Microsporidia are highly specialized obligate intracellular parasites that can infect a wide variety of animals ranging from protists to mammals. The classical concept of the parasite invasion into a host cell involves its polar tube acting as a needle-syringe system. However, recent studies show microsporidian spores can also gain access to host cells by phagocytosis. The present study investigated the phagocytic uptake process of causative agent of the pebrine disease, Nosema bombycis, in several insect cell lines. We observed KOH-treated spores and cold-storaged spores can be easily uptaken by all the studied cell types 4 h post inoculation. In contrast, large numbers of freshly recovered spores remained in the culture medium. To further investigate the intracellular fates of KOH-treated spores and cold-storaged spores, electron and fluorescence microscopy were performed. No intracellular germination or subsequent parasite development were observed. Intracellular spores can be detected in host cells by polyclonal antibody 7 d post inoculation, suggesting phagocytized N. bombycis could not be digested by these non-professional phagocytes. Our results suggest that, phagocytic uptake of N. bombycis spores might represent a defense mechanism of the host cells and the intact spore wall barrier enable freshly recovered spores to keep resistance to this mechanism.

Prohexadione-calcium (Pro-Ca) has been shown to positively regulate crop tolerance to saline-alkali stress.  However, the optimal concentration for Pro-Ca application and the mechanisms through which it enhances saline-alkali tolerance and yield in soybean remain unclear.  This study aimed to determine the optimal concentration of exogenously applied Pro-Ca and revealed the mechanisms underlying Pro-Ca’s effect on remediation and yield response in soybean under saline-alkali stress.  The results indicated that saline-alkali stress negatively impacted the morphological and physiological traits of soybean seedlings by triggering the production of reactive oxygen species (ROS), leading to oxidative damage of the grana lamellae due to excessive accumulation of Na⁺.  An application of 100 mg L⁻¹ Pro-Ca was found to be optimal, promoting dry matter accumulation and normalized difference vegetation index (NDVI) by significantly reducing Na⁺ uptake under saline-alkali stress.  Moreover, integrated physiological, ultrastructural, and transcriptomic analyses indicated that Pro-Ca significantly enhanced the ascorbate-glutathione (AsA-GSH) cycle by up-regulating the expression of related genes to enhance the activities of ascorbate peroxidase (APX), glutathione reductase (GR), dehydroascorbate reductase (DHAR), monodehydroascorbate reductase (MDHAR), and the AsA/DHA and GSH/GSSG ratios to quench ROS, thereby protecting both thylakoid and mitochondrial membrane from degradation.  The differentially expressed genes (DEGs) encoding ascorbate and aldarate metabolism were significantly (P<0.05) enriched in the integral component of membrane.  Furthermore, Pro-Ca treatment up-regulated the expression of genes encoded photosystems under saline-alkali stress, which reduced the photoinhibition and stomatal limitation (L_s) and mitigating damage photosystem and preventing yield reduction.  In summary, foliar application of Pro-Ca could efficiently enhance soybean seedlings tolerance to saline-alkali stress by inhibiting Na⁺ influx, enhancing the AsA-GSH cycle, maintaining biomembrane system, and improving photosynthetic efficiency.