Revista de Biología Tropical, ISSN: 2215-2075, Vol. 73: e63718, enero-diciembre 2025 (Publicado Jul. 29, 2025)

Morphology, histology and histochemistry of the gastrointestinal tract

of the fish Prochilodus magdalenae (Characiformes: Prochilodontidae)

Andrés Felipe Monsalve-Blandón1; https://orcid.org/0009-0005-7791-6089

Irina López-Montaño1; https://orcid.org/0009-0000-1250-8635

Ana Estrada-Posada2; https://orcid.org/0000-0003-3585-3719

Mariana Gutiérrez-Espinosa3; https://orcid.org/0000-0001-6127-9955

Gersson Vásquez-Machado4; https://orcid.org/0000-0002-4737-7038

Jonny Yepes-Blandón1*; https://orcid.org/0000-0001-6276-5488

1. Grupo de Investigación en Organismos Acuáticos Nativos y Exóticos (GIOANE), Universidad de Antioquia,

Medellín, Antioquia, Colombia; afelipe.monsalve@udea.edu.co, irina.lopez@udea.edu.co, jonny.yepes@udea.edu.co

(Correspondencia*)

2. ISAGEN S.A. E.S.P, Medellín, Antioquia, Colombia; aestrada@isagen.com.co

3. Centro de Estudios e Investigación en Acuicultura CEIAC, Guamal, Meta, Colombia; marianacgutierreze@gmail.com

4. HISTOLAB, Bogotá D.C., Cundinamarca, Colombia; gmvasquezm@unal.edu.co

Received 30-I-2025. Corrected 21-IV-2025. Accepted 24-VI-2025.

ABSTRACT

Introduction: The Bocachico Prochilodus magdalenae, an endemic herbivorous/detritivorous fish from the

Magdalena River basin, is the region’s most important fishing resource, with high ecological and socioeconomic

significance. Its populations have declined significantly due to fishing pressure and anthropogenic environmental

stressors, including wastewater discharge, infrastructure projects, and material extraction. Understanding the

characteristics of its digestive tract is a crucial knowledge gap, hindering the comprehension of nutritional and

physiological processes vital for aquaculture and both in situ and ex situ conservation efforts.

Objective: To characterize the morphological, histological, and histochemical features of P. magdalenae’s gastro-

intestinal tract (GIT) and relate them to its feeding habits.

Methods: Fifteen adult specimens from the Sogamoso River (Santander, Colombia) were analyzed using stan-

dard morphological measurements and histological and histochemical staining techniques for tissue structure

and mucin distribution analysis. Descriptive statistics were used for morphological analysis.

Results: Specimens averaged 24.66 cm standard length, 291.77 g weight, and 1.02 condition factor, with a mean

GIT weight of 12.02 g. The digestive tract showed four distinct regions: oropharyngeal cavity, esophagus (mean

length 1.26 cm), stomach, and intestine (mean length 88.18 cm). The esophagus contained abundant acinar

glands, producing neutral and acidic mucins, with acidic mucins predominating. The U-shaped stomach showed

simple columnar epithelium with tubular gastric glands producing only neutral mucins. The intestine, lined

with simple columnar epithelium, exhibited differential mucin distribution: high neutral mucins anteriorly and

predominantly acidic mucins in middle and posterior segments.

Conclusions: The gastrointestinal characteristics align with its feeding habits, particularly the elongated intes-

tine and thick pyloric muscular layer. The distinct mucin distribution pattern, especially the carboxylated acidic

mucins in the middle and posterior intestinal segments, supports nutrient absorption functions. These findings

provide crucial reference points for fish welfare assessment, pathogen response in aquaculture, and environmen-

tal impact evaluation in natural habitats.

Key words: histochemistry; histology; mucins; mucous cells; detritivorous fish.

https://doi.org/10.15517/rev.biol.trop..v73i1.63718

VERTEBRATE BIOLOGY

2Revista de Biología Tropical, ISSN: 2215-2075 Vol. 73: e63718, enero-diciembre 2025 (Publicado Jul. 29, 2025)

INTRODUCTION

The Bocachico (Prochilodus magdalenae),

an endemic herbivorous/detritivorous fish from

the Magdalena River basin, is the region’s most

important fishing resource, with high ecological

and socioeconomic significance. In Colombia,

total fishing of P. magdalenae has significantly

decreased, dropping from 38 000 tons in 1978

to 6 000 tons in 1999 (Mojica et al., 2012),

representing an 84 % decline. By 2018, this

quantity reduced to 47.9 % of the total capture

(Doria-Gonzalez et al., 2020), raising concerns

among environmental authorities regarding the

utilization of this hydrological resource due to

its vulnerable conservation status in Colom-

bian territory (Mojica et al., 2012). This species

holds substantial socio-economic importance

and contributes to ecological equilibrium and

nutrient mobilization (Albus, 2012; Allan &

Castillo, 2007; Couto et al., 2021; Flecker, 1996).

Is also important to note that fish represent

the most diverse and abundant vertebrates

globally, encompassing approximately 34 000

known species to date (Castro & Vari, 2004;

Reis et al., 2016). In the Neotropical region, it

is estimated that over 4 500 species inhabit its

waters (Burns, 2021; Flecker, 1996; Wilson &

Castro, 2010). This considerable diversity arises

from evolutionary adaptations allowing them

to exploit nutrient availability across various

environments and trophic levels (Castro & Vari,

2004; Reis et al., 2016).

One of the most significant anatomical

and physiological adaptations in fish is their

capability to consume detritus as a nutrient

source (detritivorous fish) (Agostinho et al.,

2008; Benedito et al., 2018; Bowen, 2022; Burns,

RESUMEN

Morfología, histología e histoquímica del tracto gastrointestinal

del pez Prochilodus magdalenae (Characiformes: Prochilodontidae)

Introducción: El Bocachico Prochilodus magdalenae, un pez herbívoro/detritívoro endémico de la cuenca del río

Magdalena, es el recurso pesquero más importante de la región, con alta importancia ecológica y socioeconómi-

ca. Sus poblaciones han disminuido significativamente debido a la presión pesquera y los estresores ambientales

antropogénicos, incluyendo vertimientos, proyectos de infraestructura y extracción de materiales. Comprender

las características de su tracto digestivo es un vacío crucial de conocimiento, que dificulta el entendimiento de los

procesos nutricionales y fisiológicos vitales para la acuicultura y la conservación in situ y ex situ.

Objetivo: Caracterizar los aspectos morfológicos, histológicos e histoquímicos del tracto gastrointestinal (TGI)

de P. magdalenae y relacionarlos con sus hábitos alimenticios.

Métodos: Se analizaron quince especímenes adultos del río Sogamoso (Santander, Colombia) mediante medicio-

nes morfológicas estándar y técnicas de tinción histológicas e histoquímicas para el análisis de estructura tisular

y distribución de mucinas. Se empleó estadística descriptiva para el análisis morfológico.

Resultados: Los especímenes promediaron 24.66 cm de longitud estándar, 291.77 g de peso y 1.02 de factor de

condición, con un peso medio del TGI de 12.02 g. El tracto digestivo mostró cuatro regiones distintas: cavidad

orofaríngea, esófago (longitud media 1.26 cm), estómago e intestino (longitud media 88.18 cm). El esófago con-

tenía abundantes glándulas acinares que producen mucinas neutras y ácidas, con predominio de las ácidas. El

estómago en “U” mostró epitelio columnar simple con glándulas gástricas tubulares con solo mucinas neutras. El

intestino, revestido de epitelio columnar simple, exhibió distribución diferencial de mucinas: mucinas neutras en

la parte anterior y mucinas ácidas en los segmentos medio y posterior.

Conclusiones: Las características gastrointestinales se alinean con los hábitos alimenticios, particularmente el

intestino elongado y la gruesa capa muscular pilórica. El patrón de distribución distintivo de mucinas, espe-

cialmente las mucinas ácidas carboxiladas en segmentos intestinales medios y posteriores, sustenta funciones

de absorción de nutrientes. Estos hallazgos proporcionan puntos de referencia cruciales para la evaluación del

bienestar de los peces, la respuesta a patógenos en acuicultura y la evaluación del impacto ambiental en hábitats

naturales.

Palabras clave: histoquímica; histología; mucinas; células mucosas; peces detritívoros.

Revista de Biología Tropical, ISSN: 2215-2075, Vol. 73: e63718, enero-diciembre 2025 (Publicado Jul. 29, 2025)

2021; Natale et al., 2025; Wilson & Castro,

2010). Detritus consists of solid residues result-

ing from the decomposition of organic matter

(Benbow et al., 2020; Bowen, 2022) and plays

a crucial role in the food chain by releasing

nutrients during decomposition that can be

utilized by various species of microorganisms,

invertebrates, and fish (Bowen et al., 2006;

Bowen, 2022).

Another adaptation in fish involves modi-

fications in their gastrointestinal tract (GIT).

Detritivores typically have a longer intesti-

nal length compared to their stomach, while

carnivores exhibit a shorter intestinal length

(Morales-González, 2023; Osorio-Urtecho,

2018; Salinas-Torres, 2011). Additionally, the

GIT of fish consists of four concentric layers:

the mucosal layer, submucosal layer, muscular

layer, and serosal layer, from innermost to out-

ermost, respectively. The mucosal layer, which

borders the GIT lumen, serves the function of

providing physical, chemical, and immunologi-

cal protection to the tissue (Cao & Wang, 2009;

Genten et al., 2008; Vidal et al., 2020).

This protective role is achieved through

the secretion of glycoproteins known as mucins

by goblet cells, constituting the major fraction

of mucus and forming a natural barrier on the

epithelial surface (Koshio, 2015). Acidic and

neutral mucins are secreted in the mucosal

layer of the esophagus, stomach, and intestine.

These present a protection function by trapping

water-soluble particles and defending them

against pathogens. Furthermore, they protect

the epithelium from the pH of gastric juices

and act as lubricants, facilitating food passage

(Koshio, 2015; Rodrigues & Cargnin-Ferreira,

2017; Vidal et al., 2020).

The study of gastrointestinal tract’s (GIT)

characteristics has been of great interest in Neo-

tropical fish research (Lasso et al., 2011), since

it varies depending on the type of diet and the

microenvironments in which the individuals

develop (dos Santos et al., 2015). Investiga-

tions have been conducted on various fish spe-

cies such as Blanquillo (Sorubim cuspicaudus)

(Oviedo-Montiel, 2021), Escalar (Pterophyllum

scalare) (Ramírez-Espitia et al., 2020), Dentón

(Megaleporinus muyscorum) (Mosquera-Ramos

& Gómez-Ramírez, 2024), among others. In the

case of the Characiformes order, the Prochi-

lontidae family comprises several species like

Prochilodus lineatus (found in Argentina, Para-

guay, and Uruguay) (Bowen, 2022), Prochilodus

reticulatus (Venezuela) (Vega-Contreras et al.,

2017), Prochilodus nigricans (in the Amazon

Basin), and for the Magdalena-Cauca basin, the

Sinú and Atrato rivers, the P. magdalenae (Moji-

ca et al., 2012; WWF Colombia & AUNAP,

2020). These species are of considerable impor-

tance for artisanal and commercial fishing and

food security in river-side communities.

Although this species holds ecological,

social, and economic significance, there is

still a lack of studies addressing its anatomy,

histology, and histochemistry of the diges-

tive tract, hindering the understanding of

its nutritional and physiological processes

(Vega-Contreras et al., 2017).

The assessment of morphometric param-

eters, such as length and weight, at both whole-

body (Hedayati et al., 2020) and organ-specific

levels (Ramírez-Espitia et al., 2020), along with

the analysis of body indices such as the hepato-

somatic index (Favero et al., 2022) and gonad-

osomatic index (Hasim et al., 2021), emerges as

fundamental tools for determining the anatom-

ical structure and health status of fish. These

methodologies, complemented by histological

and histochemical analyses, provide crucial

information that can contribute to enhancing

species knowledge, as well as aiding in the devel-

opment of conservation strategies (Serrano-

López et al., 2021), detection of contaminants

in their tissues (Liu et al., 2023), and improve-

ment of adaptation to commercial aquaculture

environments (Abdel-Tawwab et al., 2015).

These body indices not only provide infor-

mation about the general condition of the

organism but are also intrinsically linked to the

function and structure of the digestive tract.

The condition factor (K) reflects the overall

nutritional status, which directly indicates the

digestive system’s efficiency and ability to pro-

cess food (Radkhah & Eagderi, 2015). The hep-

atosomatic index (HSI) is particularly relevant,

4Revista de Biología Tropical, ISSN: 2215-2075 Vol. 73: e63718, enero-diciembre 2025 (Publicado Jul. 29, 2025)

as the liver plays a crucial role in nutrient

metabolism and bile production -both essential

components of digestion- (Favero et al., 2022).

The gonadosomatic index (GSI), in turn, helps

to assess how reproductive cycles may influ-

ence both the allocation of energy resources

and the histological structure of the digestive

tract (Hasim et al., 2021). Finally, carcass yield

(CY) provides valuable information about body

composition, which results from the efficiency

of nutrient digestion and absorption (Cabrera-

Páez et al., 2008). The integration of these indi-

ces with histological and histochemical analyses

allows for a more comprehensive understand-

ing of the structure–function relationship of

the gastrointestinal tract (Hedayati et al., 2020;

Ramírez-Espitia et al., 2020), especially in her-

bivorous/detritivorous species such as P. m ag da -

lenae, in which these processes are particularly

specialized (Serrano-López et al., 2021; Liu et

al., 2023; Abdel-Tawwab et al., 2015).

Based on the detritivorous/herbivorous

feeding habits of P. magdalenae, this study

hypothesized that the gastrointestinal tract

would exhibit specific adaptations: an elongated

intestine with an intestinal coefficient greater

than 2.5 (Wilson & Castro, 2010), enhanced

muscular development in the pyloric region

for mechanical processing of detritus (Burns,

2021), and differential distribution of mucins

along the digestive tract (Díaz et al., 2008). The

study predicted neutral mucins would predom-

inate in anterior regions for protection, while

acidic mucins would be more abundant poste-

riorly to facilitate nutrient absorption (Genten

et al., 2008; Vidal et al., 2020). These adapta-

tions would reflect the species’ feeding strategy

through specialized structures for increased

absorption surface area and protection against

abrasion from ingested material (Bowen, 2022;

Wilson & Castro, 2010).

Consequently, this study aims to increase

the fundamental understanding of the gastroin-

testinal tract by delineating the morphological

and histochemical attributes of the digestive

tract wall and the mucous cells within the

mucosal layer, thus advancing comprehension

of the digestive processes in this species.

MATERIAL AND METHODS

All animal handling procedures were con-

ducted in compliance with the standards for

laboratory animal use as outlined by the Com-

mittee on Care and Use of Laboratory Animals

of the National Research Council (National

Academies, USA), eighth edition (Albus, 2012)

and were also authorized by Resolution 0955

of May 27, 2020, issued by AUNAP, granting

research permission to Piscícola San Silvestre.

Fish sampling: A total of 22 adult bocachi-

co individuals (5 males and 17 females) were

collected from their natural habitat in the

Sogamoso River, a tributary branch of the Mag-

dalena River basin, and then transferred to the

Piscícola San Silvestre (PSS). These specimens

were strategically distributed for different anal-

yses: 15 individuals for morphometric mea-

surements and body indices calculation, 5 for

detailed histological and histochemical analy-

ses, and 2 for macroscopic photographic docu-

mentation. All specimens were maintained in

captivity for a maximum of 3 days to prevent

any physiological changes due to artificial feed-

ing and to ensure that data represented their

natural condition. Following a 24-hour quaran-

tine period in filtered water and two salt baths

(20 ppm for 30 seconds), they were relocated to

land ponds at a density of one fish per square

meter. Periodically, Nutrimon® fertilizers (triple

15-15-15, providing nitrogen, phosphorus, and

potassium) were administered to maintain opti-

mal levels of primary productivity (phyto and

zooplankton). Additionally, they were supple-

mented with commercially balanced food for

tilapia containing 34 % CP (1.0 % of biomass),

distributed in two daily rations.

Performance parameters / Morphomet-

ric relationships / Body indices: In the mor-

phometric characterization of P. magdalenae,

15 specimens were employed. Measurements

of total length (TL), standard length (SL),

and intestinal length (IL) in centimeters (cm)

were recorded using a fish-measuring board.

Furthermore, measurements of total weight

Revista de Biología Tropical, ISSN: 2215-2075, Vol. 73: e63718, enero-diciembre 2025 (Publicado Jul. 29, 2025)

(TW), eviscerated weight (EW), and the indi-

vidual weight of the following organs: esopha-

gus, stomach, intestine, liver, and gonads, were

taken in grams (g) using an OHAUS® EB series

scale. Moreover, the liver, gonads, and gills were

collected and weighed individually in order to

calculate the somatic indices and to support

further physiological and histological analyses

Additionally, the photographic registry was

conducted utilizing an EOS Rebel T3i camera

equipped with a Canon EF-S 18-55 mm f/4-5.6

IS STM lens to analyze anatomical features.

Subsequently, based on these measurements,

the following morphometric relationships were

computed following the criteria established by

(Cabrera-Páez et al., 2008; Day et al., 2014).

Furthermore, with the purpose of obtain-

ing an alternative morphometric index to the

intestinal coefficient, one not only dependent

on body lengths, but we also employed the

Zihler index as outlined in the publication by

Day et al. (2014).

The determination of these morphometric

and body indices was carried out to estab-

lish relationships between the general physi-

ological state and the characteristics of the

digestive tract. The condition factor (K) was

calculated to evaluate the nutritional status of

specimens, which provides information on the

digestive system’s effectiveness in obtaining

nutrients (Agbugui & Oniye, 2013). The intes-

tinal coefficient (IC) and Zihler index (ZI) were

determined to quantify specific morphological

adaptations related to herbivorous/detritivo-

rous feeding habits. The hepatosomatic index

(HSI) was included due to the close functional

relationship between the liver and the diges-

tive tract regarding metabolism and nutrient

processing. The gonadosomatic index (GSI)

was calculated to examine the possible influ-

ence of reproductive status on the distribution

of energy resources and its impact on digestive

structure and function. Finally, carcass yield

(CY) was determined to indicate the efficiency

in converting food to body biomass. These

parameters together allow contextualizing the

histological and histochemical findings within

the general physiological state of the studied

specimens. The integration of these indices

with histological observations enables a more

comprehensive analysis of how the digestive

tract structure, including epithelial organiza-

tion, mucin distribution patterns, and tissue

specialization, correlates with overall physi-

ological condition. For instance, variations in

K and HSI can be associated with changes in

mucin production and epithelial characteris-

tics, while IC and ZI provide context for inter-

preting specialized adaptations in the mucosal

layer related to nutrient absorption efficiency,

particularly important in detritivorous species

where extensive surface area and specialized

mucin secretions are critical for optimal diges-

tive function (Pereira et al., 2014).

Histological and histochemical analyses:

For the morphostructural characterization of

the gastrointestinal tract, 22 randomly sampled

individuals were employed. They were anesthe-

tized with 10 ppm eugenol and euthanized by

immersion in an ice bath. Opercula were then

removed, and the left lateral wall dissection was

conducted. Samples of the gastrointestinal tract

were fixed in 10 % buffered formalin at a ratio

of 1:10 for 24 hours.

The tissues underwent dehydration in eth-

anol at sequential concentrations, followed by

cleaning with toluene or an equivalent solvent.

Subsequently, they were embedded in hot par-

affin, molded, and stored at room temperature

as blocks. Sections of 5 to 6 μm thickness were

6Revista de Biología Tropical, ISSN: 2215-2075 Vol. 73: e63718, enero-diciembre 2025 (Publicado Jul. 29, 2025)

obtained using a Leica RM2125 RTS rotary

microtome. Following standard procedures,

these sections were subsequently stained with

hematoxylin and eosin (H&E). The respective

images were captured using an Olympus® CX21

optical microscope and a Basler® ACA5472-

17UC COLOR digital camera to qualitatively

evaluate the digestive system structure (Verma

et al., 2020; Vidal et al., 2020).

To detect neutral mucins, we conducted

Periodic Acid-Schiff (PAS) staining, and for

acidic mucins, Alcian Blue (AB) staining at pH

1.0 and pH 2.5 was utilized (Vidal et al., 2020).

Subsequently, the conjugated AB pH2.5 + PAS

technique was applied to detect the associa-

tion between neutral and acidic mucins. Simi-

larly, the specific detection of epithelial-origin

mucins and acidic mucins in the gastrointes-

tinal tract was achieved through mucicarmine

staining (Kumar & Kiernan, 2010). Finally,

Masson’s trichrome (MT) staining was carried

out for the identification of connective tissue.

Transmission Electron Microscopy

(TEM): Fragments of gastrointestinal tissue

(4-5 mm) were extracted and immersed in 2.5

% buffered glutaraldehyde in PBS (Phosphate

Buffered Saline) for fixation. Following fixa-

tion, the samples were post-fixed in 1 % osmi-

um tetroxide and 3 % uranyl acetate, gradually

dehydrated, and infiltrated with a plastic resin

mixed with acetone (1:1) before embedding in

SPURR resin (Electron Microscopy Sciences,

Fort Washington, PA, USA). The embedded tis-

sue blocks were then sectioned using a Sorvall

MT2-B ultramicrotome. Moreover, semithin

sections (1 µm) were stained with toluidine

blue and examined to identify suitable areas

for ultrathin sectioning (Graham & Orenstein,

2007). These identified areas were cut with a

diamond blade to a thickness of 80 - 100 nm,

resulting in a yellow-gold interference color,

and placed on 200 mesh copper grids. Sub-

sequently, the sections were contrasted with

uranyl acetate and lead citrate, followed by

examination and photography using a JEOL

1400 Plus transmission electron microscope at

the Department of Pathology, Hospital Univer-

sitario Fundación Santa Fe de Bogotá.

Statistical analysis: For the study, descrip-

tive statistics were used to characterize the sam-

ple regarding morphological and zootechnical

parameters, body parameters and indices were

expressed as mean ± STD (standard deviation)

(Moreno et al., 2019).

Histochemical analysis: Histochemical

findings were analyzed based on tissue staining

intensity for each staining technique. Inten-

sity levels were determined through qualitative

inspection, and the staining intensity results

were detailed in a distribution table of acidic

and neutral mucins in the gastrointestinal tract,

similar to previous studies where specific mor-

phological and histochemical adaptations have

been observed in the digestive tract mucosa

of other species such as Brycon amazonicus

(Vidal et al., 2020).

RESULTS

Macroscopic description of gastrointes-

tinal tract morphology: The gastrointestinal

tract of P. magdalenae (Fig. 1) is an elon-

gated digestive system comprising four dis-

tinct regions: the oropharyngeal cavity, the

esophagus, the stomach, and the intestine.

The esophagus is a medium-sized tubular seg-

ment, approximately 2 cm long, characterized

by thick muscular walls. The stomach has a

U-shaped configuration, featuring three dis-

cernible regions: cardiac, fundic, and pyloric

(Fig. 1D). The initial segment corresponds to

a tubular portion posterior to the esophagus,

which undergoes dilation to form a short sac-

like structure (middle-fundic region), followed

by a longer portion with considerably thick

walls and a prominent muscular tunic (muscu-

lar stomach-pyloric region). The intestine of P.

magdalenae is a lengthy, tightly coiled structure

that occupies nearly the entire celomic cav-

ity (Fig. 1B, Fig. 1C), with spirals arranged in

paired fashion and curved in a levorotatory

way (Fig. 1A, Fig. 1D). The anterior intestine

Revista de Biología Tropical, ISSN: 2215-2075, Vol. 73: e63718, enero-diciembre 2025 (Publicado Jul. 29, 2025)

exhibits a visibly larger diameter compared to

the other segments, with thinner walls.

The morphometric measurements collect-

ed are detailed in Table 1, which presents the

maximum, minimum, and average values with

their respective standard deviations for all body

parameters evaluated in the specimens studied.

The body indices, including the condi-

tion factor K, intestinal coefficient (IC), Zihler

index (ZI), gonadosomatic index (GSI), hepa-

tosomatic index (HSI), and carcass yield (CY),

were calculated based on the data obtained

from Table 1. Table 2 presents a summary of the

estimated body indices.

Fig. 1. A. Lateral view of the coelomic cavity of adult P. magdalenae showing the esophagus (Es), cardiac stomach region (CS),

fundic stomach region (FS), pyloric stomach region (PS), pyloric ceca (PC), anterior intestine (AI), middle intestine (MI),

posterior intestine (PI), and rectum (R). B. Anatomy of the digestive tract and cardiorespiratory organs of the oropharyngeal

cavity displaying rectum (R), posterior intestine (PI), middle intestine (MI), pyloric ceca (PC), esophagus (Es), liver (L), gill

arch (GA), and gill rakers (GR). C. Detailed anatomy of the gastrointestinal tract showing posterior intestine (PI), middle

intestine (MI), cardiac stomach region (CS), esophagus (Es), fundic stomach region (FS), pyloric stomach region (PS),

anterior intestine (AI), and pyloric ceca (PC).

8Revista de Biología Tropical, ISSN: 2215-2075 Vol. 73: e63718, enero-diciembre 2025 (Publicado Jul. 29, 2025)

Microscopic description of histology and

histochemistry of the esophagus: The aver-

age length of the esophagus in P. magdalenae

was 1.26 ± 0.95 cm. In the anterior section, a

mucosa covered initially by a non-keratinized

stratified squamous epithelium was observed

(Fig. 2B). However, as it progresses towards

the middle region, digitiform folds protruding

into the lumen are observed, lined by a simple

columnar epithelium (ec), and again, stratified

squamous epithelium is observed in the area

near the stomach. Throughout the length of

this organ, abundant mucous acinar glands are

present. Below the basement membrane of the

epithelium, a thick layer of loose connective

tissue (tcl) was observed, corresponding to the

lamina propria-submucosa (lp-sm), as no mus-

cular layer of the mucosa was observed; there-

fore, there is no separation between the lamina

propria and the submucosa (sm). Externally,

the muscular layer, consisting of two layers of

skeletal striated muscle, is observed, with the

inner circular layer evident in the first third of

the esophagus and the outer longitudinal layer

which is thicker and more noticeable. Covering

the muscular layer, an adventitia is observed as

a thin layer of loose connective tissue.

Transmission electron microscopy (TEM)

(Fig. 3) enabled a more detailed observation

of the cellular morphology of the superficial

layer of the epithelium, comprising thin cells

with elongated central nuclei characteristic of

a stratified squamous epithelium. These cells

exhibit higher electron density than the goblet

cells in the same tissue, and microvilli can also

be observed on the apical surface of these cells

(Fig. 3A, Fig. 3B). Additionally, large goblet

cells containing mucin granules were observed,

with some of these cells seen releasing part of

these mucins into the lumen.

Through immunohistochemical staining,

mucous acinar glands, some of them slightly

oval-shaped, were observed (Fig. 2B, Fig. 2C).

Staining with PAS, mucicarmine, AB pH 1.0,

Table 1

Body weights of 15 individuals of bocachico (P. magdalenae).

Maximum (g) Minimum (g) Average (g)

Total Weight 440.0 221.0 291.77 ± 65.04

Eviscerated Weight 356.0 179.0 249.24 ± 58.06

Intestine 7.34 0.05 4.81 ± 1.31

GIT 32.02 2.54 12.02 ± 8.01

Liver 3.18 1.06 2.20 ± 0.47

Gonads 4.33 1.25 2.39 ± 1.02

Bladder 3.05 0.98 1.79 ± 0.60

Stomach 4.12 1.32 2.88 ± 0.55

Esophagus 1.26 0.11 0.60 ± 0.27

Table 2

Body indices of 15 individuals of bocachico (P. magdalenae).

Index Minium Maximum Average Deviation

K 0.84 1.18 1.02 0.09

IC 2.43 3.16 2.87 0.19

ZI 1.10 1.49 1.33 0.10

GSI 0.50 1.61 0.81 0.29

HIS 0.43 0.92 0.75 0.07

CY 65.28 94.80 85.20 7.78

Condition factor (K), intestinal coefficient (IC), Zihler index (ZI), hepatosomatic index (HSI), carcass yield (CY), and

gonadosomatic index (GSI).

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Fig. 2. Histological Organization and Dynamics of Mucins in the Middle Esophagus of P. magdalenae. A. Detailed anatomy

of the esophagus showing esophagus (ES) and cardiac stomach region (CS). The esophageal epithelium appears as a simple

columnar epithelium (EC) with scattered goblet cells (GC), overlaying the basement membrane and the lamina propria (LP)

composed of loose connective tissue (LCT). The tissue also contains acinar glands (AG) visible in the submucosal region, and

a layer of smooth muscle (SM) underlying all structures. Goblet cells exhibit the presence of neutral mucins (marked with *)

and acidic mucins (marked with +), as observed in staining descriptions: B. Simple columnar epithelium with scattered goblet

cells, accompanied by numerous longitudinal folds towards the lumen, increasing the absorptive surface. Thin lamina propria

and thick muscular layer (H&E 100x). C. Simple columnar epithelium with goblet cells, showing a broad lamina propria with

abundant, blue-stained loose connective tissue (MT 40x). D. Numerous goblet cells with intensely magenta-stained apical

portions, indicating a high presence of neutral mucins (PAS 40x). E. Goblet cells with intensely blue-stained apical portions,

revealing significant amounts of acidic mucins (Mucicarmine 40x). F. Intense blue staining in goblet cells, demonstrating a

high concentration of sulphated and carboxylated acidic mucins (AB pH 2.5 40x). G. Slight blue staining in some goblet cells,

indicating a low presence of sulphated acidic mucins (AB pH 1.0). H. Combination allowing the distinction of goblet cells

with neutral mucins (magenta) and sulfated/carboxylated acidic mucins (blue) (PAS/AB pH 2.5).

and AB pH 2.5 facilitated the visualization

of neutral mucins (Fig. 2D, Fig. 2E, Fig. 2F,

Fig. 2G). Staining for sulfated and carboxyl-

ated acidic mucins was exclusively present in

mucus cells, while labeling for neutral mucins

was observed in both mucus and epithelial

columnar cells. Co-localization of acidic and

neutral mucins was observed with PAS/AB

pH 2.5 staining (Fig. 3H), with acidic mucins

predominating.

Histology and histochemistry of the

stomach: The initial segment displayed a wide

mucosal layer characterized by numerous short

folds projecting towards the lumen. It was lined

by a simple columnar epithelium (SCE) and

featured abundant tubular gastric glands con-

taining oxyntic peptic cells. Sparse loose con-

nective tissue was observed between the glands,

corresponding to the lamina propria (Fig. 4B,

Fig. 4C). A thin layer of smooth muscle, cor-

responding to the muscularis mucosae, was

evident towards the end of the mucosal layer.

Additionally, a thin layer of loose connec-

tive tissue (submucosal layer) was observed

externally to the mucosal layer. Below this, two

10 Revista de Biología Tropical, ISSN: 2215-2075 Vol. 73: e63718, enero-diciembre 2025 (Publicado Jul. 29, 2025)

layers of skeletal striated muscle, consisting

of an inner circular layer and an outer lon-

gitudinal layer (muscular layer), were noted.

Moreover, an outer layer of loose connective

tissue, known as the serosal layer, was observed

externally (Fig. 4C).

Furthermore, macroscopically, the cen-

tral section appears as a short sac-like struc-

ture (Fig. 4A), representing a transitional zone

where the mucosal layer thins out. It is lined by

a simple columnar epithelium, under which lies

a thin layer of loose connective tissue (lamina

propria), accompanied by blood vessels, some

mucosal folds, and gastric pits. This mucosa is

thinner than the preceding one, attributable to

the absence of glands with oxyntic peptic cells

cells. Posterior to the lamina propria, the muco-

sal layer exhibited a thin layer of smooth mus-

cle corresponding to the muscularis mucosae.

Beneath the mucosal layer, a layer of connective

tissue corresponding to the submucosal layer,

composed of loose connective tissue (LCT),

was observed, with the muscular layer located

externally to it. In this segment of the stomach,

the muscular layer displays up to three layers of

smooth muscle: an inner oblique layer, a middle

circular layer, and an outer longitudinal layer.

Beyond the muscular layer, a thin layer of loose

connective tissue representing the serosal layer

was identified.

In the observations made using TEM

(Fig. 5), epithelial cells with abundant gran-

ules exhibiting pronounced electron density

towards the apical zone were observed. Addi-

tionally, intraepithelial cells, possibly lympho-

cytes migrating through the epithelium, were

noted within the epithelial layer. Furthermore,

invaginations were observed, contributing to

the epithelium’s lobulated appearance.

The third gastric segment corresponded to

a region with a thick muscular wall (Fig. 6A),

consisting of a mucous membrane lined by a

simple columnar epithelium and abundant sim-

ple straight tubular glands. Beneath the epithe-

lium, there was a thin layer of loose connective

tissue, either the lamina propria or submucosa,

as no muscular layer of the mucosa separating

the lamina propria from the submucosa was

observed. Externally to this, there was a very

thick layer of smooth muscle corresponding

to the muscularis externa, which was the wid-

est layer in this segment of the digestive tract.

External to the muscularis externa, there was

a thin layer of connective tissue corresponding

Fig. 3. Transmission electron microscopy (TEM) of the esophagus of P. magdalenae reveals: A. Flat epithelial cells (FEC) with

microvilli (MV), goblet cells (GC), and mucin granules (MG) inside the goblet cell, with (M) representing mucin release into

the lumen. The nucleus is labeled as (N). B. Mucin release (M) and mucin granules (MG) in the cytoplasmic region of the

cell are observed in greater detail, with (N) indicating the nucleus.

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to the serous membrane. Mucin dynamics in all

stomach regions exclusively corresponded to

neutral mucins.

Histology and histochemistry of the intes-

tine: The average length of the intestine in P.

magdalenae was 88.18 ± 7.35 cm. Histologically,

four segments can be distinguished: anterior,

glandular, middle, and posterior. All segments

exhibit a mucosal tunica with digitiform folds

projecting into the lumen, with shorter folds

in the posterior segment. The mucosal tunica

in all segments is lined by a simple columnar

epithelium, composed of enterocytes and goblet

cells. In the posterior segment, there is a reduc-

tion in the number of columnar cells and an

increase in mucus-producing cells. Beneath the

epithelium in the anterior, middle, and poste-

rior segments, a layer of loose connective tissue

corresponding to the lamina propria-submu-

cosa was observed, as no muscularis mucosae

separating these layers from the connective

tissue was found. Additionally, externally, two

layers of smooth muscle -the inner circular and

the outer longitudinal- were observed, forming

the muscular tunica.

External to the muscular tunica, there

is a thin layer of loose connective tissue

Fig. 4. Histological organization and mucin dynamics in the stomach of P. magdalenae. A. Detailed anatomy of the

gastrointestinal tract showing posterior intestine (PI), middle intestine (MI), cardiac stomach region (CS), esophagus (Es),

fundic stomach region (FS), pyloric stomach region (PS), anterior intestine (AI), and pyloric ceca (PC). B. Section showing

a simple columnar epithelium (SCE) and numerous tubular gastric glands (TGG), H&E staining, 100x. C. Section displaying

the lamina propria (LP) composed of loose connective tissue (LCT), MT staining, 40x. D. Section showing the presence of

neutral mucins (*), PAS staining, 40x. E. Section showing complete absence of acidic mucins, Mucicarmine staining, 40x.

F. Section displaying complete absence of acidic mucins, AB pH 2.5 staining, 40x. G. Section showing complete absence

of acidic mucins, AB pH 1.0 staining, 40x. H. Section displaying the presence of neutral mucins (*) and absence of acidic

mucins, PAS/AB pH 2.5 staining, 40x. Abbreviations: posterior intestine (PI), middle intestine (MI), cardiac stomach region

(CS), esophagus (ES), fundic stomach region (FS), pyloric stomach region (PS), anterior intestine (AI), pyloric ceca (PC),

simple columnar epithelium (SCE), tubular gastric glands (TGG), lamina propria (LP), loose connective tissue (LCT),

Hematoxylin and Eosin (H&E), Masson’s Trichrome (MT), Periodic Acid-Schiff (PAS), and Alcian Blue (AB).

12 Revista de Biología Tropical, ISSN: 2215-2075 Vol. 73: e63718, enero-diciembre 2025 (Publicado Jul. 29, 2025)

corresponding to the serosal tunica. The ante-

rior segment of the intestine opens into multi-

ple finger-like structures that terminate blindly,

corresponding to the pyloric caeca with a muco-

sal tunica exhibiting digitiform folds projecting

into the lumen, lined by a simple columnar

epithelium (Fig. 7B, Fig. 7C). Beneath this layer

lies a thin layer of loose connective tissue, the

lamina propria, and externally, the muscular

tunica with two layers of smooth muscle, the

inner circular and the outer longitudinal, and

peripherally, a thin layer of loose connective tis-

sue, the serosal tunica. Posterior to the anterior

segment of the intestine in this species, there is

an area where the mucosal tunica, unlike the

other regions of the intestine, does present a

muscular layer of the mucosa, separating the

lamina propria from the submucosal tunica,

with abundant acinar glands producing mucus

in the latter. Beyond the submucosal tunica,

the muscular and serosal tunicas are similar to

those reported in the other segments.

When evaluating the ultrastructure of

the intestinal epithelium across various seg-

ments using Transmission Electron Micros-

copy (TEM) (Fig. 8), a greater abundance

of enterocytes is evident in the anterior and

middle regions. The presence of a simple

columnar epithelium is confirmed, character-

ized by abundant microvilli (brush border)

prominently displayed on the apical surface of

the epithelial cells.

Histochemical analysis revealed the pres-

ence of neutral and sulfated acidic mucins, as

well as carboxylated acidic mucins, with mod-

erate intensity in the anterior and middle seg-

ments, attributable to the low number of mucus

cells in these areas. A slight predominance of

neutral mucins was observed in the anterior

portion, whereas in the middle and posterior

segments, there was a predominance of acidic

mucins, primarily carboxylated (Fig. 9). Table 3

summarizes the observed dynamics of acidic

and neutral mucins in the digestive tract of P.

magdalenae, also emphasizing crucial histologi-

cal aspects of this species regarding the types of

cells involved in mucin secretion.

DISCUSSION

The Fulton condition factor (K) is an

index utilized to assess the impact of diverse

Fig. 5. Ultrastructural analysis of the gastric cardiac region epithelium of P. magdalenae. Transmission electron micrographs

of the gastric cardiac region showing: A. Simple columnar epithelium (SCE) with intraepithelial cells (*), epithelial

invagination (h), and nuclei (N), scale bar: 10 μm. B. Higher magnification view of the epithelium displaying secretory

granules and an intraepithelial cell (*), with epithelial surface indicated by arrow (h), scale bar: 5 μm.

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environmental conditions on fish health, par-

ticularly in relation to their nutritional status. K

values exceeding 1.0 are indicative of favorable

environmental and feeding conditions condu-

cive to fish development, whereas values below

1.0 suggest less favorable conditions (Radkhah

& Eagderi, 2015). In this study, the K factor was

computed to evaluate the well-being of captured

individuals, yielding an average of 1.02 ± 0.09.

This value is notably lower than those reported

by Vega-Contreras et al. (2017) at three distinct

sites along the Sogamoso River, where average

values of 1.21 ± 0.03, 1.24 ± 0.06, and 1.20 ±

0.05 were documented. Contreras-Almazo et

al. (2019) contend that the heightened presence

of anthropogenic debris may have precipitated

the premature sexual maturation of P. m a g d a -

lenae, a phenomenon previously observed in

P. lineatus and associated with K factor values

surpassing 1.0 (Alonso et al., 2015; Cañas-Alva

et al., 2020; Cifuentes et al., 2012). Poten-

tial explanations for the variance in K values

Fig. 6. Histological and dynamic organization of mucins in the pyloric region of the stomach of P. magdalenae. A.

Macroscopic view showing the presence of a prominent muscular wall (PM) and a mucosal tunica (MT), which delineate the

fundic stomach (FS) and its connection with the pyloric caeca (PC). Scale bar: 10 mm. B. Microscopic view of the mucosa

lined with a simple columnar epithelium (SCE) and simple straight tubular glands (STG). H&E staining, 10x. Scale bar: 1

μm. C. The detail shows a simple columnar epithelium (SCE), simple straight tubular glands (STG), and the presence of loose

connective tissue (LTC), specifically the lamina propria (LP). MT staining, 10x. D. The section shows simple straight tubular

glands (STG) and the presence of neutral mucins (indicated by *). PAS staining, 40x. E. Section displaying simple columnar

epithelium (SCE) and simple straight tubular glands (STG) with complete absence of acidic mucins. Mucicarmine staining,

10x. Scale bar: 1 μm. F. Section showing simple straight tubular glands (STG) with complete absence of acidic mucins. AB

pH 2.5 staining, 40x. Scale bar: 20 μm. G. Section displaying simple straight tubular glands (STG) with complete absence

of acidic mucins. AB pH 1.0 staining, 40x. Scale bar: 20 μm. H. Section showing simple straight tubular glands (STG) with

the presence of neutral mucins (indicated by *) and absence of acidic mucins. PAS/AB pH 2.5 staining, 40x. Scale bar: 20

μm. Abbreviations: prominent muscular wall (PM), mucosal tunica (MT), fundic stomach (FS), pyloric caeca (PC), simple

columnar epithelium (SCE), simple straight tubular glands (STG), loose connective tissue (LTC), lamina propria (LP),

Hematoxylin and Eosin (H&E), Masson’s Trichrome (MT), Periodic Acid-Schiff (PAS), Alcian Blue (AB).

14 Revista de Biología Tropical, ISSN: 2215-2075 Vol. 73: e63718, enero-diciembre 2025 (Publicado Jul. 29, 2025)

between this study’s findings for P. magdalenae

and those of Contreras-Almazo et al. (2019),

encompass factors such as the fish’s residence

in ponds supplied with tributary water that has

undergone prior filtration, seasonal disparities,

and the controlled feeding regimen employed

within the San Silvestre fishery (Ragheb, 2023).

The morphometric indices analyzed in this

study provide valuable context for interpreting

the histological and histochemical findings in

P. magdalenae’s digestive tract. The condition

factor (K) of 1.02 ± 0.09 indicates adequate but

not optimal nutritional status, which directly

influences the efficiency of the digestive system

and potentially affects mucin production pat-

terns observed in the different GIT segments

(Kroon et al., 2017). This K value, while suf-

ficient for normal physiological function, is

lower than previously reported values for this

species (Vega-Contreras et al., 2017), suggest-

ing potential differences in digestive efficiency

that align with our histochemical observations

Fig. 7. Histology of the intestine of P. magdalenae. The histological organization of the intestine of P. magdalenae is

characterized by: A. Anterior intestine section showing a simple columnar epithelium (JSCE) with enterocytes (E) and the

muscular tunica (MT), H&E staining, 40x. B. Middle intestine section showing a simple columnar epithelium (JSCE) with

enterocytes (E) and the muscular tunica (MT), H&E staining, 40x. C. Posterior intestine section showing a simple columnar

epithelium (JSCE) with enterocytes (E) and the muscular tunica (MT), H&E staining, 40x. D. Anterior intestine section

revealing a simple columnar epithelium (JSCE) with enterocytes (E), the lamina propria (LP), and the muscular tunica

(MT), MT staining, 40x. E. Middle intestine section displaying a simple columnar epithelium (JSCE) with enterocytes (E),

lamina propria (LP), mucosa/muscularis (MM), and acinar glands-mucin (AG-M), MT staining, 40x. F. Posterior intestine

section displaying a simple columnar epithelium (JSCE) with goblet cells (GC), enterocytes (E), and the lamina propria (LP),

MT staining, 40x. Scale bars: 20 μm. Abbreviations: simple columnar epithelium (JSCE), enterocytes (E), goblet cells (GC),

lamina propria (LP), muscular tunica (MT), mucosa/muscularis (MM), acinar glands-mucin (AG-M), Hematoxylin and

Eosin (H&E), Masson’s Trichrome (MT).

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of mucin distribution. The average hepatoso-

matic index (HSI) of the fish in the study was

0.75 ± 0.07. Considering that HSI values below

1.0 are associated with low energy reserves,

which may be linked to situations such as

reduced food availability, environmental stress,

presence of contaminants, or the onset of the

reproductive stage (Cabrera Páez et al., 2008),

the observed low energy reserves in the fish

suggest potential preparation for the reproduc-

tive stage or conditions in captivity that differ

from their natural environment. Factors such

as food quality and availability, as well as physi-

cochemical water conditions, may contribute

to these differences (Contreras-Almazo et al.,

2019; Mancera-Rodríguez et al., 2016), because

in natural environments, food availability is

often linked to the presence of organic matter

and macrophytes (Mojica et al., 2012). The

hepatosomatic index (HSI) of 0.75 ± 0.07 is par-

ticularly relevant for digestive function inter-

pretation, as the liver plays a crucial role in bile

production -essential for lipid digestion- and

detoxification of compounds from the diges-

tive tract (Figueiredo-Silva et al., 2013). This

relatively low HSI value suggests limited energy

reserves, which may influence the specialized

mucin secretion patterns we observed, particu-

larly the predominance of carboxylated acidic

mucins in the posterior intestinal segments that

Fig. 8. Epithelial ultrastructure of the intestine of P. magdalenae. Transmission electron micrographs showing the

ultrastructural organization of the intestinal epithelium of P. magdalenae as follows: A. Anterior intestine showing goblet

cells (GC) with secretory granules, scale bar: 5 μm. B. Middle intestine displaying goblet cells (GC), the brush border of the

intestinal epithelium (BB), and enterocyte nuclei (N), scale bar: 10 μm. C. Middle intestine section showing the brush border

(BB) with well-developed microvilli (MV), scale bar: 5 μm. D. Posterior intestine displaying microvilli (MV), goblet cells

(GC), and nuclei (N), scale bar: 5 μm. Abbreviations: GC: goblet cells; MV: microvilli; N: nucleus; BB: brush border.

16 Revista de Biología Tropical, ISSN: 2215-2075 Vol. 73: e63718, enero-diciembre 2025 (Publicado Jul. 29, 2025)

enhance nutrient absorption efficiency (Cabal-

lero et al., 2004).

The gonadosomatic index (GSI) of 0.81 ±

0.29 indicates potential reproductive activity

that affects nutrient allocation within the organ-

ism. During reproductive periods, resources

may be diverted from digestive processes to

gonadal development, potentially altering

mucin production patterns and the histologi-

cal structure of the digestive tract (Zaldúa &

Naya, 2014). This relationship between repro-

ductive status and digestive histology requires

Fig. 9. Mucin histochemistry of the intestine of P. magdalenae. The histochemical characterization of mucins in the intestine

of P. magdalenae is presented as follows: A. Anterior intestine showing the presence of neutral mucins (*) in goblet cells,

PAS staining, 40x. B. Anterior intestine displaying epithelial acidic mucins (<) in goblet cells, Mucicarmine staining, 40x. C.

Anterior intestine showing carboxylated acidic mucins (+), AB pH 2.5 staining, 40x. D. Anterior intestine showing sulfated

acidic mucins (@), AB pH 1.0 staining, 40x. E. Anterior intestine displaying colocalization of acidic and neutral mucins (!),

PAS/AB pH 2.5 staining, 40x. F. Middle intestine showing the presence of neutral mucins () in goblet cells, PAS staining, 40x.

G. Middle intestine displaying epithelial acidic mucins (<) in goblet cells, Mucicarmine staining, 40x. H. Middle intestine

showing carboxylated acidic mucins (+), AB pH 2.5 staining, 40x. I. Middle intestine showing sulfated acidic mucins (@), AB

pH 1.0 staining, 40x. J. Middle intestine displaying colocalization of acidic and neutral mucins (!), PAS/AB pH 2.5 staining,

40x. K. Posterior intestine showing the presence of neutral mucins (*) in goblet cells, PAS staining, 40x. L. Posterior intestine

displaying epithelial acidic mucins (<) in goblet cells, Mucicarmine staining, 40x. M. Posterior intestine showing carboxylated

acidic mucins (+), AB pH 2.5 staining, 40x. N. Posterior intestine showing sulfated acidic mucins (@), AB pH 1.0 staining,

40x. O. Posterior intestine displaying colocalization of acidic and neutral mucins (!), PAS/AB pH 2.5 staining, 40x. Scale bars:

20 μm. Abbreviations: Periodic Acid-Schiff (PAS), and Alcian Blue (AB).

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Table 3

Dynamic of acidic and neutral mucins in the digestive tract of P. magdalenae.

Staining Type of mucins Esophagus –

Cranial portion

Esophagus –

Middle portion

Esophagus –

hind portion

Stomach –

Antral

region

Stomach –

Fundic

region

Stomach

– Pyloric

region

Foregut Midgut Hindgut

PAS Neutral mucins +++ GC

and epithelium

+++

GC and epithelium

+++

GC and epithelium

+++

epithelium

+++

epithelium

+++

Epithelium

Few GC

+ ++

pH 1.0

Sulfated acidic

mucins

+++ GC +++ GC +++ GC - - - ++

Few CC

++ ++

pH 2.5

Carboxylated

acidic mucins

+++ GC +++ GC +++ GC - - - ++

Few CC

++ ++

PAS/AB

pH 2.5

Acid and neutral

mucins

+++ predominance

of acidic mucins

and slight neutral

marking

+++ predominance of

acidic mucins and slight

neutral marking. EC

exclusively neutral dialing

+++ predominance of

acidic mucins and slight

neutral marking. EC

exclusively neutral dialing

+++

exclusively

neutral

+++

exclusively

neutral

+++

exclusively

neutral

Few GCs,

both acidic and

neutral (slightly

predominant)

Predominance

of carboxylated

acidic mucins

Predominance of

carboxylated acidic

mucins

Mucicarmin Epithelial acidic

mucins

++ GC ++ GC ++ GC - - - + + ++

The intensity of staining was assessed as (–) absence, (+) mild, (++) moderate and (+++) intense, (GC) goblet cells.

18 Revista de Biología Tropical, ISSN: 2215-2075 Vol. 73: e63718, enero-diciembre 2025 (Publicado Jul. 29, 2025)

further investigation in P. magdalenae. Finally,

the carcass yield (CY) of 85.20 ± 7.78 reflects

overall body composition and provides indi-

rect evidence of digestive efficiency, as higher

values indicate better conversion of ingested

nutrients into body mass (Salem et al., 2018).

The CY values observed support our findings

regarding the specialized intestinal structure

with its elongated form (intestinal coefficient of

2.87 ± 0.19) and differential mucin distribution

that maximizes nutrient extraction from detri-

tal material (Jiao et al., 2023). Together, these

indices provide a comprehensive physiological

context that enhances our understanding of the

structure-function relationship in the digestive

tract of this detritivorous species.

Further analysis of the K factor (1.02 ±

0.09) and HSI (0.75 ± 0.07) values requires con-

sideration of multiple environmental and phys-

iological variables that could influence these

indices (Mozsár et al., 2015). The controlled

food availability in the San Silvestre fishery,

with supplementary balanced feed (34 % CP)

administered at 1.0 % of biomass, differs sig-

nificantly from the natural feeding conditions

of wild P. magdalenae populations that pri-

marily consume detritus and periphyton. This

difference in diet composition and availability

could explain the lower K values compared to

those reported by Vega-Contreras et al. (2017).

Similarly, the relatively low HSI values observed

may reflect an adaptation to captive conditions

with controlled feeding regimens rather than

energy deficiency per se. Population density

also plays a crucial role in modulating these

indices, as the experimental density of one fish

per square meter represents a significantly less

crowded environment than natural conditions,

potentially reducing competition stress but also

altering feeding behavior (Abdel-Tawwab et

al., 2015). Seasonal variables and reproductive

cycles further interact with these indices, as

demonstrated by Cifuentes et al. (2012) and

Cañas-Alva et al. (2020) in related species.

The relationship between reproductive status

and both K and HSI is particularly relevant,

as energy reserves are redistributed during

gonadal development. The GSI values (0.81

± 0.29) in our specimens suggest that some

individuals were in early gonadal development

stages, which may have influenced both hepatic

reserves and overall body condition (Nunes

et al., 2011). These interrelated factors -food

availability, population density, and reproduc-

tive status -collectively impact the digestive

physiology observed in our histological and

histochemical analyses, particularly the mucin

distribution patterns that optimize nutrient

absorption under specific energetic demands.

On the other hand, Karachle and Ster-

giou (2010) argue that a strong correlation

exists between intestinal length and the feeding

habits of fish species. Typically, carnivorous

species possess shorter intestines, while omni-

vores, herbivores, and detritivores tend to have

longer ones. Similarly, morphometric param-

eters concerning intestinal length, such as the

intestinal coefficient (CI) and Zihler’s index,

are frequently utilized to deduce the feeding

behaviors of fish species. In this study, the esti-

mated values for both indices were 2.87 ± 0.19

and 1.33 ± 0.10, respectively. The CI aligns with

what is reported for omnivorous species but is

notably close to the value of 3.0 specified for

detritivores (Karachle & Stergiou, 2010). Con-

cerning Zihler’s index (IZ), which establishes a

morphometric relationship between intestinal

length and fish body mass, it delineates the

following ranges: pure herbivores (2.0 to 2.1),

omnivores with a herbivorous inclination (2.1

to 2.9), omnivores with an animal inclination

(2.9 to 3.7), carnivores with a preference for

decapods and fish (3.7 to 4.0), and carnivores

with a preference for fish and cephalopods (4.0

to 4.5) (Day et al., 2014; Karachle & Stergiou,

2010). The estimated IZ value for P. magdale-

nae falls below 2, indicating a classification as a

pure herbivore within this system; nevertheless,

there is an insufficient number of reports for

detritivore classification and IZ values tailored

to this feeding habit.

The morphology and observed distribu-

tion of the gastrointestinal system organs in

P. magdalenae align with the compiled infor-

mation on the generalities and specificities of

digestive structures in teleosts (Genten et al.,

Revista de Biología Tropical, ISSN: 2215-2075, Vol. 73: e63718, enero-diciembre 2025 (Publicado Jul. 29, 2025)

2008; Wilson & Castro, 2010). P. magdalenae

possesses a lengthy digestive tract comprised of

four well-defined regions (oropharyngeal cav-

ity, esophagus, stomach, and intestine) similar

a B. amazonicus (Vidal et al., 2020), Schizodon

knerii (dos Santos et al., 2015), and other,

more distantly related taxa such as Tilapia

sparrmani (Okuthe & Bhomela, 2021), and

Oreochromis niloticus and Clarias gariepinus

(Awaad et al., 2014).

The secretion of neutral mucins in the

mucosa of the esophageal tract facilitates food

passage while providing protection against

abrasion damage due to the absence of saliva

and a masticatory process, as seen in other

vertebrates. Additionally, acidic mucins offer

protection against the establishment of poten-

tial pathogens in the esophageal lumen, and

contribute to food degradation before it enters

the stomach (Awaad et al., 2014; Cornick et

al., 2015; Firmino et al., 2020; Marchetti et al.,

2006; Phrompanya et al., 2019).

Glycoproteins, particularly mucins secret-

ed by goblet cells throughout the digestive tract

of P. magdalenae, play crucial roles in epithe-

lial protection against abrasion from organic

matter (Nachi et al., 1998). The abundance

of acinar glands in the esophagus that pro-

duce both acidic and neutral mucins forms a

protective barrier that shields the epithelium

from mechanical damage caused by ingested

sediments, common in detritivorous feeding

(Cyrino et al., 2008). This mucin layer acts as

a renewable physical barrier preventing direct

contact between abrasive sediment particles

and delicate epithelial surfaces, reducing cel-

lular damage and irritation, especially in the

pyloric region which functions similarly to a

gizzard in the mechanical processing of detritos

(Baldisserotto et al., 2019).

The differential distribution of mucins

facilitates both protection and efficient diges-

tion of particulate organic matter, with neu-

tral mucins predominating in anterior regions

(esophagus and anterior intestine) and acid-

ic mucins in middle and posterior intestinal

regions Acidic mucins, especially abundant in

the middle and posterior intestine, help trap

organic particles from detritus, increasing con-

tact time with digestive enzymes and enhanc-

ing nutrient extraction efficiency (Palladino

et al., 2023). Additionally, mucins may create

microenvironments that optimize the function

of digestive enzymes needed to break down

organic matter in detritus, which often contains

complex carbohydrates and other nutrients

embedded in difficult-to-digest matrices, rep-

resenting a key evolutionary adaptation that

allows P. magdalenae to efficiently exploit detri-

tal food resources (Ortiz-Ruiz et al., 2024).

In terms of ultrastructure, similarities are

observed in the arrangement of acinar glands,

the presence of microvilli towards the luminal

part, interruption of the continuity of squa-

mous epithelial tissue, and abundant granular

content of mucins when compared with spe-

cies such as Saliminus affinis (Atencio-García

et al., 2008), Sparus aurata (Abumandour &

El-Bakary, 2018) and Anguilla anguilla (Knut-

sen et al., 2021). For the genus Prochilodus,

there are no studies supporting findings at the

esophageal structure level. The stomach of P.

magdalenae, due to its macroscopic structure

and U-shaped disposition, could be classified

as fundic type, as reported by de Moraes et

al. (1997). In teleosts, the diversity of stomach

shapes and structures is vast, even absent in

families such as Cyprinidae, Labridae, Gobiidae,

Scaridae, Cyprinodontidae, and some members

of Poeciliidae (Genten et al., 2008; Ray & Ringø,

2014). The celomic organization of P. m a g d a -

lenae concerning the coilings and folds of the

gastrointestinal tract (esophagus, stomach, and

intestine) shows a significant similarity to what

has been reported for Characids (Alonso et

al., 2015). Similarly, for species like Astyanax

endy, Astyanax rutilus, Cheirodon interruptus,

Aphyocharax anisitsi, Gymnocorymbus ternetzi,

Bryconamericus thomasi, Markiana nigripin-

nis, Characidium borellii; also belonging to the

characid group (Alonso et al., 2015). The same

report also identified histological organization

patterns for the entire stomach structure very

similar to what was observed in this study for P.

magdalenae, including structural peculiarities

regarding the thickness of the muscular layer

20 Revista de Biología Tropical, ISSN: 2215-2075 Vol. 73: e63718, enero-diciembre 2025 (Publicado Jul. 29, 2025)

of the pyloric stomach (Fig. 6A) and coincid-

ing with what has been reported for P. lineatus

(Bowen, 2022).

The muscular tissue that lines the pyloric

stomach region serves functions similar to a

gizzard in herbivorous and detritivorous fish,

aiding in the degradation of plant fibers and

other non-living organic matter through the

mechanical action of muscular movements

(Burns, 2021; de Moraes et al., 1997). In the case

of P. lineatus, the retention of mineral particles

(sand) during the digestive process in the pylor-

ic region has been confirmed, facilitating the

grinding process of detritus ingested by the fish.

This retention and selection of particles suggest

muscular control that allows for the retention

and mixing of food during this process, fol-

lowed by its subsequent movement towards the

intestine (Bowen, 2022; Guzmán-Beltran et al.,

2013; Ramírez-Espitia et al., 2020). Despite the

similarities between the gastrointestinal tracts

of P. magdalenae and P. lineatus, studies are

needed to provide precise information on how

mechanical, enzymatic, and nutrient assimila-

tion processes occur in P. magdalenae.

The mucins detected in the different

regions of P. magdalenae’s stomach in this

study exclusively corresponded to neutral

mucins (Fig. 5, Fig. 6, and Table 3), as similarly

observed in other species such as B. amazonicus

(Vidal et al., 2020), Hyphessobrycon anisitsi, and

G. ternetzi (Leiv-Leknes, 2011; Leiv-Leknes,

2015). Neutral mucins play a significant role

in protecting the intestinal mucosa against

mechanical damage from elements that are dif-

ficult to digest, as is the case with herbivores

and detritivores, while also shielding the tis-

sue from damage caused by hydrochloric acid

secreted by oxynticopeptic cells and the enzy-

matic activity of the stomach (Domeneghini et

al., 2005; Genten et al., 2008). In this study, the

presence of this cell type was confirmed in the

cardiac region of P. magdalenae’s stomach.

With TEM, the epithelial tissue in the

pyloric region of P. magdalenae’s stomach con-

firmed the structure of simple columnar epi-

thelium devoid of acinar glands and a marked

granularity corresponding to the neutral mucins

detected with Periodic Acid-Schiff (PAS) stain-

ing. This morphology is consistent with species

from the characid group, including A. endy,

A. rutilus, C. interruptus, A. anisitsi, G. ternetzi,

B. thomasi, M. nigripinnis, and C. borellii (Alon-

so et al., 2015). While the aforementioned study

analyzed morphology using optical micros-

copy, it provides a detailed description that

aligns with the findings revealed by TEM in this

study. Studies in other taxonomic groups, such

as Siluriformes, including the species C. gariepi-

nus (Moawad et al., 2016), Salmoniformes with

Brachymystax tsinlingensis (Xiong et al., 2019)

and Perciriformes with Perca fluviatilis (Noail-

lac-Depeyre & Gas, 1982), report intracellular

granularities related to the secretory activity of

the tissue in the stomach epithelium observed

with TEM. At the intraepithelial level, electron-

dense cells with large nuclei and little cytoplasm

corresponding to migratory lymphocytes were

also observed, similar to what was seen with

TEM in Pimelodus pictus (Olaya et al., 2007) y

Lates niloticus (Namulawa et al., 2015).

The intestinal morphology of P. magdale-

nae, particularly its length and coiling within

the coelomic cavity, is consistent with findings

reported for various species of detritivorous

fish across different taxonomic groups and

remains consistent within the Prochilodontidae

genus (Bowen, 2022; Burns, 2021; Nachi et al.,

1998). The intestinal length, typically ranging

between 3 and 6 times the body length, the

abundant number of pyloric caeca (which in

this study correspond to the anterior portion

of the intestine), numerous epithelial folds,

and the increased volume of the coelomic cav-

ity, represent convergent evolutionary adapta-

tions in detritivorous fish. A larger surface

area is necessary for efficient nutrient absorp-

tion in a diet low in nutrients (Burns, 2021;

Nachi et al., 1998).

The observations conducted on the ultra-

structure of the intestinal epithelium of P. m a g -

dalenae revealed elongated enterocytes in the

epithelial composition, with oval nuclei exhib-

iting notable electrodensity towards the basal

part of the cell. Towards the apical region,

the cell membrane is adorned with numerous

Revista de Biología Tropical, ISSN: 2215-2075, Vol. 73: e63718, enero-diciembre 2025 (Publicado Jul. 29, 2025)

microvilli, accompanied by a cytoplasm of

higher electrodensity compared to the basal

zone of the enterocyte. Microvilli represent

a characteristic feature of the intestinal epi-

thelium in fish, augmenting the surface area

of the cell membrane projected towards the

intestinal lumen (Alonso et al., 2015; Bosi et al.,

2022; Namulawa et al., 2015). This morphology

maintains a close association with the physi-

ological traits observed in detritivorous fish

species. Similarly, the presence of some goblet

cells producing mixed mucins is observed. His-

tologically, the study of the intestine of P. m a g -

dalenae revealed the presence of neutral and

sulfated acidic mucins, as well as carboxylated

mucins, with moderate intensity in the anterior

and middle segments due to the low number of

mucus-producing cells in these segments.

In the anterior portion, a slight predomi-

nance of neutral mucins over acidic mucins

was observed. In the middle and posterior

segments, there was a predominance of acidic

mucins, mainly carboxylated. The presence of

both acidic and neutral mucins in the intestine

has been widely reported across various spe-

cies and taxonomic groups. Some studies have

yielded similar results for T. sparrmanii (Okuthe

& Bhomela, 2021), O. niloticus (Phrompan-

ya et al., 2019), and B. amazonicus (Vidal et

al., 2020). The presence of different types of

mucins in the analyzed intestinal regions in this

study is associated with the absorption of pro-

teins, ions, and other particles throughout the

intestine (Bosi et al., 2022). These mucins also

play a role in protecting the epithelium against

pathogens and facilitating food and fecal transit

by lubricating the walls of the intestinal lumen

(Genten et al., 2008). Understanding the distri-

bution of acidic and neutral mucins in the gas-

trointestinal tract of P. magdalenae is crucial for

establishing frameworks regarding fish welfare,

as observed in parasitic infestations by Entero-

myxum leei in S. aurata. Additionally, knowl-

edge of the gastrointestinal tract’s ultrastructure

would enable the detection of potential damage

caused by microorganisms to the microvilli in

different gastrointestinal tract regions (Ringø et

al., 2007), as well as determining fish exposure

to various contaminants such as herbicides or

heavy metals (Samanta et al., 2018).

The study reveals distinct morphological

and histochemical characteristics of P. magdale-

nae’s gastrointestinal tract. The stomach exhib-

its a U-shaped, fundic-type structure with three

regions (cardiac, fundic, and pyloric), contain-

ing exclusively neutral mucins for mucosal pro-

tection. A notable thick muscular layer in the

pyloric region aids in mechanical food degra-

dation. The intestine shows considerable length

(intestinal coefficient 2.87 ± 0.19), with multi-

ple organized loops occupying the celomic cavi-

ty. Mucin distribution varies along the tract: the

anterior intestine shows predominantly neutral

mucins, while middle and posterior segments

contain mainly carboxylated acidic mucins

associated with nutrient absorption. The Zihler

index (1.33 ± 0.10) and morphological features

confirm its herbivorous/detritivorous feeding

habits. These findings provide baseline data for

the conservation and commercial production

of this vulnerable endemic species.

Ethical statement: The authors declare

that they all agree with this publication and

made significant contributions; that there is no

conflict 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 supported by ISAGEN S.A.,

Piscícola San Silvestre S.A., HISTOLAB, and

Universidad de Antioquia within the frame-

work of the agreement 33/121 and 33/02168, as

a part of the Management Program for protect-

ing fish and fishing resources of the Sogamoso

River and its floodplain. The funder did not

determine the study design, collection, analysis,

data interpretation, or the decision to submit it

for publication. Except for A.L.E-P, all authors

declare no other competing interests.

22 Revista de Biología Tropical, ISSN: 2215-2075 Vol. 73: e63718, enero-diciembre 2025 (Publicado Jul. 29, 2025)

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