Revista de Biología Tropical, ISSN: 2215-2075, Vol. 71: e54253, enero-diciembre 2023 (Publicado Oct. 25, 2023)

Trophic morphology and diet of the endangered fish

Tlaloc hildebrandi (Cyprinodontiformes: Profundulidae)

Sara Elizabeth Domínguez-Cisneros1; https://orcid.org/0000-0003-1707-2556

Abraham Kobelkowsky2; https://orcid.org/0000-0003-2716-7233

Ernesto Velázquez-Velázquez1*; https://orcid.org/0000-0003-1884-0502

1. Museo de Zoología, Instituto de Ciencias Biológicas, Universidad de Ciencias y Artes de Chiapas, Tuxtla Gutiérrez,

Chiapas, México; sara.dominguez@unicach.mx, ernesto.velazquez@unicach.mx (*Correspondence)

2. Laboratorio de Peces, Departamento de Biología, Universidad Autónoma Metropolitana, Unidad Iztapalapa, Ciudad

de México, México; akd@xanum.uam.mx

Received 25-II-2023. Corrected 23-VI-2023. Accepted 02-X-2023.

ABSTRACT

Introduction: Tlaloc hildebrandi is a freshwater killifish, endemic to Southern Mexico and under threat of extinc-

tion; the knowledge of the trophic morphology and diet is needed by conservation managers.

Objective: To analyse and describe the anatomy of the visceral skeleton, visceral musculature, digestive tract and

its adjoining glands of T. hildebrandi; as well as its diet.

Methods: We performed the trophic anatomy on 20 adult specimens of both sexes, through manual dissection;

as well as gut content analysis in 60 individuals to describe the diet.

Results: As notable characters of the visceral skeleton of T. hildebrandi we found the posterior notch of the

premaxillary, the presence of the “coronoid cartilage”, the tricuspid shape of the gill rakers of the first branchial

arch, and the presence of the coronomeckelian bone; some outstanding characters of the visceral musculature

are the origin of the retractor dorsalis muscle from the first four vertebral centra, and the division of the pharyn-

goclavicularis externus muscle into two sections. The notable characters of the digestive tube are the absence of

stomach and pyloric caeca, and the presence of the “intestinal valve”. Insects (IVI = 66.6 %) and ostracods (13 %

IVI) were the dominant prey items of the Tlaloc hildebrandi diet; larvae and adults of the family Chironomidae

were the most dominant insects in the diet (53 % IVI ).

Conclusions: The organization of the digestive system of T. hildebrandi corresponds to the general morphologic

pattern of the Cyprinodontiformes; however, we register as new information for these fish, the presence of the

“coronoid cartilage” and the “intestinal valve”. The structures of the trophic morphology and the components of

the diet, confirms us that T. hildebrandi is a carnivorous-insectivorous fish.

Key words: Chiapas killifish; trophic anatomy; visceral cavity; digestive tract; food analysis.

RESUMEN

Morfología trófica y dieta de Tlaloc hildebrandi (Cyprinodontiformes: Profundulidae),

especie amenazada de extinción

Introducción: Tlaloc hildebrandi es un killi de agua dulce, endémico del sur de México y bajo amenaza de

extinción; el conocimiento de la morfología trófica y la dieta son necesarios para los administradores de la

conservación.

https://doi.org/10.15517/rev.biol.trop..v71i1.54253

VERTEBRATE BIOLOGY

2Revista de Biología Tropical, ISSN: 2215-2075 Vol. 71: e54253, enero-diciembre 2023 (Publicado Oct. 25, 2023)

Objetivo: Analizar y describir la anatomía del esqueleto visceral, la musculatura visceral, el tracto digestivo y las

glándulas adyacentes de T. hildebrandi; así como los componentes de su dieta.

Métodos: Mediante la técnica del descarnado manual, realizamos la descripción de la anatomía trófica en 20

especímenes adultos de ambos sexos, y el análisis del contenido estomacal en 60 individuos para describir la dieta.

Resultados: Como caracteres sobresalientes del esqueleto visceral de T. hildebrandi está la escotadura posterior

del premaxilar, la presencia del “cartílago coronoides”, la forma tricúspide de las branquiespinas del primer arco

branquial y la presencia del hueso coronomeckeliano; como caracteres de la musculatura visceral sobresalen el

origen del músculo retractor dorsalis de los cuatro primeros centros vertebrales, y la división del músculo pharyn-

goclavicularis externus en dos secciones. Los caracteres notables del tubo digestivo son la ausencia de estómago

y de ciegos pilóricos y la presencia de la “válvula intestinal”. Los insectos (IVI = 66.6 %) y ostrácodos (13 % IVI)

fueron los componentes dominantes de la dieta de T. hildebrandi; particularmente las larvas y adultos de la familia

Chironomidae fueron los insectos más abundantes en la dieta (53 % IVI).

Conclusiones. La organización del sistema digestivo de T. hildebrandi corresponde al patrón morfológico general

de los Cyprinodontiformes, sin embargo, se registra como nueva información para estos peces, la presencia del

cartílago coronoides y la válvula intestinal. Las estructuras de la morfología trófica y los componentes de la dieta

nos confirman que T. hildebrandi es un pez carnívoro-insectívoro.

Palabras clave: Chiapas killis; anatomía trófica; cavidad visceral; tracto digestivo; análisis alimenticio.

INTRODUCTION

Tlaloc hildebrandi (Miller, 1950), an

endemic fish of Chiapas, México (Velázquez-

Velázquez & Schmitter-Soto, 2004), is a small-

bodied profundulid fish (maximum size: 110.66

mm standard length (Domínguez-Cisneros et

al., 2017). Also known as the “escamudo de

San Cristóbal” or the “Chiapas killifish”, it

has been cataloged as an Endangered spe-

cies (P) under Mexican legislation (NOM-

059-SEMARNAT-2010) (Secretaría de Medio

Ambiente y Recursos Naturales, 2019) and also

in the International Union for Conservation of

Nature (IUCN) Red List of Threatened Species

(EN), due to its restricted distribution, invasive

species, and habitat vulnerability (Schmitter-

Soto & Vega-Cendejas, 2019).

This species is the only native fish to

inhabit the mountains of Chiapas, at altitudes

ranging from 2 110 to 2 360 m.a.s.l. Tlaloc

hildebrandi inhabits the Fogotico River and

its tributaries and in the upper reaches of the

Grijalva-Usumacinta system, in Chiapas, Méxi-

co (Beltrán-López et al., 2021; Domínguez-

Cisneros et al., 2017; Velázquez-Velázquez &

Schmitter-Soto, 2004). It exhibits a restricted

extent of occurrence, because the number of

locations where this species occurs is limited to

three, based on the recognition of at least three

evolutionarily significant units (Beltrán-López

et al., 2021; Velázquez-Velázquez et al., 2016).

The morphology of the mouth and diges-

tive tract of the fishes, together with the iden-

tification of the food content in the intestine,

allows the interpretation of the trophic biology

of the species (Karachle & Stergiou, 2010; Keast

& Webb, 1966; Wootton, 1998). Likewise, the

analysis and descriptions of its components

contribute to understanding its functions and

its role in the life history of the organism (Drewe

et al., 2004; Hale, 1965; Kapoor et al., 1976).

The digestive system of teleost fishes is

composed of the digestive tract and its associ-

ated glands (Moyle & Cech, 2000). The diges-

tive tract begins at the mouth and finishes at

the anus and may be divided into four general

sections based on functional and histological

criteria: bucco-pharyngeal cavity, esophagus,

stomach, and intestine. The stomach is the

most highly diversified region of the gut in tele-

ost fishes and has also undergone a number of

independent secondary losses, with stomach-

less fishes accounting for approximately 20 % of

teleost species (Wilson & Castro, 2011).

Among the few studies on the anato-

my of the digestive system of the Mexican

Cyprinodontiformes are those of Kobelkowsky

(2005) on Goodeidae and Hernández et al.,

(2009) on Rivulidae, Fundulidae, Poeciliidae,

Revista de Biología Tropical, ISSN: 2215-2075, Vol. 71: e54253, enero-diciembre 2023 (Publicado Oct. 25, 2023)

and Cyprinodontidae. Studies devoted to the

description of the digestive system of Profun-

dulidae are still lacking. Trophic morphology

and food habit studies of fishes are necessary

to understand the role they play in the tro-

phic food web (Gerking, 1994). These data

are essential to account as an important biotic

factor when making conservation and manage-

ment decisions. Understanding the relationship

between diet and morphological traits is criti-

cally relevant to the successful management of

threatened species such as T. hildebrnadi. The

specific aims of this study are: 1) to describe

the anatomy of the digestive system and 2) to

provide a quantitative description of the diet of

T. hildebrandi. In order to establish their feed-

ing habits, we take into account the relationship

between diet and trophic morphology.

MATERIALS AND METHODS

Material examined: Specimens deposited

in the Fish Collection of the Zoology Muse-

um of the University of Sciences and Arts

of Chiapas were analyzed (MZ-UNICACH);

twenty individuals of both sexes between 44.63-

102.06 mm standard length (SL) were used for

anatomical analysis: Tlaloc hildebrandi; MZ-

UNICACH 4301, 3; MZ-UNICACH 4346, 4;

MZ-UNICACH 4296, 4; MZ-UNICACH 4331,

6; MZ-UNICACH 4348,1; MZ-UNICACH

4330,1; MZ-UNICACH 4328, 3. All specimens

from the Amarillo River, in the municipal-

ity of San Cristóbal de Las Casas, Chiapas,

México. Fish were fixed in a 10 % formalin

solution and subsequent preserved in a 70 %

ethanol solution.

In order to expose the visceral musculature

and the visceral skeleton, manual dissection

and disarticulation technique were used (Bemis

et al., 2004; Kobelkowsky, 2005). The external

mouth morphology was described. In order to

expose the visceral musculature, the skin from

the cephalic region was removed. After the

analysis of these structures to allow the obser-

vation of the mandibular apparatus, palatine

series, the opercular series, hyoid apparatus

and branchial apparatus, the visceral muscles

were manually removed. The visceral cavity was

exposed, removing the body wall. In order to

examine the branchial musculature, the pecto-

ral girdle was removed. Osteological terminol-

ogy follows Gregory (1933), Parenti (1981) and

Costa (2006); the terminology of the visceral

musculature follows Winterbottom (1973). The

illustrations were done by means of a camera

lucida mounted in a stereoscopic microscope

Wild M3Z.

The gut content of 60 adult specimens of

both sexes between 24.8-114.2 mm standard

length (SL) were used for diet analysis. In each

case, the food content of the anterior region of

the intestine was squashed on a graduated slide

to a uniform depth and the area of the squash

was measured (Castillo-Rivera et al.,1996),

according to the method for measuring small

stomach volumes (Hyslop, 1980). Gut contents

were sorted and identified to the lowest taxo-

nomical level using standard taxonomic keys

(Merrit & Cummins, 1984; Needham & Need-

ham, 1962). Composition of gut content was

interpreted using two indices: percent frequen-

cy of occurrence (% F) that shows the percent-

age of guts that had a certain prey type present.

Percent numerical importance (% N) gives the

proportion of a prey group compared to the

total number of prey items examined for each

species. These two indices were also used to

generate an index of relative importance (IRI)

(McCune & Grace, 2002). These values were

totaled for all items and a %IRI is presented.

Intestinal index (Ii) was calculated using

the equation of Nikolski (1963); which defines

this index as the ratio of the length of the intes-

tine to the standard length of the fish. Accord-

ing to this index, fish can be classified into three

food categories: carnivores Ii < 1; omnivores Ii

between 1 and 2; herbivores Ii > 2.

RESULTS

Visceral skeleton: Jaws: The mouth of

T. hildebrandi has strong jaws, with lower jaw

strongly upwards (Fig. 1A). Premaxilla and

maxilla form the upper jaw, while dentary, angu-

loarticular, retroarticular and coronomeckelian

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form the lower jaw (Fig. 1B). The coronoid

cartilage is located on the external face of the

coronoid processes of the dentary and angu-

loarticular (Fig. 1B, Fig. 2A, Fig. 2D). The pre-

maxilla has an S shape and a short ascending

process with blunt border. The buccal border of

the premaxilla carries an external row of slim

conical, medium-sized teeth, and a numerous

group of smaller internal teeth. The rest of

the premaxilla is wide and abruptly descends

to form a wide notch on its posterior margin

where it contacts the coronoid cartilage (Fig.

2A, Fig. 2B).

The maxilla is an elongate and narrow

bone, with the anterior margin contacting the

superior edge of the premaxilla (Fig. 2A, Fig.

2B, Fig. 2C). The external surface of the maxilla

has a concave area where it joins the anterior

border of the palatine (Fig. 1B).

The coronoid process of dentary is elon-

gate and narrow, ending in a blunt angle, joined

to the external surface of the anguloarticular.

Fig. 1. Cephalic region of Tlaloc hildebrandi. A. Lateral view. B. Visceral skeleton: mandibular skeleton, palatine series,

mandibular suspension and opercular series. Arrow point to coronoid cartilage.

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Externally on the two bones it is the coronoid

(Fig. 2D). The dentary also has a short ventral

process in the lower side of the bone. The

anterior-dorsal margin of the dentary has two

rows of teeth, a single outer row of medium

sized-teeth, and several internal rows of smaller

teeth. The dentary has a branch of the lateral

line system (Fig. 2A, Fig. 2D).

The anguloarticular forms a large triangu-

lar process that inserts in the dentary anteriorly,

and forms the coronoid process joined dorsally

with the dentary (Fig. 2E). In its postero-infe-

rior angle the anguloarticular has a fossa that

Fig. 2. Mandibular skeleton of Tlaloc hildebrandi. A. Left lateral view of upper and lower jaws. B. Ventral view of the upper

jaw. C. Dorsal view of the upper jaw. D. Left lateral view of the lower jaw. E. Medial view of the lower jaw.

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joins with the condyle of the quadrate (Fig.

1B, Fig. 2D). The retroarticular is a small bone

located posteroventral to the anguloarticular

(Fig. 2D, Fig. 2E). The coronomeckelian is a

small and triangular bone that firmly joins the

medial surface of the anguloarticular (Fig. 2E).

Palatine series: The palatine series is

formed by the palatine and the entopterygoid

(Fig. 1B). The palatine is flat and straight, and

joined anteriorly to the maxilla.

The palatine makes contact through a car-

tilaginous portion of the quadrate, and medially

with parts of the entopterygoid. The entoptery-

goid is thin; its internal border is concave and

forms an anterior process that covers parts of

the internal surface of the palatine, contact-

ing the quadrate medially and the symplectic

posteriorly.

Mandibular suspension: The mandibular

suspensorium is formed by the hyomandibula,

symplectic and quadrate (Fig. 1B). The hyo-

mandibula articulates by means of condyles

with the sphenotic, pterotic, and prootic bones

of the neurocranium, and the opercle and

preopercle bones of the opercular series. The

anteromedial hyomandibula margin is undu-

lated and approximately vertical, and contacts

the lateral surfaces of the preopercle and sym-

plectic. The symplectic is broad and obliquely

oriented, with a robust medial section, and the

ventral margin is acute and located between

the entopterygoid and the quadrate (Fig. 1B).

The quadrate is relatively large and irregular-

shaped, the anterior portion is truncate and

enlarged anterodorsally, the posterior portion

has a thin retroarticular process connected with

the medial surface of the preopercle.

Opercle: The opercular series is formed

by the preopercle, opercle, interopercle and

subopercle bones (Fig. 1B). The preopercle is

L-shaped, with a large median shelf wrapping

around the symplectic, a convex anterior mar-

gin, and a laterosensory canal groove on its pos-

terior margin. The opercle is triangular, with a

convex superior edge, a truncate anterodorsal

process, and a straight posteroventral margin.

The subopercle is large with an acute anterior

end. The interopercle is elongate, with a point-

ed anterior end, and a ligamentous attachment

to the retroarticular.

Branchial apparatus: The branchial appa-

ratus is formed by two basibranchial, three

hypobranchial, five ceratobranchial, four epi-

branchial, and three pharyngobranchial bones

(Fig. 3A, Fig. 3B, Fig. 3C, Fig. 3D, Fig. 3E). The

articular faces of the first two basibranchials

contact the hypobranchials. The hypobran-

chials are rectangular-shaped and gradually

become smaller posteriorly (Fig. 3B, Fig. 3D).

Ceratobranchials 1-4 (Fig. 3D) have a groove

on their lateral surface to accommodate the

branchial arteries and are the largest bones,

of the branchial apparatus. The fourth cerato-

branchial bone has teeth in its dorsal surface.

The fifth ceratobranchial or lower pharyngeal

bone is robust with its anterior extreme convex

and the posterior extreme truncate, ventrally it

forms a condyle where it joins to the pharyngo-

clavicularis externus muscle. Its dorsal surface is

covered by teeth (Fig. 3D, Fig. 3E).

Epibranchials (Fig. 3A, Fig. 3B) are small

bones and joined by the levator externalis mus-

cle. The first epibranchial is bifurcated with

each branch contacting a cartilage, and lacking

a dorsal process. Epibranchials two and four

possess a dorsal process. The dorsal process of

the third epibranchial joins the anterior process

of the fourth epibranchial.

The pharyngobranchials or upper pharyn-

geal bones (Fig. 3C) are three flat elements of

the branchial apparatus, and have numerous

teeth on their ventral surfaces. The second pha-

ryngobranchial bone is the largest and the third

pharyngobranchial is the smallest, which unites

posteriorly with the retractor dorsalis muscle.

The anterior arm of the first branchial arch

of T. hildebrandi has 13 to 15 gill rakers. The gill

rakers of this species do not have teeth and are

found on both margins of branchial arches 1 to

3, with the exception of the internal surface of

branchial arch 4 (Fig. 3F). The gill rakers are

triangular-shaped, large and thin, with three

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cusps on the internal border of the first arch. In

contrast, they are triangular-shaped, small, and

without cusps in the rest of the branchial arches.

The teeth of T. hildebrandi are conical and

slightly curved backwards. Teeth are found

on the following bones: premaxilla, dentary,

ceratobranchial 4, and pharyngobranchials 1

to 3. The teeth in the premaxilla and dentary

are arranged in two groups. The first row is a

single group of large teeth with about the same

Fig. 3. Branchial skeleton of Tlaloc hildebrandi. A. Left lateral view of the branchial apparatus. B. Dorsal view of the branchial

apparatus. C. Ventral view of the upper pharyngeal bones. D. Ventral view of the branchial apparatus. E. Dorsal view of

the lower pharyngeal bone. F. Lateral view of a gill raker on the external row of the first branchial arch; numbers 1-5,

ceratobranquiales.

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size; the second row lies behind the first, and

has numerous irregularly arranged and smaller

teeth (Fig. 2A, Fig. 2B).

The teeth of ceratobranchial 4 are located

(Fig. 3B, Fig. 3F) on the anterior portion of the

bone. The teeth of the lower pharyngeal bones

are larger on the medial border and gradually

become smaller at the external edge (Fig. 3E).

The teeth of pharyngobranchial 1 are confined

to its posterior edge; teeth cover the majority

of the ventral surface of pharyngobranchial 2,

and the entire surface of pharyngobranchial 3

(Fig. 3C).

Hyoid apparatus: The hyoid apparatus

is formed by the glossohyal, basihyal, uro-

hyal, dorsal and ventral hypohyals, ceratohyal,

epihyal, interhyal, and six branchiostegal-ray

bones (Fig. 4A). The basihyal is enlarged and

joins anteriorly to the glossohyal. The urohyal

is enlarged, laterally compressed, triangular

in a lateral view, and the posterior portion is

convex (Fig. 4B). Dorsal hypohyal absent; ante-

rior extension of anterior ceratohyal ventral to

ventral hypohyal. The ceratohyal is elongate

and narrow in the middle where it joins the two

branchiostegal rays and posteriorly joins three

branchiostegal rays (Fig. 4A).

The epihyal is triangular-shaped and artic-

ulates anteriorly with the ceratohyal by a suture,

and it also contacts the sixth branchiostegal ray.

The epihyal joins the interhyal at its antero-

posterior margin. The interhyal is cylindrical-

shaped, thin and contacts the preopercle by

means of a cartilage.

Visceral musculature: The visceral mus-

culature of T. hildebrandi is organized into

mandibular, hyoid and branchial arches. The

most complex mandibular muscle is the adduc-

tor mandibulae (Fig. 5) which is divided into

four sections: A1, A2, A3, and Aw (Fig. 5C). Its

origin is from several areas of the mandibular

suspension and preopercular, its insertion is

on both the upper and lower jaws. The levator

arcus palatini muscle originates from the lateral

process of the sphenotic and inserts on the lat-

eral surface of the hyomandibula. The adductor

arcus palatini muscle originates from the paras-

phenoid and it inserts on the entopterygoid and

palatine (Fig. 5A, Fig. 5C). The intermandibu-

laris muscle (Fig. 4D) is narrow and inserts on

the internal surface of both dentaries near the

mandibular symphysis.

The protractor hyoideus muscle (Fig. 4D)

originates from the dentary and inserts on

the ceratohyal, reaching the third branchioste-

gals. The retractor dorsalis muscle (Fig. 4C) is

formed by several sections that originate from

precaudal vertebrae 1-4 and inserts on the pos-

terior border of the third pharyngobranchial.

The dilatator operculi muscle originates from

the sphenotic process and inserts on the oper-

cular anterodorsal process. The levator operculi

muscle originates from the pterotic and inserts

on the internal surface of the opercular, near its

superior margin (Fig. 5B, Fig. 5C). The levatores

externi muscles originate from the sphenotic

and insert on all four epibranchial bones. The

levatores interni muscles are situated medially

to the levatores externi (Fig. 4B). The sterno-

hyoideus muscle (Fig. 4B) originate from the

cleithrum and inserts on the posterior border of

the urohyal. The pharyngoclavicularis externus

muscle (Fig. 4B) is divided into two sections

(1 and 2) and originates from the anteroventral

portion of the cleithrum and inserts on the

ventral surface of the lower pharyngeal bone.

The pharyngoclavicularis internus muscle (Fig.

4B) originates from the anterior surface of the

cleithrum and inserts on the ventral surface of

the lower pharyngeal bone.

Visceral cavity: The visceral cavity of T.

hildebrandi (Fig. 6A) is wide and enclosed by

the precaudal vertebrae, the first hemal arch,

the anal fin musculature, the infracarinalis

media muscle, the pelvic girdle, infracarinales

anteriores muscles, the pectoral girdle, pleural

ribs, epipleural ribs and axial musculature. The

liver and gas bladder occupy about 50 % of the

visceral cavity; the pancreatic tissue gradually

invades the liver along the branches of the por-

tal vein. The combined hepatic and pancreatic

tissue are collectively called the hepatopancreas.

The digestive tube, spleen and gonads occupy

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Fig. 4. Hyoid and branchial musculature of Tlaloc hildebrandi. A. Lateral left view of the buccopharyngeal cavity and

hyoid apparatus. B. Lateral left view of the branchial and hyoid musculature; LE, levatores externi, LI, levatore interni; 1-2,

pharyngoclavicularis externus C. Lateral view of the retractor dorsalis muscle; numbers 1-5, precaudal vertebrae D. Ventral

view of the gular region.

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Fig. 5. Mandibular musculature of Tlaloc hildebrandi. A. Left lateral view of the cephalic region. B. Adductor mandibulae

muscle projecting section A1, A2 and A3 C. Mandibular and opercular musculature.

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the remaining space. The gas bladder does not

have a pneumatic duct and makes contact with

the dorsal surfaces of the stomach and gonads.

The gonads develop between the gas bladder

and the intestine. The female genital opening

and the male urogenital opening are located

immediately behind the anus.

The digestive tube of T. hildebrandi is

formed by the esophagus and the intestine.

The stomach is missing (Fig. 6B, Fig. 6C). The

esophagus begins from the upper and lower

pharyngeal bones and it is funnel-like (Fig.

7A, Fig. 7B); numerous thin longitudinal folds

and grooves form the lining. The first third of

Fig. 6. Visceral cavity of Tlaloc hildebrandi. A. Left lateral view; ER, epipleural ribs B. Left lateral view of the digestive tube

and liver. C. Right lateral view of the digestive tube and liver.

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intestine is wide and irregular folds form the

lining (Fig. 7B). Between the esophagus and

intestine is the connection of the common

biliary duct, being the feature indicating the

absence of the stomach.

The intestine is relatively short, and dis-

plays a single fold located below the anterior

portion of intestine. The intestinal index was

determined as 0.3 (0.24-0.45). The rest of the

intestine is straight and oriented posteriorly

(Fig. 6B, Fig. 6C, Fig. 7A, Fig. 7B). The surface

of the intestinal lumen is covered by folds and

shallow grooves in a reticulate pattern. An

intestinal valve is posteriorly situated, dividing

the intestines into anterior and posterior por-

tions (Fig. 7B, Fig. 7C). The last portion of the

posterior intestine is the rectum, which may be

recognized by the presence of longitudinal and

shallow grooves and folds.

The liver has two lobes; the left lobe is

always larger and covers a greater portion of

the stomach on its left side, and the initial part

of the intestine (Fig. 6B). The right lobe of the

liver is smaller and forms a groove to accom-

modate the gall bladder. The gall bladder is

spherical-shaped (Fig. 6C).

Fig. 7. Digestive tube of Tlaloc hildebrandi. A. Ventral view. B. Ventral view of the frontal section of the digestive tube. C.

Section of the intestines with the intestinal valve in ventral view. D. The intestinal valve divides the intestine in anterior

(prevalvular) and posterior (postvalvular) intestine.

Revista de Biología Tropical, ISSN: 2215-2075, Vol. 71: e54253, enero-diciembre 2023 (Publicado Oct. 25, 2023)

Diet: Of the total analyzed intestines, 11.7

% (7) was found empty and 88.3 % (53) were

found with food. Eighteen trophic categories

were recognized, being insects and crustaceans

the two largest taxa, together account for 97 %

of dominance (IVI). Organic matter (MON)

and unidentified fish scales and insects (INI)

were determined as trophic categories. Accord-

ing to Table 1, insects of the order Diptera,

Ephemeroptera, Hymenoptera and Coleoptera

were the dominant prey items (IVI = 66.6 %)

DISCUSSION

The anatomy of the digestive system of

Tlaloc hildebrandi largely corresponds to that

of teleost fishes. However, T. hildebrandi has the

“coronoid cartilage”, registered here for the first

time; the close location of this cartilage to the

posterior notch of the premaxillary, probably

limits the backward movement of jaws. The

anterior extension of the quadrate functionally

compensates for the absence of an ectoptery-

goid, which is a character of Cyprinodonti-

formes (Parenti, 1981).

The number of bones that carry teeth in T.

hildebrandi is relatively reduced compared with

other teleosts (Wainwright, 1989), including

those of the gill rakers. However, T. hildebrandi

exhibits teeth on fourth ceratobranchial bone.

The number of gill rakers in the first-

branchial arch is within the range of other

profundulid species (Domínguez-Cisneros et

al., 2023; Nelson et al., 2016). However, the

presence of large, thin and tricuspid gill rakers

on the first-branchial arch is a new observation

in T. hildebrandi.

The adductor mandibulae muscle is com-

posed of four sections as in many other teleosts

(Kenaley et al., 2019; Winterbottom, 1973); the

large volume of section A1 suggests a strong

movement of jaws during mouth closure.

The relative high number of sections of the

retractor dorsalis muscle suggests an effective

swallowing in contrast to what happens in other

teleosts (Wainwright, 1989).

The pharyngoclavicularis externus in T.

hildebrandi is formed by two elements (1 and

2), but in many teleost fishes this muscle has

a single element, as in G. atripinnis (Kobel-

kowsky, 2005).

The organography of the visceral cavity

of T. hildebrandi is the typical of that of teleost

fishes, as illustrated in Chirostoma estor (Kobel-

kowsky & Figueroa, 2018) and Eugerres mexi-

canus (Kobelkowsky & Terán-Martínez, 2020).

The absence of the stomach in T. hildeb-

randi observed in the present study coincides

with that of Hale (1965) in Poecilia reticu-

lata and that of Wilson and Castro (2011) in

Table 1

Diet composition of Tlaloc hildebrandi.

Food type Diet composition (%)

% N % FO % IRI

Insects 66.6

Diptera 55.9 50.4 53.1

Chironomidae

Chaoboridae 2.4 5.0 3.7

Simulidae 1.7 1.4 1.6

Dixidae 0.9 1.4 1.2

Syrphydae 1.6 1.4 1.5

Stratiomyidae 0.2 1.4 0.8

Ephemeroptera 3.4 4.3 3.8

Hymenoptera 0.2 0.7 0.4

Coleoptera 0.3 0.7 0.5

Crustaceans 31.3

Ostracoda 14.0 13.5 13.7

Amphipoda 6.2 5.7 5.9

Isopoda 9.6 8.5 9.1

Cladocera 2.0 1.4 1.7

Copepoda 0.2 1.4 0.8

Diplopoda 0.4 0.7 0.6

INI 0.6 0.7 0.7

MON 0.3 0.7 0.5

Fish scales 0.1 0.7 0.4

Bold value indicates the sum of each diet group (% IRI).

of the Tlaloc hildebrandi diet; ostracods (13 %

IVI) were recognized in the diet as a secondary

prey category and the rest of the prey compo-

nents as accidental (less than 10 %). The insects’

larvae of the Chironomidae family (Diptera),

accounted for 53 % (IVI) of the Chiapas killifish

diet (Table 1).

14 Revista de Biología Tropical, ISSN: 2215-2075 Vol. 71: e54253, enero-diciembre 2023 (Publicado Oct. 25, 2023)

Cyprinodontiformes. The thickening of the

first portion of the intestine in Goodea atripin-

nis (Kobelkowsky, 2005) is misnamed as stom-

ach. The character denoting the absence of the

stomach is the communication of the common

bile duct to the digestive tube at the junction of

the esophagus with the intestine.

Gut type and modes of intestinal morphol-

ogy have been used for the categorization of

fish in relation to feeding (Karachle & Stergiou,

2010). Furthermore, it has been shown that for

a given body length, gut length of herbivorous

fish is larger than that of omnivores, and that

of omnivores larger than that of carnivorous

species (Karachle & Stergiou, 2010; Nikolski,

1963). The values estimate of intestinal index

in T. hildebrandi was of 0.24-0.45; this Ii-values

varied within the expected range (<1) for car-

nivores species (Karachle & Stergiou, 2010;

Ward-Campbell et al., 2005). In this sense, T.

hildebrandi diet in wild environments consists

predominantly of insects.

Teleost fishes commonly have pyloric

caeca (Sano, 2021), however they are absent in

T. hildebrandi. The presence of the intestinal

valve is a remarkable character of the intestine

in T. hildebrandi, as is also observed in some

flatfish species of Paralichthyidae (Gisbert et

al., 2004; Kobelkowsky & Rojas-Ruiz, 2017)

and of species of Sparidae (Cataldi et al., 1987).

These fish are carnivores, which are also char-

acterized by the presence of a very short intes-

tine. Although there are substantial changes

to the bony elements of the feeding apparatus

within cyprinodontiforms (Hernández et al.,

2009) changes in intestinal morphology, may

be just as important in enhancing novel feeding

modes. One of these structures is the intestinal

valve; the intestinal valve appears as a constric-

tion of the intestinal mucosa dividing the intes-

tine in two regions, the prevalvular (anterior)

and postvalvular (posterior) intestine (Gisbert

et al., 2004). It is possible that the valve act then

passively avoiding the reflux of the intestinal

material, as well as directing the gut contents to

the rectum (Nachi et al., 1998; Oliveira-Ribeiro

& Fanta, 2000).

Stomach content analysis is widely used to

determine food composition, feeding strategies,

trophic position, energy flow (Hyslop, 1980).

Food habit and trophic morphology studies of

fishes are necessary to understand the role they

play in the food web (Gerking, 1994; Luczkov-

ich et al., 1995; Pease et al., 2020). We found

that T. hildebrandi fed mainly on insects (66.6

% IRI), especially Chrironomus larvae (> 50 %

IRI). The feeding behavior of T. hildebrandi is

scarcely documented; Velázquez-Velázquez et

al. (2007) described aspects related to feeding,

reproduction and growth of the Chiapas kil-

lifish; they concluded that it is an insectivorous

fish, with a specialized diet, based mainly on

insect larvae.

The descriptive and comparative mor-

phology has played an important role in the

reconstruction of the evolutionary history

and classification of cyprinodontiform fishes,

often providing useful phylogenetic informa-

tion at different taxonomic levels (Costa, 2006;

Dominguez-Cisneros et al., 2023; Ghedotti

& Davis, 2013; González-Díaz et al., 2014).

In addition, morphological characterization

provides a good approximation of feeding

modes or types of prey that are used dif-

ferentially by species (Ornelas-García et al.,

2018). The descriptive nature of our results

provides an effective method to analyze the

relationship between diet and morphology of

cyprinodontiform fishes. This study showed a

significant correlation between ecomorpho-

logical traits and trophic habits (diet com-

position); this information may be useful

for ecological niche studies (Calixto-Rojas

et al., 2021), among members of the family

Profundulidae and other groups within the

Cyprinodontoidei suborder.

Our study provided clear evidence that

diet-morphology specialization occurs in this

fish species studied. The food content in the

intestine, mainly of insects, the robustness of

the jaws, the low number of gill rakers and the

shortness of the intestine, are aspects that allow

us to conclude that T. hildebrandi is carnivore

fish of entomophagous type.

Revista de Biología Tropical, ISSN: 2215-2075, Vol. 71: e54253, enero-diciembre 2023 (Publicado Oct. 25, 2023)

Ethical statement: the authors declare that

they all agree with this publication and made

significant contributions; that there is no con-

flict of interest of any kind; and that we fol-

lowed all pertinent ethical and legal procedures

and requirements. All financial sources are fully

and clearly stated in the acknowledgments sec-

tion. A signed document has been filed in the

journal archives.

ACKNOWLEDGMENTS

This work was funded by a research grant

from the Fondo Mixto Consejo Nacional de

Ciencia y Tecnología/Gobierno del Estado de

Chiapas (CHIS-2005-C03-072). We thank Man-

uel Anzueto Calvo and Adán Gómez González,

for their collaboration during field and labora-

tory work. The Program para el Desarrollo

Profesional Docente (PRODEP) awarded a sup-

port scholarship for graduate studies to SEDC.

We thank Tania Kobelkowsky-Vidrio for the

English translation.

REFERENCES

Beltrán-López, R. G., González-Díaz, A., Soria-Barreto,

M., Garduño-Sánchez, M. A., Xochitla-Castrejón,

C., Rodiles-Hernández, R., & Ornelas-García, C. P.

(2021). Genetic diversity and structure of one of the

most endangered freshwater fish species in Mexico:

Tlaloc hildebrandi (Miller, 1950) and recognition of

its evolutionarily significant units. PeerJ, 9, e11952.

https://doi.org/10.7717/peerj.11952

Bemis, W. E., Hilton, J. E., Brown, B., Arrindell, R., Rich-

mond, A. M., Little, C. D., Grande, L., Forey, P. L.,

& Nelson, G. J. (2004). Methods for Preparing Dry,

Partially Articulated Skeletons of Osteichthyans, with

Notes on Making Ridewood Dissections of the Cra-

nial Skeleton. Copeia, 2004(3), 603–609.

Calixto-Rojas, M., Lira-Noriega, A., Rubio-Godoy, M.,

Pérez-Ponce de León, G., & Pinacho-Pinacho, C.

D. (2021). Phylogenetic relationships and ecological

niche conservatism in killifish (Profundulidae) in

Mesoamerica. Journal of Fish Biology, 99(2), 396–410.

Castillo-Rivera, M., Kobelkowsky, A., & Zamayoa, V.

(1996). Food resource partitioning and trophic mor-

phology of Brevoortia gunteri and B. patronus. Journal

of Fish Biology, 49, 1102–1111.

Cataldi, E., Cataudella, S., Monaco, G., Ross, A. I., &

Tancioni, L. (1987). A study of the histology and

morphology of the digestive tract of the sea-bream,

Sparus aurata. Journal of Fish Biology, 30, 135–145.

Costa, W. (2006). Descriptive morphology and phyloge-

netic relationships among species of the Neotropical

annual killifish genera Nematolebias and Simpsoni-

chthys (Cyprinodontiformes: Aplocheiloidei: Rivuli-

dae). Neotropical Ichthyology, 4(1),1–26.

Domínguez-Cisneros, S., Velázquez-Velázquez, E., Anzue-

to-Calvo, M., Gómez, G. A., Liévano, T. J., & Mata-

moros, W. (2017). Ampliación de la distribución

geográfica del popoyote de San Cristóbal Tlaloc

hildebrandi (Miller 1950) (Cyprinodontiformes: Pro-

fundulidae). Lacandonia, 11(2), 13–18.

Domínguez-Cisneros, S. E., Domínguez-Domínguez, O.,

Velázquez-Velázquez, E., & Pérez-Rodríguez, R.

(2023). Redescription and diagnoses of the genera

Profundulus and Tlaloc (Cyprinodontiformes: Pro-

fundulidae), Mesoamerican endemic fishes. Neotropi-

cal Ichthyology, 21(1), e220089.

Drewe, K. E., Horn, M. H., Dickson, K. A., & Gawlicka, A.

(2004). Insectivore to frugivore: ontogenetic changes

in gut morphology and digestive enzyme activity in

the characid fish Brycon guatemalensis from Costa

Rican rain forest streams. Journal of Fish Biology,

64(4), 890–902.

Gerking, S. D. (1994). Feeding ecology of fish. Academic

Press.

Ghedotti, M. J., & Davis, M. P. (2013). Phylogeny, classifica-

tion, and evolution of salinity tolerance of the North

American topminnows and killifishes, family Fundu-

lidae (Teleostei: Cyprinodontiformes). Fieldiana Life

and Earth Sciences, 7, 1–65.

Gisbert, E., Piedrahita, R., & Conklin, D. (2004). Ontogene-

tic development of the digestive system in California

halibut (Paralichthys californicus) with notes on fee-

ding practices. Aquaculture, 232, 455–470.

González-Díaz, A. A, Díaz-Pardo, E., Soria-Barreto, M.,

& Martínez-Ramírez, E. (2014). Diferencias Osteo-

lógicas entre los Subgéneros Profundulus y Tlaloc

(Teleostei: Profundulidae). International Journal of

Morphology, 32(3), 1074–1078.

Gregory, W. K. (1933). Fish skulls: a study of the evolution

of natural mechanisms. Transaction America of the

American Philosophical Society, 23(2), 75–81.

Hale, P. A. (1965). The morphology and histology of the

digestive systems of two freshwater teleosts, Poecilia

reticulata and Gasterosteus aculeatus. Journal of Zoo-

logy, 146(2), 32–49.

Hernández, L. P., Gibbs, A. C., & Ferry-Graham, L. (2009).

Trophic apparatus in Cyprinodontiform fishes:

Functional specializations for picking and scraping

behaviors. Journal of Morphology, 270, 645–661.

16 Revista de Biología Tropical, ISSN: 2215-2075 Vol. 71: e54253, enero-diciembre 2023 (Publicado Oct. 25, 2023)

Hyslop, E. J. (1980). Stomach contents analysis-a review of

methods and their application. Journal of Fish Biology,

17(4), 411–429.

Kapoor, B. G., Smit, H., & Verighina, I. A. (1976). The ali-

mentary canal and digestion in Teleosts. Advances in

Marine Biology, 13, 109–239.

Karachle, K. P., & Stergiou, K. (2010). Intestine mor-

phometrics of fishes: a compilation and analysis of

bibliographic data. Acta Ichthyologica et Piscatoria,

40(1), 45–54.

Keast, A., & Webb, D. (1966). Mouth and body form relative

to feeding ecology in the fish fauna of a small lake,

Lake Opinicon, Ontario. Journal of Fisheries Research

Board of Canada, 23(12), 1845–1874.

Kenaley, P. C., Mareckib, C. M., & Lauderb, V. G. (2019).

The role of an overlooked adductor muscle in the

feeding mechanism of rayfinned fishes: Predictions

from simulations of a deep-sea viperfish. Zoology,

135, 125678.

Kobelkowsky, A. (2005). General anatomy and sexual

dimorphism of Goodea atripinnis (Teleostei: Goodei-

dae). In M. C. Uribe, & H. J. Grier (Eds.), Viviparous

fishes (pp. 483–498). New Life Publications.

Kobelkowsky, A., & Figueroa, L. G. (2018). Anatomía del

sistema digestivo del pescado blanco Chirostoma

humboldtianum (Teleostei: Atherinopsidae). Hidro-

biológica, 28(1), 37–50.

Kobelkowsky, A., & Rojas-Ruiz, M. I. (2017). Anatomía

comparada del sistema digestivo de los lenguados

Syacium papillosum y Syacium gunteri (Pleuronecti-

formes: Paralichthyidae). Revista de Biología Marina

y Oceanografía, 50(2), 255–273.

Kobelkowsky, A., & Terán-Martínez, J. (2020). Anatomy

of the visceral cavity of Eugerres mexicanus (Teleos-

tei: Gerreidae). Revista de Biología Tropical, 68(1),

189–199.

Luczkovich, J. J., Norton, S. E., & Gilmore, R. G. Jr. (1995).

The influence of oral anatomy on prey selection

during the ontogeny of two percoid fishes, Lagodon

rhomboides and Centropomus undecimalis. Environ-

mental Biology of Fishes, 44, 79–95.

McCune, B., & Grace, J. B. (2002). Analysis of Ecological

Communities. MjM Software Design.

Merrit, R. W., & Cummins, K. W. (1984). An introduction

to the aquatic insects of North America. Kendall Hunt.

Miller, R. R. (1950). Profundulus hildebrandi, a new cypri-

nodontid fish from Chiapas, Mexico. Copeia, 1, 22–30.

Moyle, P. B., & Cech, J. J. (2000). Fishes. An introduction to

Ichthyology. Pearson Prentice Hall.

Nachi, M. A., Hernandez-Blazquezb, F. J., Barbieric, R. L.,

Leite, R. G., Ferri, S., & Phan, M. T. (1998). Intestinal

histology of a detritivorous (iliophagous) fish Pro-

chilodus scrofa (Characiformes, Prochilodontidae).

Annales des Sciences Naturelles, 2, 81–88.

Needham, P. R., & Needham, J. G. (1962). A guide to the

study of Fresh-water Biology. Holden Day.

Nelson, J. S., Grande, T. C., & Wilson, M. V. H. (2016).

Fishes of the World. John Wiley & Sons.

Nikolsky, G. V. (1963). The ecology of fishes. Academic

Press.

Oliveira-Ribeiro, C. A., & Fanta, E. (2000). Microsco-

pic morphology and histochemistry of the digestive

system of a tropical freshwater fish Trichomycterus

brasiliensis (U.itken) (Siluroidei, Trichomycteridae).

Revista Brasileira de Zoologia, 17(4), 953–971.

Ornelas-García, C. P., Córdova-Tapia, F., Zambrano, L.,

Bermudez-González, M. P., Mercado-Silva, N., Men-

doza-Garfias, B., & Bautista, A. (2018). Trophic spe-

cialization and morphological divergence between

two sympatric species in Lake Catemaco, Mexico.

Ecology and Evolution, 8, 4867–4875.

Parenti, L. R. (1981). A phylogenetic and biogeographic

analysis of Cyprinodontiform fishes (Teleostei, Athe-

rinomorpha). Bulletin of the American Museum of

Natural History, 168, 335–557.

Pease, A. A., Soria-Barreto, M., González-Díaz, A., &

Rodiles-Hernández, R. (2020). Seasonal Variation in

Trophic Diversity and Relative Importance of Basal

Resources Supporting Tropical River Fish Assembla-

ges in Chiapas, Mexico. Transactions of the American

Fisheries Society, 149(6), 753–769.

Sano, Y., Kambe, H., & Kihara, M. (2021). Morphological

changes of pyloric caeca and their relevancy to moti-

lity in laboratory-reared Kurosoi rockfish (Sebastes

schlegelii Hilgendorf), using an in vitro assay method.

Aquaculture, 539(2021), 736604.

Schmitter-Soto, J., & Vega-Cendejas, M. (2019). Tlaloc hil-

debrandi. IUCN Red List of Threatened Species, Swit-

zerland. http://dx.doi.org/10.2305/IUCN.UK.2019-2.

RLTS.T169366A1274187.en

Secretaría de Medio Ambiente y Recursos Naturales.

(2019). Modificación del Anexo Normativo III, Lista de

especies en riesgo de la Norma Oficial Mexicana NOM-

059-SEMARNAT-2010, Protección ambiental-Especies

nativas de México de flora y fauna silvestres-Categorías

de riesgo y especificaciones para su inclusión, exclusión

o cambio-Lista de especies en riesgo 2010. Diario Ofi-

cial de la Federación, México.

Velázquez-Velázquez, E., López-Vila, J. M., Gómez-Gon-

zález, A., Romero-Berny, E., Lievano-Trujillo, J. L., &

Matamoros, W. (2016). Checklist of the continental

Revista de Biología Tropical, ISSN: 2215-2075, Vol. 71: e54253, enero-diciembre 2023 (Publicado Oct. 25, 2023)

fishes of the state of Chiapas, Mexico, and their distri-

bution. ZooKeys, 632, 99–120.

Velázquez-Velázquez, E., & Schmitter-Soto, J. (2004). Con-

servation status of the San Cristobal pupfish Profun-

dulus hildebrandi Miller (Teleostei: Profundulidae) in

the face of urban growth in Chiapas, Mexico. Aquatic

Conservation: Marine and Freshwater Ecosystems, 14,

201–209.

Velázquez-Velázquez, E., Domínguez, R. E., Domínguez, C.

S., Hernández, S. J., Rodríguez, M. R. (2007). Mono-

grafia de Profundulus hildebrandi Miller, 1950, pez

endémico de Chiapas. Universidad de Ciencias y Artes

de Chiapas, México.

Wainwright, P. C. (1989). Functional Morphology of the

Pharyngeal Jaw Apparatus in Perciform Fishes: An

Experimental Analysis of the Haemulidae. Journal of

Morphology, 200, 231–245.

Ward-Campbell, B. M. S., Beamish, F. W. H., & Kongchaiya,

C. (2005). Morphological characteristics in relation to

diet in five coexisting Thai fish species. Journal of Fish

Biology, 67, 1266–1279.

Wilson, J. M., & Castro, L. F. C. (2011). Morphological

diversity of the gastrointestinal tract in fishes. In

M. Grosell, A. P. Farrell, & C. J. Brauner (Eds.), The

multifunctional gut of fish (pp. 1–55). Academic Press.

Winterbottom, R. (1973). A descriptive synonymy of the

striated muscles of the Teleostei. Proceeding of the

Academy of Natural Sciences of Philadelphia, 125(12),

225–317.

Wootton, R. J. (1998). Ecology of teleost fishes. Kluwer Aca-

demic Publishers.