholm) 0: 1–19 (August 2016)
toad, Rhinella marina (Amphibia: Bufonidae), induced by Ortleppas-
caris sp. larvae (Nematoda: Ascarididae). – Journal of Parasitology
99: 250–256.
Spornitz, U. M. 1975a. Studies on the liver of Xenopus laevis. I. The
ultrastructure of the parenchymal cell. – Anatomy and Embryology
146: 245–264.
Spornitz, U. M. 1975b. Studies on the liver of Xenopus laevis. II. The
ultrastructure of the peritoneal cover and the perihepatic layer. –
Anatomy and Embryology 146: 265–277.
Spornitz, U. M. 1978. Studies on the liver of Xenopus laevis. III. The
ultrastructure and the glycogen content of the developing liver. –
Anatomy and Embryology 154: 1–25.
Suganuma, T., Katsuyama, T., Tsukahara, M., Tatematsu, M.,
Sakakura, Y. and Murata, F. 1981. Comparative histochemical
study of alimentary tracts with special reference to the mucous neck
cells of the stomach. – The American Journal of Anatomy 161: 219–
238.
Summers, A. P. and O’Reilly, J. C. 1997. A comparative study of
locomotion in the caecilians Dermophis mexicanus and Typhlonectes
natans (Amphibia: Gymnophiona). – Zoological Journal of the Lin-
nean Society 121: 65–76.
Taylor, E. H. 1968. The Caecilians of the World: A Taxonomic
Review, pp. 1–848. University of Kansas Press, Kansas.
Teipel, H. 1932. Beitrag zue Kenntnis der Gymnophionen. XVI. Die
Zunge. – Zeitschrift f€ ur Anatomie und Entwicklungsgeschichte 98:
726–746.
Wake, M. H. 1976. The development and replacement of
teeth in viviparous caecilians. – Journal of Morphology 148: 33–
63.
Wake, M. H. 1978. Comments on the ontogeny of Typhlonectes obesus,
particularly its dentition and feeding. – Pap
eis Avulsos de Zoologia
32: 1–13.
Wake, M. H. 1980. Fetal tooth development and adult replacement
in Dermophis mexicanus (Amphibia: Gymnophiona): Fields versus
clones. – Journal of Morphology 166: 203–216.
Wake, M. H. 1992. Patterns of peripheral innervation of the tongue
and hyobranchial apparatus in caecilians (Amphibia: Gymno-
phiona). – Journal of Morphology 212: 37–53.
Wake, M. H. and Schwenk, K. 1986. A preliminary report on the
morphology and distribution of taste buds in Gymnophiones,
with comparison to other amphibians. – Journal of Herpetology 20:
254–256.
Wake, M. H. and Wurst, G. Z. 1979. Tooth crown morphology in
caecilians (Amphibia: Gymnophiona). – Journal of Morphology 159:
331–341.
Wiedersheim, R. 1879. Die Anatomie der Gymnophionen,
pp. 73–76. Gustav Fischer Verlag, Jena.
Wilkinson, M. 1991. Adult tooth crown morphology in the
Typhlonectidae (Amphibia). – Journal of Zoological Systematics and
Evolutionary Research 29: 304–311.
Xie, Z. H., Zhong, H. B., Li, H. J. and Hou, Y. J. 2011. The struc-
tural organization of the liver in the Chinese fire-bellied
newt (Cynops orientalis). – International Journal of Morphology 29:
1317–1320.
Zaher, M., El-Ghareeb, A.-W., Hamdi, H. and AbuAmod, F. 2012.
Anatomical, histological and histochemical adaptations of the avian
alimentary canal to their food habits: I-Coturnix coturnix. – Life
Science Journal 9: 253–275.
Zapata, A., Gomariz, R. P., Garrido, E. and Cooper, E. L. 1982.
Lymphoid organs and blood cells of the caecilian Ichthyophis
kohtaoensis. – Acta Zoologica 63: 11–16.
© 2016 The Royal Swedish Academy of Sciences
Acta Zoologica (Stockholm) 0: 1–19 (August 2016)
Zuasti, A., Jimenez-Cervantes, C., Garcıa-Borr
on, J. C. and Ferrer,
C. 1998. The melanogenic system of Xenopus laevis. – Archives of
Histology and Cytology 61: 305–316.
Zylberberg, L. 1972. Donnees histologiques sur les glandes linguales
d’Ichthyophis glutinosus (L.) (Batracien gymnophione). – Archives
D’Anatomie Microscopique et de Morphologie Experimentale 61:
227–241.
© 2016 The Royal Swedish Academy of Sciences
Muikham et al.  Digestive system of Supachai’s caecilian
Zylberberg, L. 1977. Histochemistry and ultrastructure of amphibian
lingual glands and phylogenetic relations. – The Histochemical Jour-
nal 9: 505–520.
Zylberberg, L. 1986. L’epithelium lingual de deux Amphibiens
Gymnophiones: Typhlonectes compressicaudus et Ichthyophis
kohtaoensis. – M
emoires de la Soci
et
e Zoologique de France 43:
83–84.
19