Crayfish plague is a water mold, an oomycete pathogen native to southeastern North America, where local crayfish species tolerate it thanks to millions of years of coevolution. When North American crayfish were shipped to Europe for aquaculture and stocking, they brought the fungus with them as an invisible passenger. The pathogen reproduces asexually by releasing zoospores, microscopic swimming spores that home in on new hosts using chemical cues. For European native crayfish, which never evolved defenses against it, infection is usually catastrophic: the fungus invades the cuticle, melanization responses fail, and mass mortality follows within days to weeks.

The consequences for Europe’s freshwater ecosystems have been devastating. Native crayfish are keystone species and ecosystem engineers, large omnivores that structure food webs and shape streambeds. Crayfish plague swept across Europe in two major waves, the first beginning in northern Italy in 1859 and the second in the late 1980s, driven by introduced signal crayfish (Pacifastacus leniusculus) and spinycheek crayfish (Faxonius limosus). Populations of the white-clawed crayfish have declined by an estimated 50 to 80 percent since the 2000s. Five major genotype groups of A. astaci (A through E) have been described, with group A representing the oldest European lineage and the others associated with different North American carriers, each differing in virulence and transmission efficiency.

Croatia’s history with the pathogen stretches back to the late nineteenth century, when the first plague-related mass mortalities were recorded, with another outbreak documented in the late 1960s. Molecular confirmation came only in 2016, and that initial survey covered a limited number of localities using earlier qPCR assays with known specificity issues. To fill the gap, the new study turned to a remarkable resource: archived crayfish specimens preserved in 96 percent ethanol, collected over 27 years from both the Black Sea and Adriatic Sea drainages. The researchers analyzed 238 specimens representing all seven crayfish species present in the country, four native and three invasive.

The technical approach was meticulous. Tissue pools of eyes, abdominal cuticle, and appendage setae were dissected under bleach-decontaminated conditions to prevent cross-contamination. DNA was extracted with a modified insect kit protocol, tripling reagent volumes to fully submerge the tissue. Detection relied on a TaqMan qPCR assay targeting a 131-base-pair fragment of the pathogen’s internal transcribed spacer region, with synthetic gBlocks gene fragments serving as positive controls and contamination sentinels. Every presumptive positive was confirmed by Sanger sequencing against the GenBank database, and an exogenous internal positive control flagged any PCR inhibition from the decades-old ethanol-preserved samples.

The headline result: 91 of 238 samples, 38.2 percent, tested positive for A. astaci. The pathogen was detected in both major drainages, in all seven host species, and throughout the entire sampling period. Positives ranged from 33.8 percent in stone crayfish (Austropotamobius torrentium) to 42.3 percent in narrow-clawed crayfish (Pontastacus leptodactylus) among native species, and reached 50 percent in signal crayfish among invasives. Metadata accompanying the archived material also revealed four suspected plague outbreaks: Kotluša Stream in 2019 and the Krupa River in 2020, both involving stone crayfish; the Bregana River in 2025, involving stone crayfish again; and the Otuča River in 2019, involving noble crayfish (Astacus astacus).

Genotyping proved harder than detection, as expected. The researchers integrated three methods: a pentaplex qPCR assay distinguishing genotype groups A through E using five fluorescently labeled probes, mitochondrial haplotyping of the rnnS and rnnL ribosomal genes, and microsatellite genotyping. Only 11 of the 91 positive samples yielded interpretable consensus genotype profiles, because genotyping requires substantially higher pathogen loads than species-level detection, and archived specimens were mostly collected from apparently healthy populations where pathogen loads were low. DNA degradation from years in ethanol further reduced amplification success.

Those 11 profiles told a striking story. Genotype group A, the old European lineage, appeared in six samples across three native species and both drainages, including two outbreak localities, with the earliest record extending back to 2007. Group A is generally considered less virulent, but the study shows it can still trigger mass mortalities when hosts are stressed by pollution or elevated water temperatures. Genotype group B, typically carried by signal crayfish, was confirmed in two native species, noble crayfish and stone crayfish, including a 2025 outbreak in the Bregana River, a locality outside the signal crayfish’s known invasion range. Two previously unreported profiles also emerged: an SSR-UP-like genotype in a signal crayfish from the Drava River, the first such record in an invasive host, and a putatively novel genotype in a narrow-clawed crayfish from the Danube River that yielded discordant results across methods.

The detection of group B in native crayfish far from any known signal crayfish population is particularly alarming. Similar patterns, where NICS-associated genotypes appear in native populations years before their carriers are identified, have been reported in Switzerland, the Czech Republic, and Ireland. The researchers suggest two explanations: spillover from undetected invasive crayfish upstream, or transfer via contaminated fishing gear. Either way, the finding implies that A. astaci strains can reach highly susceptible native populations through pathways that current monitoring does not capture. The marbled crayfish (Procambarus virginalis), present in Croatia since 2013, tested positive in two samples from Šoderica Lake, but their genotypes could not be resolved, leaving open the question of whether genotype group D lurks undetected. Similarly, several positive spinycheek crayfish samples could not be genotyped, so genotype group E may also be present but invisible to current methods.

The study’s authors argue that Croatia needs a coordinated, genotype-resolved national monitoring framework, modeled on systems in Norway, Switzerland, and Ireland. Although crayfish plague has been legally notifiable in Croatia since 1997, practical reporting of wild crayfish mortality events remains limited, hampered by field observers who are not trained to recognize outbreaks and by diagnostic pathways that depend on authorized veterinarians visiting sites before carcasses degrade. The researchers call for targeted training of field personnel, integration of samples from protected-area staff and researchers into the official diagnostic pathway, continuous biosafety education for fishermen, and the adoption of environmental DNA screening as a non-invasive early-warning tool. Priority zones should include invasion fronts and contact zones between native and invasive crayfish, such as the Drava–Mura system and the Korana–Mrežnica area. As this study demonstrates, the pathogen’s true distribution may still be broader than documented, and only sustained, genotype-informed surveillance can reveal where it will strike next.

Subject of Research: Time capsules of infection: Updating the distribution of Aphanomyces astaci in Croatia using archived crayfish specimens

Article Title: Time capsules of infection: Updating the distribution of Aphanomyces astaci in Croatia using archived crayfish specimens

Article References: Škugor, T., Baudry, T., Maguire, I., Hudina, S., Jemmi, E., Delaunay, C., Pisano, S. R. R., Temunović, M., Schmidt-Posthaus, H., & Bielen, A. (2026). Time capsules of infection: Updating the distribution of Aphanomyces astaci in Croatia using archived crayfish specimens. NeoBiota, 109, 247-270. https://doi.org/10.3897/neobiota.109.200821

Image Credits: AI Generated

DOI: 10.3897/neobiota.109.200821

Keywords: Time, capsules, infection, Updating, distribution, Aphanomyces, astaci, Croatia, archived, crayfish, specimens, scientific research

Now a research team at Guangdong Haid Group has shown that a surprisingly simple environmental lever—dimming the lights—can help break that constraint. Writing in Frontiers in Physiology, the researchers report that in crayfish held at a sweltering 36 degrees Celsius, reducing light intensity to roughly half of full brightness significantly improved weight gain and survival while slightly delaying gonad development. The finding matters because gonad development in crustaceans diverts energy away from somatic growth, so any intervention that keeps animals in a growth-oriented physiological state, without invasive surgery or hormonal treatment, could translate directly into better yields for an industry worth billions.

The study rested on two linked experiments. First, the team reared juvenile female crayfish, each weighing about 6.2 grams, at six temperatures ranging from 16 to 36 degrees Celsius for 60 days, using a gradual ramping method that raised water temperature by roughly 2 degrees every 24 hours until each group reached its designated target. The temperatures were chosen to mirror the natural seasonal warming that crayfish pond water experiences from spring into summer in Hubei Province, China. Second, at the hottest temperature of 36 degrees, the researchers exposed crayfish to three light regimes created by layering shading netting over fluorescent lamps: a control at 800 to 1,000 lux representing 100 percent relative light intensity, a moderate treatment at 400 to 500 lux representing 50 percent, and a dim treatment at 150 to 200 lux representing 20 percent, all under a 12-hour light and 12-hour dark cycle designed to simulate the shade found in ponds during high-temperature periods.

The temperature experiment told a clear story of trade-offs. Crayfish weight increased significantly as water warmed, but declined above 28 degrees and dropped significantly at 36 degrees. Survival followed a similar pattern: at temperatures above 28 degrees, the survival rate fell significantly, from 96.7 percent down to 72.2 percent. Gonad development, meanwhile, accelerated with heat. Below 24 degrees, more than 85 percent of gonads remained at the non-developmental stage, with no signs of maturation. Above 28 degrees, between 10 and 30 percent of gonads were fully mature, and at 36 degrees, few undeveloped animals remained. In other words, the very temperatures that push crayfish toward sexual maturity also compromise their survival, creating a dilemma for farmers facing ever-hotter summers.

Behind these visible changes lay a molecular cascade. The researchers measured vitellogenin, or VTG, the yolk precursor protein that crustaceans synthesize and transport to developing oocytes, in the hemolymph, the crustacean equivalent of blood. VTG concentrations rose steadily with temperature and were significantly higher at 32 and 36 degrees than at lower temperatures. Gene expression of vtg in both the hepatopancreas, the digestive organ that serves as the primary VTG factory, and the ovary climbed in parallel, with expression at 32 to 36 degrees running 10 to 40 times higher than below 24 degrees. Notably, vtg expression in the hepatopancreas exceeded that in the ovary at every temperature, indicating that the hepatopancreas is the dominant source of the vitellogenin circulating in the blood.

The endocrine regulators told an equally revealing story. Crustaceans control gonad development through a unique neuroendocrine system centered on the X-organ-sinus gland complex in the eyestalk, which secretes members of the crustacean hyperglycemic hormone family. Two members are especially important: vitellogenesis-inhibiting hormone, or VIH, which suppresses yolk protein synthesis, and molt-inhibiting hormone, or MIH, which restrains molting and thus growth. Above 24 degrees, VIH expression fell significantly while MIH expression rose significantly, consistent with the observed acceleration of gonad maturation. The researchers are careful to note that these correlations do not by themselves prove causation, but the pattern aligns with earlier work on the Japanese marsh shrimp, in which gonad-inhibiting hormone expression declined as temperatures rose.

The light experiment then showed that this endocrine machinery responds to brightness as well as heat. At 50 percent relative light intensity, crayfish gained significantly more weight and survived at higher rates than controls under full light, while their gonad development was slightly lower. Nearly half of the gonads in the moderate-light group remained at early developmental stages, about twice the proportion seen in controls. The 20 percent light treatment, by contrast, produced gonad compositions similar to the control group, suggesting that the benefit of shading has a sweet spot rather than a simple dose-response. Hemolymph VTG concentrations were significantly lower in both shaded groups than in the controls, and vtg expression declined significantly in the ovary under reduced light, although hepatopancreatic vtg expression did not differ significantly between treatments.

The endocrine data under reduced light pointed in the same direction. As light intensity decreased, VIH expression in the eyestalk rose significantly, while MIH expression showed no significant change between treatments. The researchers speculate that at high temperatures, VIH in red swamp crayfish is particularly sensitive to reductions in light intensity, and that this sensitivity restrains ovarian VTG synthesis, slowing the accumulation of yolk precursor in the developing eggs. Because the ovary can synthesize VTG locally as well as receive it from the hepatopancreas, the selective decline in ovarian vtg expression under dim light suggests that shading acts on the ovary’s own contribution to vitellogenesis rather than on the systemic supply from the hepatopancreas.

The broader context sharpens the significance of these results. Crustacean farmers have long manipulated the eyestalk endocrine system to control reproduction, most commonly through eyestalk ablation, a surgical procedure that removes the source of inhibitory hormones and promotes molting and gonad development. But ablation is invasive and stressful, and the search for non-invasive alternatives has driven interest in environmental regulation. Light is known to influence the light-sensitive eyestalk tissue, and photoperiod effects on crustacean reproduction are well documented, but the specific role of light intensity has remained poorly studied. This work adds intensity to the toolkit, showing that the same neuroendocrine pathway that ablation disrupts can be nudged in the opposite direction simply by changing how bright the water is.

For an industry confronting warming waters, the practical implications are immediate. In pond culture, crayfish must be harvested before sexual maturity because from July to September, high temperatures drive many animals into reproduction, and the increasing average temperature in the Yangtze River basin year by year is expected to exacerbate the problem. The study suggests that during hot periods, farmers could reduce light intensity to around 50 percent of ambient, perhaps through shade netting similar to the polyethylene layers used in the experiment, and thereby promote growth and survival while delaying the onset of gonad development and reducing the rate of VTG accumulation in the ovaries. The authors caution that their conclusions are restricted to growth and ovarian developmental endpoints, since spawning, fecundity, and gamete quality were not measured, and the effects were demonstrated under controlled experimental conditions rather than full-scale pond systems. Still, the idea that a sheet of shade netting could rewire the endocrine balance between growth and reproduction in one of the world’s most farmed crustaceans is a striking reminder that sometimes the most powerful aquaculture interventions are not chemical or surgical at all, but simply a matter of changing the environment the animals live in.

Subject of Research: Controlled gonad development: effects of environmental factors on sexual maturity in red swamp crayfish, Procambarus clarkii

Article Title: Controlled gonad development: effects of environmental factors on sexual maturity in red swamp crayfish, Procambarus clarkii

Article References: Jiao, J.-G., Yun, B., & Qian, X.-Q. (2026). Controlled gonad development: effects of environmental factors on sexual maturity in red swamp crayfish, Procambarus clarkii. Frontiers in Physiology, 17, Article 1956361. https://doi.org/10.3389/fphys.2026.1956361

Image Credits: AI Generated

DOI: 10.3389/fphys.2026.1956361

Keywords: aquaculture, crayfish, light intensity, temperature, vitellogenin, endocrinology, Frontiers in Physiology, Controlled, gonad, development, effects, environmental