The life cycle begins when a host ingests an oocyst, a resilient, wall-encased structure containing four sporozoites. In the gastrointestinal tract the oocyst excysts, releasing sporozoites that invade the epithelial cells lining the gut. Cryptosporidia are obligate intracellular parasites: they attach to the surface of a gut cell and pull the host membrane around themselves to form a parasitophorous vacuole, inside which they replicate. The sporozoite develops into a meront, duplicating its nucleus and organelles, and after roughly twelve hours divides into eight motile merozoites. These egress and invade neighbouring epithelial cells, repeating the asexual cycle. Remarkably, live-cell imaging of Cryptosporidium parvum infecting HCT-8 cells, together with single-cell RNA sequencing data from in vitro and in vivo infections, indicates that merozoites from the third round of asexual replication are committed directly to sexual development, apparently without a morphologically distinct type II meront of the kind described in other species such as C. muris and C. mortiferum.

Why and how the parasite commits to sexual stages after exactly three asexual rounds remains unknown. What is clear is that the committed merozoite invades a new epithelial cell and differentiates into either a macrogamont, which develops a single female gamete, or a microgamont, which produces sixteen male gametes. Mature males egress and seek out females, which never leave their host cell and instead mature intracellularly until fertilised. Fertilisation produces a zygote that undergoes meiosis, constructs the oocyst wall and completes sporogony to yield four haploid sporozoites within the protective shell. Because standard co-culture of C. parvum with HCT-8 cells does not support observation of fertilisation or subsequent zygote and oocyst development, sexual commitment, fertilisation, meiosis, sporogony and oocyst construction remain among the most mysterious phases of the life cycle.

Sexual commitment itself is mandatory for the third generation of merozoites and appears to be governed by an intrinsic, non-chromosomal mechanism. Live imaging has shown that merozoites from a single meront give rise to both sexes: of the eight merozoites in a meront, approximately two to three become male and the remaining five to six become female. This produces an excess of small, motile microgametes, presumably improving the odds that a female will be found and fertilised. One known player is the transcription factor Myb-M, expressed early in male development; experimental overexpression of Myb-M in C. parvum is sufficient to drive differentiation into males. No equivalent trigger for female commitment has been identified, leading the authors to suggest that female fate may be the default programme of sexual differentiation.

Female parasites, once committed, undertake one of the most demanding developmental programmes in the parasite’s repertoire. They carry a large nucleus, reflecting their status as the most transcriptionally and translationally active life cycle stage. Early in development, females express putative transporters, membrane proteins and secreted proteins hypothesised to mediate fertilisation, possibly facilitating male attachment and entry. Later, they mount two distinct waves of gene expression devoted to building the oocyst wall. The wall itself is a multilayered structure: an external glycocalyx, an outer layer of acid-fast lipids, and an inner layer of glycoproteins. These components are synthesised and stockpiled in female-exclusive organelles called Wall Forming Bodies, or WFBs, of which there are two subtypes distinguished by electron density. The prevailing hypothesis is that WFB Type I contents are secreted first to build the outer layer and WFB Type II contents follow to form the inner layer.

Only a handful of WFB components are confirmed. COWP1 is currently the only validated marker for WFB Type I, while COWP6 and COWP8 show punctate localisation in developing females, suggesting they also reside in these organelles. Transcriptomic and oocyst proteomic datasets have generated a comprehensive candidate list of WFB proteins, but how the organelles are synthesised, what exactly they contain, and how their secretion is regulated remain open questions. The single-cell atlas of C. parvum reveals that the first expression wave contains the Cryptosporidium Oocyst Wall Protein family and accessory enzymes such as proteases and protein modifiers for assembling the proteinaceous shell, while the second wave expresses glycosyltransferases and other secreted proteins including mucins and C-type lectins. These components are secreted into the space between the parasite plasma membrane and the host cell membrane, allowing the wall to be built while the parasite is still inside the gut cell. Having wall components ready just in time presumably keeps pace with the rapid turnover of epithelial host cells.

Females also provision their offspring. Genetic crosses using isolates tagged with different fluorescent proteins have shown that maternal cytoplasmic proteins persist in the sporozoites of first-generation progeny, including proteins involved in assembling dense granule organelles, which remodel the host cell after invasion. Females additionally express the proteins of the crystalloid body, a structure whose function is unknown but is likely involved in sporogony. Post-transcriptional control may be equally important: the maternal RNA-binding protein F-RBP, expressed in early females, binds female RNA transcripts, and conditional knockouts in vivo produce sterile sporozoites. F-RBP may therefore be essential for regulating maternally inherited RNA required to generate infectious sporozoites in the next generation.

Transcriptional regulation of female development is only beginning to be mapped. Two female-specific Apetala-2-type DNA-binding proteins have been identified: ApiAP2-1 is required for female differentiation, and ApiAP2-F is required for oocyst shedding. Insulinase-like protease 1 also contributes to female development and oocyst shedding. Intriguingly, females show reduced expression of ribosomal proteins, indicative of translational repression, and the authors hypothesise that epigenetic mechanisms such as chromatin remodelling may operate, consistent with the expansion of the female nucleus. Whatever the precise combination of mechanisms, they are fundamental to completing the life cycle and represent attractive targets for intervention.

What happens after fertilisation is even less understood. Fertilisation is presumed to trigger oocyst wall formation, sporogony and meiosis, but no culture system permits direct observation, so the signals regulating post-fertilisation development remain unknown across the coccidia. In Cryptosporidium, meiotic divisions appear to precede and partially overlap with deposition of the oocyst wall. Genes related to meiosis and DNA repair are expressed by females, including the DNA meiotic recombinase DMC1, detected in female nuclei. Under experimental conditions, C. parvum chromosomes show Mendelian inheritance with high frequencies of meiotic recombination, and frequent sex between genetically distinct parasites could accelerate the evolution and spread of traits such as virulence or drug resistance, as suggested by clofazimine resistance observed between C. parvum subtypes. Unlike other apicomplexans, Cryptosporidium oocysts are shed fully sporulated and immediately infectious, ready either to be excreted into the environment or to excyst in situ and reinfect the current host, a process that underpins the chronic disease seen in vulnerable patients.

A long-standing model proposes two oocyst types: thin-walled oocysts that excyst within the host to drive autoinfection, and thick-walled oocysts that are excreted and transmitted. The evidence for thin-walled oocysts rests on a few electron micrographs, and single-cell data show no transcriptional signature suggesting that some females are programmed to produce thin-walled oocysts. The authors instead hypothesise that all oocysts are thick-walled and environmentally robust, but that a small number excyst in the gut before excretion; given the exponential asexual cycle, even rare reinfections can sustain heavy parasite burdens. Improved culture systems, including organoid co-culture, are now making post-fertilisation biology experimentally accessible for the first time. Because oocyst biogenesis is exclusive to females and conserved across coccidia, dissecting this female-driven process could expose factors essential for transmission and open new avenues for drugs or vaccines. As the authors argue, it is time to stop overlooking Cryptosporidium females, the true engines of the parasite’s success.

Subject of Research: The biology of female sexual stages and oocyst formation in Cryptosporidium parasites

Article Title: Mysterious girls: The biology of female Cryptosporidium parasites

Article References: Stevens, S., Goddard, P., Walzer, K. A., & Pawlowic, M. C. (2026). Mysterious girls: The biology of female Cryptosporidium parasites. PLOS Pathogens, 22(10), e1014678. https://doi.org/10.1371/journal.ppat.1014678

Image Credits: AI Generated

DOI: 10.1371/journal.ppat.1014678

Keywords: Cryptosporidium, parasitology, oocyst, sexual development, gametogenesis, apicomplexa, Cryptosporidium parvum, transmission, diarrhoeal disease, Wall Forming Bodies, meiosis, global health

The work was led by Isabelle S. van der Meulen and Sabine E. Hannema of Amsterdam University Medical Center, together with colleagues spanning endocrinology, urology, psychology, sexology and gynaecology. This breadth of expertise was deliberate. The authors argue that questions about puberty suppression and sexuality cannot be answered from within a single discipline, because puberty itself is a phenomenon that unfolds simultaneously across the endocrine system, the developing brain, the genital organs and the psychosocial life of an adolescent. The recommendations were further shaped by input from a round table session held at the 2025 conference of the European Professional Association for Transgender Health (EPATH), which brought together clinical and community perspectives on how these conversations should be handled in practice.

At the physiological level, GnRH analogues work by suppressing the hypothalamus–pituitary–gonadal axis, the hormonal cascade that normally drives the physical transformations of puberty. By halting the release of gonadotropins, the medications pause the production of sex steroids and thereby interrupt the development of secondary sexual characteristics such as breast growth, voice deepening, genital development and changes in body composition and distribution. The rationale is to give adolescents with persistent gender incongruence time to explore their identity without the distress of unwanted, and in many cases irreversible, pubertal changes. When treatment is stopped, endogenous puberty resumes; when followed by gender-affirming hormone treatment, a puberty aligned with the adolescent’s gender identity is induced instead.

However, the same sex steroid hormones that drive physical pubertal changes also play roles in genital maturation, neural development and possibly in sexual response and pleasure. Androgens, for example, contribute to clitorophallus growth and penile tissue remodeling, while estrogens influence vaginal and vulvar development. Suppression of these hormones during a critical developmental window therefore raises legitimate biological questions about whether later genital sensitivity, sexual function or capacity for pleasure might be affected. The authors stress that the existing evidence base is too sparse to answer these questions definitively, and that this uncertainty is precisely why structured, evidence- and experience-based counselling is so important for adolescents and their families who are weighing these decisions.

The recommendations generated by the expert group centre on providing developmentally appropriate information about puberty, puberty suppression and sexual development. Clinicians are encouraged to explain, in language suited to the adolescent’s age and cognitive stage, how the hypothalamus–pituitary–gonadal axis works, what GnRH analogues do to it, and what is known and unknown about the consequences for sexual maturation. Importantly, the experts also highlight psychosocial dimensions of sexual development, including the effects of stigma and minority stress, which research in sexual minority populations has shown to be strongly linked to psychological well-being and, by extension, to sexual well-being. Counselling that ignores these social realities, the authors suggest, would present an incomplete picture of what shapes a young person’s sexual development.

Another core recommendation concerns individualized decision making around fertility and future surgical options. Puberty suppression at early pubertal stages limits the development of gametes, complicating options for fertility preservation, and it also affects the tissues available for later gender-affirming genital surgery, such as vaginoplasty. Decisions about when to start suppression, whether to pause treatment to allow sperm or oocyte maturation, and how to counsel about surgical implications must therefore be tailored to each adolescent’s circumstances, values and goals. The experts emphasize shared decision making, in which adolescents, families and clinicians weigh benefits, risks and uncertainties together, supported by decision aids and ethics support tools that have been developed and evaluated in recent years within transgender care.

Equally central is the promotion of positive, open communication about sexuality itself. The authors note that sexuality remains a taboo topic in many clinical settings, including youth mental health care, and that transgender adolescents frequently report wanting their healthcare providers to talk with them about sexuality and gender diversity rather than avoiding the subject. The recommendations call for clinicians to proactively address sexual desire, arousal, pleasure, relationships and masturbation as normal components of adolescent development, rather than treating sexuality purely as a domain of risk or dysfunction. Tools designed to open these conversations with young people, including question-based conversational aids developed for gender-diverse youth, are highlighted as practical resources that can help clinicians overcome their own hesitation and make these discussions routine rather than exceptional.

On the research side, the article identifies major knowledge gaps and sets out priorities for filling them. Longitudinal studies following TGD individuals from the start of puberty suppression well into adulthood are needed, ideally combining quantitative measures with qualitative approaches that capture lived experience. The first exploratory studies in this area have offered a measure of reassurance: they found similar sexual well-being in young adults who received puberty suppression during adolescence and those who received gender-affirming hormone treatment without prior suppression. Research on sexual functioning after vaginoplasty, and on the influence of puberty suppression timing on later sexual function, has begun to map this terrain, but the authors caution that these are early, often small studies that cannot yet support firm causal conclusions. Research frameworks adapted from other fields, such as the target trial framework for observational data, may help strengthen causal inference in this challenging population.

A further priority is the development of TGD-specific instruments to assess sexual development, sexual function and sexual satisfaction. The authors argue that existing sexual health measures, which were largely designed for cisgender populations, fail to capture the diversity and intersectionality of sexual experiences among TGD individuals, including the interplay of gender dysphoria and gender euphoria with desire, pleasure and body satisfaction. Without valid, sensitive measurement tools, the field cannot accurately track outcomes or evaluate whether and how puberty suppression affects sexual well-being over the life course. Genital sensitivity itself, including erogenous sensation in surgically constructed and hormone-influenced genital tissue, is named as an area requiring both psychophysical and patient-reported investigation.

Ultimately, the expert recommendation is framed not as a verdict on puberty suppression but as a framework for honest, compassionate and scientifically grounded care. By integrating evidence where it exists, transparently acknowledging uncertainty where it does not, and centering the voices and experiences of TGD adolescents themselves, the authors aim to support individuals considering puberty suppression, their families and their clinicians in optimizing both care and long-term well-being. The call for longitudinal, interdisciplinary research signals that the scientific conversation is far from over, and that the coming decade of work will be critical in transforming clinical experience into rigorous evidence about how best to support the sexual development and sexual well-being of transgender and gender-diverse young people.

The biological rationale for caution rests on well-established endocrine principles. Sex steroids act not only as activators of sexual response but also as organizers of tissue development during defined windows of maturation, a concept long recognized in classical studies of pubertal staging in boys and girls. Analogies can also be drawn from conditions such as congenital hypogonadotropic hypogonadism, in which individuals experience absent or delayed endogenous puberty and require hormonal induction; research in these populations has examined psychosexual development under long-term treatment and offers a useful comparative lens for understanding how pubertal hormone exposure relates to later sexual function.

The clinical context for these recommendations has been shaped by several prior expert documents. The European Society for Sexual Medicine has issued position statements addressing hormonal management and sexual function in adolescent and adult trans people, as well as sexual well-being after gender-affirming surgery, and a working group of the European Society for Paediatric Endocrinology has published expert opinion on the endocrine management of transgender and gender-diverse adolescents. The new recommendation complements these efforts by focusing specifically on the intersection of puberty suppression and sexuality, an area those documents addressed only briefly.

Conceptual framing also matters. The World Health Organization defines sexuality broadly, encompassing sex, gender identities and roles, sexual orientation, eroticism, pleasure, intimacy and reproduction, and the ICD-11 now classifies gender incongruence outside mental and behavioural disorders. Within this framing, sexual well-being is understood as a positive dimension of health rather than merely the absence of dysfunction, which reinforces the authors’ argument that counselling should address pleasure and satisfaction, not only risk.

The evidence base itself has grown out of decades of longitudinal cohort research, most notably the Amsterdam Cohort of Gender Dysphoria, which has documented diagnostic and treatment trajectories since the earliest days of the Dutch protocol. Recent pilot studies from this tradition have begun comparing sexual function in adults treated with gender-affirming hormones with or without prior pubertal suppression, and examining sexual satisfaction following vaginoplasty in relation to the timing of suppression. Although preliminary, these studies represent the first direct empirical tests of questions that were previously addressed only through theory, and they provide the methodological foundation on which the recommended larger, longitudinal and mixed-methods research programme can now be built.

Subject of Research: The effects of puberty suppression with GnRH analogues on sexual development and sexual well-being in transgender and gender-diverse adolescents

Article Title: Puberty suppression and sexual well-being in transgender and gender-diverse individuals: evidence- and experience-based counselling and future research directions

Article References: van der Meulen, I. S., van den Boogaard, E., Özer, M., Bungener, S. L., Vrouenraets, L. J., & Hannema, S. E. (2026). Puberty suppression and sexual well-being in transgender and gender-diverse individuals: evidence- and experience-based counselling and future research directions. Nature Reviews Urology. https://doi.org/10.1038/s41585-026-01182-3

Image Credits: AI Generated

DOI: 10.1038/s41585-026-01182-3

Keywords: puberty suppression, GnRH analogues, transgender health, gender-diverse adolescents, sexual well-being, sexual development, fertility preservation, shared decision making, gender-affirming care, EPATH, Nature Reviews Urology, expert recommendation