Myxozoans constitute an economically important group of microscopic metazoan parasites that cause diseases in a wide variety of commercially important fish species [1]. These parasites are distributed worldwide, infecting fish in both marine and freshwater environments [1][2]. In freshwater ecosystems, myxozoans have complex life cycles involving alternating forms in fish and aquatic oligochaete worms, designated as myxosporean and actinosporean stages respectively [3][4]. Myxosporeans can infect any organ of the fish body, causing various types of damage [1]. The extent of tissue and organ damage in the host is influenced by biotic factors such as the species of myxozoan, its life cycle stage, infection intensity, and the host’s reaction, as well as abiotic factors [5]. Compared to coelozoic species, histozoic myxosporeans are more frequently found to attack the gills [6]. Beyond their pathogenic effects, myxozoans play crucial ecological roles in aquatic ecosystems [1]. As parasites, they regulate fish populations by affecting host survival, growth, and reproduction. Severe infections can lead to mass mortality events, altering community structures and influencing trophic interactions [2][3]. Despite their widespread presence and ecological importance, myxozoan research remains limited in certain regions, including Cameroon. Studies on the diversity, distribution, and life cycle dynamics of myxozoans in Cameroonian freshwater systems are scarce [5]. The impact of myxozoan infections on local fish species, many of which are critical to artisanal and commercial fisheries, remains poorly understood.

With the increasing interest in fish culture and production, awareness of parasites affecting fish health, growth, and survival has also increased [7]. Cyprinid fish of the genus Labeobarbus Rüppel, 1836 are widely distributed in Africa, particularly in the rivers of Cameroon, such as the Moliwè River in Limbé. These fish play a crucial role in river ecosystems and are an important source of sustenance for local communities [8]. Labeobarbushabereri (Steindachner, 1912), commonly known as yellow fish, attains a large size and is one of the most economically important fish species in Cameroon. High reproductive capacity, rapid growth, widespread, and commercial acceptance make L.habereri a promising species for fish farming in Cameroon.

The popularity of L. habereri as a food source reinforces the need for parasitological studies. Understanding the ecological aspects of myxozoan species parasitizing this fish is essential. Disease outbreaks affect the productivity of both natural and artificial aquatic systems, negatively impacting the livelihoods of communities dependent on these resources. Mixed myxozoan infections are common, especially in cyprinid fish [9]. Ecological studies provide information that helps understand how these parasites communities are structured, as well as the processes involved in maintaining their structure [10]. This observation, directly applicable to myxosporean parasites of fish, not only allows for a better understanding of the biology of parasitic species but also helps in developing management strategies.

This study aims to examine the prevalence, mean cyst load, and microhabitat selection determinants of five species of myxosporean found in the gills of L. habereri from the Moliwè River in Cameroon.

Between March 2018 and February 2020, specimens of L.habereri (Family Cyprinidae) were collected from the Moliwè River near the village of Moliwè, adjacent to Limbé in the southwest region of Cameroon. The region exhibits a tropical climate characterized by two main seasons: an extended rainy season from March to November and a dry season from December to February. The Moliwè River is typified by variable water flows, substrates that range from rocky to sandy, and abundant native riparian vegetation. The fish were caught at different sites from coordinates 04˚ 03' 38.278'' N - 09˚ 14' 45.516'' E to 04˚ 04' 04.328'' N - 09˚ 16' 43.396'' E and at these sites, standardized gill nets with predetermined mesh sizes were deployed during peak fish activity periods, either in the early morning or late afternoon, for a fixed duration of 2 - 4 hours. Immediately after capture, specimens were measured, sexed, and preserved in a 10% formalin solution for a minimum of 48 hours prior to dissection.

In the laboratory, prior to identify the exact location of the parasites on the gills, each fish specimen was dissected under an Olympus BO61 stereomicroscope with the aim of extracting the eight (08) holobranchs, each half of which comes from one side (left or right) of the fish’s body. Holobranchs were divided arbitrarily following the method of [11] with slight modifications (Figure 1). The holobranchs were individually placed in Petri dishes filled with tap water and numbered I, II, III, and IV in the antero-posterior direction. Each holobranch was further subdivided: the gill filaments were divided into anterior hemibranch (AH) towards the operculum and posterior hemibranch (PH) away from the operculum. Each hemibranch was then subdivided dorsoventrally into ventral portion (VP), median portion (MP), and dorsal portion (DP), based on the number of gill filaments. Additionally, based on the length of the gill filaments, each hemibranch was divided into proximal zone (PZ), middle zone (MZ), and distal zone (DZ). This division resulted in 72 microhabitats per fish side, which were examined under a binocular microscope to search for and count myxozoans cysts. After locating the cysts, some were taken and gently crushed between a slide and coverslip in a drop of water, then examined using a ×100 objective of an Ivymen light microscope. Myxozoans species were identified following the guidelines of [12]. The number of cysts and their exact locations were recorded for each parasite species.

Figure 1. Schematic diagram of an hemibranch showing longitudinal and transversal subdivisions.

Parasite prevalence or infection rate (%), expressed as a percentage, was estimated by dividing the number of L. habereri individuals infected with one or more individuals of a particular myxosporean species by the total number of L. habereri examined [13]. Mean intensity ( x ¯ ), as referred to by [13], was calculated as the average number of cysts of a specific myxosporean species found among infected L. habereri individuals in the sample, divided by the number of L. habereri infected with that myxosporean species. These epidemiological indices were assessed at the microhabitat level using the same approach. To compare prevalence, the chi-square test was employed (χ²). To compare multiple groups, the Kruskal–Wallis H test was employed, while pairwise comparisons were performed using the Mann-Whitney U test. These non-parametric methods were selected due to their robustness against violations of normality, reduced sensitivity to outliers, and applicability to ordinal or skewed data. This approach provides a reliable framework for assessing differences in central tendency even when the data do not satisfy the assumptions required for parametric tests. All statistical analyses were performed using the SPSS program for Windows version 26.0, with significance set at p < 0.05.

Examination of 760 L. habereri specimens’ gills revealed the presence of Myxoboluspaludinosus Reed, Basson and Van, 2002; Myxobolusmakombensis Feudjio-Dongmo, Lekeufack-Folefack, Tene-Fossog, Fomena, Wondji, Yurakhno, Alomar and Mansour, 2022; Myxobolus sp., Myxobolusnyongana (Fomena, Bouix and Birgi, 1985) Fomena and Bouix, 1997 and Thelohanellusassambai Fomena, Marquès, Bouix & Njiné, 1994. Prevalence varied from 14.61% (T. assambai) to 98.16% (M. paludinosus), while mean cyst load ranged between 7.18 ± 0.64 (M. nyongana) and 598.56 ± 29.92 (M. paludinosus) (Table 1). At the xenocommunity level, there was no significant difference in infestation rate (χ² = 0.001; p = 1) or mean cyst load (U = 266,129; p = 0.17) between the two sides of the host’s gill. However, the distribution of myxozoans species was balanced between the left and right sides of the gill, except for M. makombensis, which had a significantly higher average cyst count on the left side (Table 1). Parasite prevalence significantly varied (p < 0.001) regardless of the gill side, with the following patterns: M. paludinosus > M. makombensis > M. nyongana > Myxobolus sp. = T. assambai for the left side, and M. paludinosus > M. makombensis > M. nyongana > Myxobolus sp. > T. assambai for the right side. Similarly, average cyst accumulation differed significantly (p < 0.001) on both gill sides, showing the same distribution pattern on both gill side: M. paludinosus > T. assambai > M. makombensis > Myxobolus sp. = M. nyongana (Table 1).

Examination of the gills of L. habereri revealed the presence of five species of myxozoans: M. paludinosus, M. makombensis, Myxobolus sp., M. nyongana, and T. assambai. According to [14], most known myxozoans species infect the gills, likely because releasing myxospores into the external environment is easier from this organ [15][16]. The heterogeneous structure of the gills allows myxozoans cysts to develop in the gill filaments, gill lamellae, and the cartilaginous arch. Furthermore, the extensive vascularization of the gills favors the development of histozoic myxozoans species because the capsule formed around the myxozoans plasmodium does not sufficiently inhibit the diffusion of oxygen nutrients to the parasite [17]. [16] Report that myxozoans often infest the gills because they are rich in blood and promote gas exchange. The infestation of the gills by myxozoans is not without effects; [18] report histological changes in the gills due to the presence of myxozoans. These changes include inflammation, hypertrophy of lamellar cells, fusion of neighboring gill filaments, and lifting of the gill epithelium.

Polyparasitism in the gills has previously been reported in Cameroon in Enteromiusmartorelli Roman, 1971 [19], Ctenopomapetherici Günther, 1864 [20], and Schilbemystus Linné, 1758 [21]. Occurrence of polyparasitism can be explained by the low parasite densities in natural environments, which leave niches available on the gills of the host fish, facilitating simultaneous infestation by several myxozoans species. Furthermore, multispecies infection does not lead to competition as long as space and resources are sufficient [22][23].

In L. habereri, parasite prevalence ranged from 14.61% to 98.16%, and the mean cyst load varied from 7.18 to 598.56 cysts, confirming the observations of [24] that the epidemiological indices of myxozoans in fish fluctuate significantly in natural environment. Additionally, [25] also notes that the proportion of hosts affected by a parasites species is not fixed and may vary across its geographic range.

Overall, the epidemiological indices of myxozoans species in L. habereri did not vary significantly between the left and right side of the gills. The equitable distribution of myxozoans on both sides of the gills has been reported by [10][20][21] and [24]. This distribution is likely due to the bilateral symmetry of the gill biotope, which results in similar volumes of water flowing through both sides of the gills [26], ensuring an equitable supply of actinospores to both sides.

This study indicates that the first two to three holobranchs of L. habereri are more infected by certain myxozoans species, with M. paludinosus cysts accumulating less in holobranch IV. Several authors have also revealed that holobranch IV is the least affected by myxozoans species in Cameroon, they are: [27] for Myxobolusnyongana and Myxobolusnjinei Fomena, Bouix and Birgi, 1935; [20] for Henneguyapethericii Fomena, Lekeufack Folefack and Bouix, 2008 and Myxoboluspethericii Fomena, Lekeufack Folefack and Tang II, 2007; and [21] for Henneguyacamerounensis Fomena and Bouix, 1957 and Henneguyantondei Fomena, Lekeufack Folefack and Bouix 2008. The preference for holobranchs I, II, and III is likely due to respiratory water currents and the larger surface area available for colonization [26][28]. Holobranch IV, being covered by other holobranchs, receives a reduce water current and offers a smaller surface area for actinospores fixation and cyst development.

The distribution of myxozoans on hemibranchs varies according to species, with some preferring the anterior hemibranch of certain arches and others the posterior hemibranch [29]. In this study, M. makombensis cysts were significantly more numerous on the anterior hemibranch of holobranchs III and IV, similar to M. pethericii in C. petherici [20] and M. nyongana and M. njinei in Enteromiusmartorelli [27]. Conversely, H. ntondei had a higher cyst load on the posterior surface of holobranchs I and III in Shilbemystus [21]. Furthermore, [24] also found that, in Labeorohita, Thelohanellusrohita mostly affects the posterior hemibranch of all holobranchs. The variability in hemibranchs colonization could be due to changes in gill conformation during the respiratory cycle [30]. The epidemiological indices of other myxozoans species and xenocommunities were similar between the anterior and posterior surfaces of the holobranchs, supporting [21], who observed an equitable colonization of both sides of the holobranchs by Henneguyacamerounensis and the xenocommunity in Schilbemystus. This equitability appears because the hemibranchs of a holobranchs generally have the same filaments structure and number. [31] affirmed that the type of hemibranch does not significantly influence the developmental capacity of parasites.

Generally, the median portion of the gill was more affected by myxozoans recorded, likely due to the larger surface area provided by more developed gill filaments. Similarly, [20] reported that the median portion of the gills of Ctenopomapetherici is more infested by M. pethericii and the xenocommunity. Similarly, M. nyongana exploits the medial portion of holobranch III in Enteromiusmartorelli [27], and this was attributed to it exposure to the high respiratory water current.

To the best of our knowledge, the unique study dealing with the variation of epidemiological indices of myxozoans on gill filament zone was done by [27], who indicated that Myxobolusbarbi (syn: Myxobolusnyongana) exploits all filamentous zones with mean cyst load decreasing progressively from the distal to the basal part. In the present study, the infection pattern of the three filamentous zones varied among myxozoans species, likely because the gill system offers different microhabitats, reducing interspecific competition [32] as long as space and resources are sufficient [23]. In addition, [33] suggest that parasite microhabitats vary according to species, developmental stage, host species, and immune response intensity.

Regardless of the fragmentation mode of the gill system, most myxozoans species recorded in L. habereri were present in the 72 selected microhabitats, with their epidemiological indices varying significantly from each other. It is likely that all fish examined were exposed to actinospores from the Moliwè River. If exposure begins at the fish’s first minute of life [34] and the epidemiological indices vary, therefore the developmental kinetics of these parasites species within the host fish differ significantly. The most critical factor affecting parasite development kinetics is the host’s immune response, which varies according to parasite species, target tissue, and host individual [35]. Niche overlap in the spatial distribution of the five species recorded in L. habereri suggests a lack of interspecific competition, consistent with observations in other myxozoans species in Cameroon [19][20] and [21]. This absence of competition can be explained by unsaturated gill niches, likely due to low diversity and low infection levels, as the maximal parasite carrying capacity is rarely reached in natural conditions [36].

This study revealed five species of myxosporeans with varying prevalence and mean cyst load on the gills of L.habereri in the Moliwè River. The distribution of parasites and variations in mean cyst load among different gill microhabitats suggest specific ecological preferences influenced by both biotic and abiotic factors. Myxoboluspaludinosus emerges as the dominant species, raising concerns about its implications for the management of L.habereri. This study emphasizes the necessity of understanding the biology and ecology of myxosporeans to inform resource management and establish disease prevention strategies in the Moliwè River. Therefore, future research should explore the mechanisms behind myxosporean distribution in L.habereri by investigating water quality, temperature gradients, and host tissue characteristics influencing microhabitat preferences. Studying the pathogenicity of Myxoboluspaludinosus and its interactions with the host’s immune system can clarify infection thresholds and immune responses. Employing advanced molecular and phylogenetic tools will refine species identification, reveal cryptic species, and elucidate evolutionary adaptations. Establishing long-term monitoring programs in the Moliwè River will uncover seasonal and environmental influences on parasite prevalence. Finally, integrating parasite biology with fisheries management can develop effective disease prevention and sustainability strategies, overall safeguarding local food security. By pursuing these future research avenues, we can better safeguard both fish populations and the livelihoods that depend on them.

L. F. G. B., A. E. T. T. and N. O. O. A. drafted the proposal, L. F. G. B. and A. E. T. T. participated in the field work and laboratory analysis, L. F. G. B., A. E. T. T. and N. O. O. A. participated in the data analysis and interpretation, F. A. contributed to the correction of the final draft of the manuscript. All authors read, corrected and approved the final manuscript.