# Genome-Wide Association Analysis and Candidate Gene Identification for Resistance to “Milky Disease” in the Chinese Mitten Crab (Eriocheir sinensis)

## TL;DR

Researchers identified genetic loci linked to resistance against milky disease in Chinese mitten crabs, offering insights for breeding disease-resistant strains.

## Contribution

The study reveals a novel mechanism where crabs may induce the pathogen into a nonculturable state, alongside identifying key genetic markers.

## Key findings

- The F05 crab family showed strong resistance with only 3% mortality and normal growth after infection.
- Genome-wide analysis identified 767 loci significantly associated with disease resistance traits.
- Resistant crabs may induce the pathogen into a viable but nonculturable state, a first-time observation in this context.

## Abstract

The Chinese mitten crab (Eriocheir sinensis) is an important aquaculture species in China. However, “milky disease” caused by Metschnikowia bicuspidata poses a serious threat to crab health and leads to substantial mortality. To support the development of disease-resistant crab strains, we evaluated disease resistance across 10 crab families. The F05 family exhibited the highest resistance, with a post-infection mortality rate of only 3%, while maintaining normal molting and growth performance. Genome-wide analyses identified 767 loci significantly associated with disease resistance. Notably, our results suggest, for the first time, that resistant crabs may mitigate disease progression by inducing the pathogen into a viable but nonculturable (VBNC) state (metabolically active but non-culturable on conventional media). These findings provide a genetic and mechanistic foundation for selective breeding programs aimed at reducing disease-related losses and improving aquaculture efficiency.

“Milky disease” of the Chinese mitten crab (Eriocheir sinensis), caused by Metschnikowia bicuspidata, leads to substantial economic losses. Despite extensive research on its pathogenesis, the genetic basis of host resistance and underlying regulatory mechanisms remain unclear, limiting the development of disease-resistant varieties. This study aimed to (1) evaluate resistance differences among 10 E. sinensis families (five highly resistant and five sensitive) via artificial challenge; (2) identify disease-resistance-associated genetic loci using genome-wide association analysis (GWAS). Our findings revealed that the F05 family exhibited the strongest resistance, with a mortality rate of only 3% and a molting rate of 73%, accompanied by superior growth performance. Whole-genome resequencing identified 10,161,545 high-quality SNPs, and GWAS detected 767 loci significantly associated with disease-resistance traits, among which two pleiotropic SNPs (Chr46:18395778 and Chr1:20680490) were simultaneously associated with both “dead or not” and “qPCR fixed amount of fungi”, reflecting their functional relevance in regulating survival and pathogen load. Notably, we propose for the first time that E. sinensis achieves resistance by inducing M. bicuspidata into a viable but nonculturable (VBNC) state, in which the fungal cells remain metabolically active but cannot form colonies on conventional culture media. This study establishes a multidimensional resistance evaluation system, clarifies the genetic basis and novel mechanism of resistance, and provides valuable molecular markers for marker-assisted breeding. The findings contribute to reducing disease-related losses and promoting sustainable development of the E. sinensis aquaculture industry.