Monitoring of Atelopus colomai in One of the Last Tropical Rainforest Patches Next to Puyo City, in the Western Amazon of Ecuador

Introduction

Amphibians constitute one of the most threatened vertebrate taxa on the planet, with approximately 41% of assessed species classified under some category of threat (1). The primary drivers of this decline encompass habitat loss and fragmentation, climate change, environmental pollution, and infectious diseases such as chytridiomycosis, which is caused by exposure to the chytrid fungus Batrachochytrium dendrobatidis (2). This pathogen has driven severe population collapses in amphibians across the Tropical Andes.

Harlequin toads of the genus Atelopus represent one of the most drastically affected groups within the global amphibian decline of the late 20th century. This genus is distributed from Costa Rica to Bolivia, comprising more than 100 species that are predominantly endemic and exhibit restricted geographic ranges (3). Numerous studies have documented severe population declines and extinctions of multiple Atelopus species, phenomena tightly linked to the spread of chytridiomycosis and anthropogenic habitat alteration (4,5).

Ecuador is among the countries with the highest diversity of harlequin frogs, with approximately 25 recorded species, the majority of which are classified as Critically Endangered or Possibly Extinct following the catastrophic population collapses of the 1980s and 1990s (6). However, over the last decade, the landscape of herpetological conservation has experienced an unexpected shift due to the rediscovery of remnant populations previously thought to be extinct (7).

The eastern slope of the Ecuadorian Andes, specifically the Upper Pastaza watershed and the Llanganates-Sangay Connectivity Corridor, represents an area of exceptional biological priority. Herpetological assessments in this region have demonstrated that complex topography and local climatic gradients drive drastic biotic turnover and remarkably high rates of microendemism. Consequently, even adjacent mountains exhibit markedly differentiated amphibian communities that are highly threatened by anthropogenic activities (8). Foothill populations of Atelopus in this region were historically treated as part of the Atelopus spumarius species complex, but recent phylogenetic analyses resolved the taxonomy of the populations inhabiting Pastaza, establishing them as a distinct species named Atelopus colomai (9).

To generate ecological data on the communities of this newly described taxon and to safeguard its remaining remnant populations, the Upper Pastaza Atelopus Project was established with support of the Amphibian Survival Alliance. This initiative aims to design and implement targeted conservation projects to ensure the long-term survival of the species.

The objective of the present study was to monitor the population of Atelopus colomai within the Coloma's Harlequin Frog Conservation Block. This was achieved by recording individuals, evaluating population structure, characterizing microhabitat use, and detecting the presence of Batrachochytrium dendrobatidis. The generated data seek to advance the ecological knowledge of the species and strengthen local conservation strategies.

Methodology

Study Area

The study was conducted in the province of Pastaza, near the city of Puyo, within the Coloma's Harlequin Frog Conservation Block. This block comprises Finca La Argentina, Finca Anders Sirén, and the Yanacocha Refuge and Zoo. The ecosystem consists of eastern foothill forest (pie-montano) characterized by high humidity levels, temperatures ranging between 18°C and 24°C, and an annual precipitation exceeding 3,000 mm. The Churuyaku River flows through the study area, which was mapped as it constitutes a critical component of the habitat of Atelopus colomai (Map 1).

Sampling Design

Monitoring of Atelopus colomai was conducted from January to July 2025, during which 24 nocturnal surveys were carried out. The sampling methodology consisted of Visual Encounter Surveys (VES) and acoustic surveys along four 500-meter transects established within the conservation block where Atelopus colomai has been historically recorded. All individuals were handled in strict accordance with established biosecurity protocols (10).

Data Collection and Individual Registration

Every observed individual was logged using the Epicollect5 mobile platform. Each specimen was assigned a unique registration number associated with the following parameters: date, time, geographic coordinates, elevation, sex, developmental stage (juvenile or adult), microhabitat, perch height, perch substrate, and photographic records (dorsal, ventral, left lateral, and right lateral views).

Epidermal swabbing was performed on each individual to determine the presence or absence of Batrachochytrium dendrobatidis within the studied population. Individual identification was achieved through photo-identification, utilizing the unique dorsal coloration patterns characteristic of each specimen. Because these patterns are unique and remain stable over time, recaptured individuals could be reliably recognized without the need for invasive marking techniques that could compromise individual welfare.

Environmental Variables and Habitat Characterization

Environmental parameters were recorded for each individual encounter. Ambient temperature and relative humidity were measured using a thermo-hygrometer, while forest canopy cover and other specific habitat attributes were evaluated via direct field observations. To further contextualize habitat ecology, syntopic amphibian species co-occurring with Atelopus colomai were also recorded.

Results

Abundance and Recaptures

Throughout the 24 nocturnal surveys conducted in the study area, a total of 87 Atelopus colomai encounters were recorded. Individual identification based on unique dorsal color patterns and line configurations allowed for the non-invasive determination of recaptures. A total of 10 recaptures were identified, yielding a total of 77 unique individuals documented during the monitoring period (Table 1).

Population Structure

The demographic structure of the population consisted of 38 adult males, 37 adult females, and two juvenile individuals (one male and one female). The observed sex ratio was close to parity at 1:1 (1.03 males per female), suggesting a balanced sex distribution within the assessed population (Table 1).

Table 1. Summary of sampling effort, abundance, population structure, reproductive evidence, and prevalence of Batrachochytrium dendrobatidis (Bd).

Spatial Distribution

The spatial distribution of Atelopus colomai within the study area revealed a marked, sex-specific habitat preference. Female records occurred with higher frequency in interior forest zones, well separated from the Churuyaku River. Conversely, males were consistently found aggregated along the riverbanks. This spatial segregation behavior was highly pronounced throughout the study.

Microhabitat Use

Individuals utilized a variety of substrates for nocturnal perching. The most frequently utilized substrate belonged to plants of the family Araceae (40 records), followed by Pteridophyta (13 records) and Melastomataceae (11 records) (Graph 1).

In terms of vertical stratification, individuals were recorded on perches ranging from ground level up to 2.5 meters in height. The highest frequency of observations clustered around 1.0 meter (15 records), followed by 1.2 meters (12 records). These data indicate a clear preference for the lower strata of the understory, although observations exceeding 2.0 meters demonstrate vertical habitat utilization (Graph 2).

Environmental Variables

During the monitoring surveys, ambient temperatures fluctuated between 17°C and 23°C, while relative humidity remained consistently high, between 91% and 99%. These microclimatic conditions are highly characteristic of pre-montane foothill forests, reflecting an environment with high moisture availability and moderate temperatures that favor the activity, survival, and reproduction of Atelopus colomai.

Reproductive Evidence

Two juvenile individuals (one male and one female) were recorded during the monitoring period, providing empirical evidence of recent recruitment within the population. The presence of these juveniles indicates that reproductive activity in Atelopus colomai is successfully ongoing within the studied area.

An amplexus event was documented in the month of May. According to existing data available (11), May is an atypical period for this behavior, as it falls outside the previously documented months for amplexus in this species.

Sympatry with Other Species

Atelopus colomai shares its ecological niche with a highly diverse assemblage of amphibians, which are photographically logged and listed in Photo 3. These co-occurring taxa reflect the remarkable species richness and ecological complexity of the study area.

Presence of Batrachochytrium dendrobatidis (Bd)

Diagnostic analysis of the epidermal swabs confirmed the presence of Batrachochytrium dendrobatidis (Bd), the fungal pathogen responsible for chytridiomycosis. The pathogen was detected in 13% of the sampled population, meaning that 10 out of the 77 unique individuals tested positive for Bd (Table 1).

The detection of Bd confirms the active circulation of the fungus within this population. Consequently, these findings highlight the critical importance of maintaining continuous epidemiological surveillance programs to evaluate the long-term effects of the pathogen on population dynamics and species prevalence.

Conclusions

The presence and short-term stability of a population of Atelopus colomai were confirmed within the foothill forests of Pastaza, inside the Coloma's Harlequin Frog Conservation Block, registering a total of 77 unique individuals and validating photo-identification as an effective, non-invasive monitoring methodology.

The population exhibits a balanced demographic structure with a 1:1 sex ratio, alongside strict spatial segregation wherein males primarily occupy the margins of the Churuyaku River and females utilize interior forest habitats.

Atelopus colomai exhibits a marked preference for low-stratum understory perches (1.0 to 1.2 m in height), preferentially selecting vegetation of the family Araceae as its primary nocturnal perch substrate.

Empirical evidence of reproductive capacity and recent recruitment was documented through the discovery of juvenile individuals and a documented amplexus event in May, expanding current knowledge regarding the reproductive phenology of this bufonid on the eastern Andean slope.

Despite recording a 13% prevalence of the pathogenic fungus Batrachochytrium dendrobatidis (Bd), the active population dynamics suggest a degree of resilience, underscoring the critical need to sustain long-term monitoring and apply strict biosecurity protocols to mitigate the impacts of chytridiomycosis.

Acknowledgements

We would like to express our sincere gratitude to the Amphibian Survival Alliance and Re:wild for their support of amphibian conservation efforts. We are especially grateful to the Instituto Nacional de Biodiversidad (INABIO) and Pablo Jarrín, and to Guadalupe Vinueza, owner of Finca La Argentina, for their openness, hospitality, and the logistical support provided during fieldwork. We also thank the members of the Biology Club of the Universidad Estatal Amazónica for their valuable assistance, dedication, and collaboration throughout the study, which were essential for the successful completion of field activities. We further acknowledge the support of Fundación EcoMinga, Julio Carrión, Javier Robayo, Marco Montero, Fundación Oscar Efrén Reyes, the Zoology Laboratory of Universidad San Francisco de Quito, and Centro Jambatu for Research and Conservation of Amphibians. Special thanks are extended to Andrea Terán and Luis A. Coloma (Centro Jambatu), and the University of Trier, for their contributions and support throughout this project.

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