Intersexuality represents a significant reproductive and developmental anomaly in gonochoristic animals, characterized by the simultaneous coexistence of male and female characteristics—either externally in primary and secondary sexual traits or internally within reproductive organs (e.g., ovitestes) (Ford 2012). Manifestations of this anomaly are diverse, ranging from subtle microscopic histological variations to marked alterations in the macroscopic structure and physiological function of gonads (Stentiford 2012). Widely documented in aquatic invertebrates, intersexuality is particularly prevalent among crustaceans, with reported occurrences across major lineages including Amphipoda, Decapoda, Isopoda, and Mysida (see Ford 2012). This widespread distribution suggests intersexuality may have deep evolutionary origins and a shared developmental biological basis, making it a key focus for understanding crustacean sex determination and phenotypic plasticity.

The Chinese mitten crab, Eriocheir sinensis Milne-­Edwards, 1853, is an economically prominent freshwater crab species in China. Since the pivotal breakthrough in its artificial propagation was achieved in 1984, extensive research has been conducted on its reproductive biology and developmental processes, resulting in a relatively comprehensive understanding of these aspects. In recent years, there has been an increasing incidence of intersex individuals in this species. Typically, these cases involve male crabs infected by the rhizocephalan parasite, Polyascus gregarius (Okada et Miyashita, 1935) (see Tang et al. 2005), while a more recent report has documented genetically female crabs undergoing masculinization (Lu et al. 2024). However, a consistent feature of all previously documented intersex E. sinensis individuals is an intermediate abdominal shape—exhibiting forms such as elliptical, rectangular, asymmetrical, or isosceles triangular—that falls between the distinct male (triangular) and female (circular) phenotypes of this species.

Notably, the present study describes the first documented case of an intersex E. sinensis with a fully typical female external phenotype, whose abdomen retains the characteristic circular shape unique to normal females. This distinct intersex phenotype, combining a complete female somatic morphology with developed male gonopods, differs fundamentally from all previously reported intersex individuals of this species. Given the complex etiology of intersexuality, the discovery of this unique individual not only enriches the phenotypic diversity of intersexuality in E. sinensis but also suggests the existence of novel underlying mechanisms driving intersex occurrence in this economically important species. Herein, we detail the morphological characteristics of this intersex specimen, aiming to provide foundational data for further investigations into the sex determination pathways and reproductive developmental plasticity of crustaceans.

An intersex individual of Eriocheir sinensis was sampled from a commercial aquaculture farm in Gaochun District, Nanjing, Jiangsu Province, China (32°1'31.8"N, 118°44'28.5"E) on 12 October 2025. Upon collection, the specimen was initially identified as a normal female. It appeared healthy and showed no signs of parasitic infection. However, when its abdomen was gently spread open for non-invasive examination of gonadal development and somatic condition, a pair of distinct males first gonopods (G1) was observed. Immediately following on-site observation, the specimen was transported to the laboratory for morphological measurements. All metrics were recorded point-to-point using a vernier caliper (precision: 0.1 mm), including carapace length, carapace width, body height, frontal width, and the distance between the first anterolateral teeth. Subsequently, the specimen was preserved in 95% ethanol and deposited in the collection of the Freshwater Fisheries Research Institute of Jiangsu Province under the registration number JSFFRI/25002 for subsequent histological analysis and long-term archival storage (Fig. 1).

The intersex specimen exhibited a fully female external phenotype with a total weight of 179.2 g. Its carapace measured 70.1 mm in length and 78.3 mm in width, presenting an overall rectangular morphology. Additionally, the body height, frontal width, and distance between the first anterolateral teeth were recorded as 38.5 mm, 17.3 mm, and 41.2 mm, respectively. Notably, the ratios of all measured morphological traits were consistent with the national standards of China for E. sinensis.

The left and right chelipeds and pereiopods of the specimen were morphologically symmetrical. Sparse, short setae occurred on the outer surfaces of the cheliped propodus and basal half of the dactylus, whereas the inner surfaces were smooth and glabrous (Fig. 1A). In contrast, among previously reported intersex individuals, only those with an elliptical abdomen showed female-like cheliped setal traits. All others with a more triangular abdomen exhibited male-like setal patterns, where setae extensively covered the full propodus and most of the dactylus base across the entire outer manus surface, extending even to the carpus-manus junction and appearing densely packed, fluffy, and plump (Lu et al. 2024).

The specimen’s abdomen displayed a circular morphology typical of normal female individuals (Fig. 1B). When the abdomen was spread open, a pair of female gonopores were identified on the fifth thoracic somite, accompanied by the second to fifth pairs of ovigerous pleopods bearing normal setae—all characteristic traits of typical female crabs (Fig. 2). However, a pair of ossified, tubular copulatory organs, namely the first gonopods (G1), was also observed, a feature consistent with that of normal males. This finding is inconsistent with previous reports on intersex crabs possessing rounded abdomens. In those cases, the G1 typically exhibits incomplete development with reduced calcification, whereas intersex individuals with a more triangular abdomen present normally calcified G1 structures (Lu et al. 2024). Notably, no second gonopods (G2) or penises—structures essential for reproductive functions—were detected. Although no invasive dissection was performed, mature female gonads were clearly visible through the first and second abdominal flaps.

The Chinese mitten crab E. sinensis exhibits striking sexual dimorphism. Tang et al. (2005) previously documented parasite-induced degeneration of male phenotypic traits in intersex E. japonica. Such Polyascus-infected individuals are commonly observed in aquaculture, where infection causes damage to the androgenic gland (AG) in males and ultimately leads to the degeneration of male gonads and reproductive organs. In contrast, the effects of such infection on females are limited to ovarian degeneration and growth retardation, with no induction of intersex phenotypes. The intersex individual identified in the present study shares similarities with the recently reported genetically female crabs undergoing masculinization (Lu et al. 2024). However, the four typical abdominal morphologies of intersex crabs described by Lu et al. (2024) display a high degree of masculinization. In addition to possessing the first gonopods (G1), second gonopods (G2) and penis, these individuals also have well-developed male gonadal tissues. Notably, the intersex individual in this study retained relatively intact female phenotypic characteristics and completely lacked both the second gonopods (G2) and penis (Pe). These features indicate the absence of a complete male reproductive tract and support its classification as a case of low-degree masculinization.

In crustaceans, crustacean female sex hormone (CFSH) is tightly associated with female sexual differentiation (Adkins-Regan 2014), whereas insulin-like androgenic gland hormone (IAG), secreted exclusively by the androgenic gland (AG), is a well-characterized peptide hormone that critically regulates male sexual differentiation (Levy et al. 2020). Previous AG implantation experiments in various female crustacean species have confirmed that induced masculinization is restricted to external sexual characteristics (Fowler and Leonard 1999). In E. sinensis, CFSH-1 initiates feminization by effectively repressing IAG expression starting from the third zoeal stage (Z3) during larval development. After the fourth zoeal stage (Z4), IAG expression surges and CFSH-1 expression declines extremely significant in male larvae, whereas an opposite expression trend is observed in female larvae (Zhu et al. 2022). Sexual differentiation of the reproductive organs subsequently initiates at the megalopa stage (Lee et al. 1994). Collectively, these findings suggest that the intersex individual identified in the present study may have been exposed to multiple stressors during the late larval developmental stage, including parasitism, environmental endocrine-disrupting chemicals (EDCs) such as androgenic compounds, and abnormal water temperatures (Rodgers-Gray et al. 2004; Lye et al. 2005). These stressors likely interfered with normal hormonal signaling pathways, resulting in a moderate upregulation of IAG expression while CFSH-1 expression remained unaltered to sustain normal female reproductive system differentiation and development. This dysregulated hormonal balance ultimately induced low-degree masculinization and the development of male first gonopods (G1). Nevertheless, the above hypothesis of moderate IAG upregulation with unaltered CFSH-1 expression currently lacks direct molecular evidence from the examined intersex specimen. Therefore, future studies are required to collect more intersex E. sinensis individuals, perform systematic histological analysis of gonads and appendages, and quantify the transcriptional levels of IAG and CFSH-1. These follow-up works will further validate the hormonal dysregulation hypothesis and clarify the molecular mechanism underlying intersex formation in E. sinensis. Overall, the present study identifies a novel type of intersex phenotype in E. sinensis and provides a valuable model for investigating the early developmental timing of sexual differentiation, sex determination pathways, and phenotypic plasticity in crustaceans.