Gut pathobiome mediates behavioral and developmental disorders in biotoxin-exposed amphibians.

Emerging evidence suggests a link between alterations in the gut microbiome and adverse health outcomes in the hosts exposed to environmental pollutants. Yet, the causal relationships and underlying mechanisms remain largely undefined. Here we show that exposure to biotoxins can affect gut pathobiome assembly in amphibians, which in turn triggers the toxicity of exogenous pollutants. We used Xenopus laevis as a model in this study. Tadpoles exposed to tropolone demonstrated notable developmental impairments and increased locomotor activity, with a reduction in total length by 4.37%-22.48% and an increase in swimming speed by 49.96%-84.83%. Fusobacterium and Cetobacterium are predominant taxa in the gut pathobiome of tropolone-exposed tadpoles. The tropolone-induced developmental and behavioral disorders in the host were mediated by assembly of the gut pathobiome, leading to transcriptome reprogramming. This study not only advances our understanding of the intricate interactions between environmental pollutants, the gut pathobiome, and host health but also emphasizes the potential of the gut pathobiome in mediating the toxicological effects of environmental contaminants.

A new vaccination regimen using adenovirus-vectored vaccine confers effective protection against African swine fever virus in swine.

African swine fever (ASF) is an acute and highly contagious lethal infectious disease in swine that severely threatens the global pig industry. At present, a safe and efficacious vaccine is urgently required to prevent and control the disease. In this study, we evaluated the safety and immunogenicity of replication-incompetent type-2 adenoviruses carrying African swine fever virus (ASFV) antigens, namely CP204L (p30), E183L (p54), EP402R (CD2v), B646L (p72), and B602L (p72 chaperone). A vaccine cocktail delivered by simultaneous intramuscular (IM) and intranasal (IN) administration robustly elicited both systemic and mucosal immune responses against AFSV in mice and swine and provided highly effective protection against the circulating ASFV strain in farmed pigs. This multi-antigen cocktail vaccine was well tolerated in the vaccinated animals. No significant interference among antigens was observed. The combined IM and IN vaccination using this adenovirus-vectored antigen cocktail vaccine warrants further evaluation for providing safe and effective protection against ASFV infection and transmission.

Phyllosphere microbiome induces host metabolic defence against rice false-smut disease.

Mutualistic interactions between host plants and their microbiota have the potential to provide disease resistance. Most research has focused on the rhizosphere, but it is unclear how the microbiome associated with the aerial surface of plants protects against infection. Here we identify a metabolic defence underlying the mutualistic interaction between the panicle and the resident microbiota in rice to defend against a globally prevalent phytopathogen, Ustilaginoidea virens, which causes false-smut disease. Analysis of the 16S ribosomal RNA gene and internal transcribed spacer sequencing data identified keystone microbial taxa enriched in the disease-suppressive panicle, in particular Lactobacillus spp. and Aspergillus spp. Integration of these data with primary metabolism profiling, host genome editing and microbial isolate transplantation experiments revealed that plants with these taxa could resist U. virens infection in a host branched-chain amino acid (BCAA)-dependent manner. Leucine, a predominant BCAA, suppressed U. virens pathogenicity by inducing apoptosis-like cell death through H2O2 overproduction. Additionally, preliminary field experiments showed that leucine could be used in combination with chemical fungicides with a 50% reduction in dose but similar efficacy to higher fungicide concentrations. These findings may facilitate protection of crops from panicle diseases prevalent at a global scale.

Ethylene enhances MdMAPK3-mediated phosphorylation of MdNAC72 to promote apple fruit softening.

The phytohormone ethylene plays an important role in promoting the softening of climacteric fruits, such as apples (Malus domestica); however, important aspects of the underlying regulatory mechanisms are not well understood. In this study, we identified apple MITOGEN-ACTIVATED PROTEIN KINASE 3 (MdMAPK3) as an important positive regulator of ethylene-induced apple fruit softening during storage. Specifically, we show that MdMAPK3 interacts with and phosphorylates the transcription factor NAM-ATAF1/2-CUC2 72 (MdNAC72), which functions as a transcriptional repressor of the cell wall degradation-related gene POLYGALACTURONASE1 (MdPG1). The increase in MdMAPK3 kinase activity was induced by ethylene, which promoted the phosphorylation of MdNAC72 by MdMAPK3. Additionally, MdPUB24 functions as an E3 ubiquitin ligase to ubiquitinate MdNAC72, resulting in its degradation via the 26S proteasome pathway, which was enhanced by ethylene-induced phosphorylation of MdNAC72 by MdMAPK3. The degradation of MdNAC72 increased the expression of MdPG1, which in turn promoted apple fruit softening. Notably, using variants of MdNAC72 that were mutated at specific phosphorylation sites, we observed that the phosphorylation state of MdNAC72 affected apple fruit softening during storage. This study thus reveals that the ethylene-MdMAPK3-MdNAC72-MdPUB24 module is involved in ethylene-induced apple fruit softening, providing insights into climacteric fruit softening.

Exogenous Ca2+ promotes transcription factor phosphorylation to suppress ethylene biosynthesis in apple.

Ethylene biosynthesis in apple (Malus domestica) fruit can be suppressed by calcium ions (Ca2+) during storage; however, the underlying mechanisms are unclear. In this study, we identified the apple transcription factor MCM1-AGAMOUS-DEFICIENS-SRF5 (MdMADS5), which functions as a transcriptional activator of the ethylene biosynthesis-related gene 1-AMINOCYCLOPROPANE-1-CARBOXYLIC ACID SYNTHASE1 (MdACS1), a partner of the calcium sensor CALCIUM-DEPENDENT PROTEIN KINASES7 (MdCDPK7). Ca2+ promoted the MdCDPK7-mediated phosphorylation of MdMADS5, which resulted in the degradation of MdMADS5 via the 26S proteasome pathway. MdCDPK7 also phosphorylated 1-AMINOCYCLOPROPANE-1-CARBOXYLIC ACID OXIDASE1 (MdACO1), the key enzyme in ethylene biosynthesis, leading to MdACO1 degradation and inhibition of ethylene biosynthesis. Our results reveal that Ca2+/MdCDPK7-MdMADS5 and Ca2+/MdCDPK7-MdACO1 are involved in Ca2+-suppressed ethylene biosynthesis, which delays apple fruit ripening. These findings provide insights into fruit ripening, which may lead to the development of strategies for extending the shelf life of fruit.

Phosphorylation of MdCYTOKININ RESPONSE FACTOR4 suppresses ethylene biosynthesis during apple fruit ripening.

The plant hormone ethylene plays a central role in the ripening of climacteric fruits, such as apple (Malus domestica). Ethylene biosynthesis in apple fruit can be suppressed by calcium ions (Ca2+); however, the underlying mechanism is largely unknown. In this study, we identified an apple APETALA2/ETHYLENE-RESPONSIVE FACTOR (AP2/ERF) transcription factor, MdCYTOKININ RESPONSE FACTOR4 (MdCRF4), which functions as a transcriptional activator of ethylene biosynthesis- and signaling-related genes, including Md1-AMINOCYCLOPROPANE-1-CARBOXYLIC ACID SYNTHASE1 (MdACS1) and MdETHYLENE-RESPONSIVE FACTOR3 (MdERF3), as a partner of the calcium sensor, calmodulin. Ca2+ promoted the Ca2+/CaM2-mediated phosphorylation of MdCRF4, resulting in MdCRF4 recognition by the E3 ubiquitin ligase MdXB3 ORTHOLOG 1 IN ARABIDOPSIS THALIANA (MdXBAT31), and consequently its ubiquitination and degradation via the 26S proteasome pathway. This in turn resulted in lower expression of MdACS1 and MdERF3 and reduced ethylene biosynthesis. Transiently overexpressing various MdCRF4 proteins with specific mutated phosphorylation sites revealed that the phosphorylation state of MdCRF4 affects the ripening of apple fruit. The results reveal that a Ca2+/CaM-MdCRF4-MdXBAT31 module is involved in Ca2+-suppressed ethylene biosynthesis, which delays apple fruit ripening. This provides insights into fruit ripening that may result in strategies for extending fruit shelf life.

The molecular mechanism on suppression of climacteric fruit ripening with postharvest wax coating treatment via transcriptome.

Wax coating is an important means to maintain fruit quality and extend fruit shelf life, especially for climacteric fruits, such as apples (Malus domestica). Here, we found that wax coating could inhibit ethylene production, chlorophyll degradation, and carotenoid synthesis, but the molecular mechanism remains unclear. The regulatory mechanism of wax coating on apple fruit ripening was determined by subjecting wax-treated apple fruits to transcriptome analysis. RNA-seq revealed that 1,137 and 1,398 genes were upregulated and downregulated, respectively. These differentially expressed genes (DEGs) were shown to be related to plant hormones, such as ethylene, auxin, abscisic acid, and gibberellin, as well as genes involved in chlorophyll degradation and carotenoid biosynthesis. Moreover, we found that some genes related to the wax synthesis process also showed differential expression after the wax coating treatment. Among the DEGs obtained from RNA-seq analysis, 15 were validated by quantitative RT-PCR, confirming the results from RNA-seq analysis. RNA-seq and qRT-PCR of pear (Pyrus ussuriensis) showed similar changes after wax treatment. Our data suggest that wax coating treatment inhibits fruit ripening through ethylene synthesis and signal transduction, chlorophyll metabolism, and carotenoid synthesis pathways and that waxing inhibits endogenous wax production. These results provide new insights into the inhibition of fruit ripening by wax coating.

Temporal metabolite responsiveness of microbiota in the tea plant phyllosphere promotes continuous suppression of fungal pathogens.

INTRODUCTION: A broad spectrum of rhizosphere bacteria and fungi were shown to play a central role for health, fitness and productivity of their host plants. However, implications of host metabolism on microbiota assembly in the phyllosphere and potential consequences for holobiont functioning were sparsely addressed. Previous observations indicated that tea plants might reduce disease occurrence in various forests located in their proximity; the underlying mechanisms and potential implications of the phyllosphere microbiota remained elusive. OBJECTIVES: This study aimed atdeciphering microbiome assembly in the tea plant phyllosphere throughout shoot development as well as elucidating potential implications of host metabolites in this process. The main focus was to explore hidden interconnections between the homeostasis of the phyllosphere microbiome and resistance to fungal pathogens. METHODS: Profiling of host metabolites and microbiome analyses based on high-throughput sequencing were integrated to identify drivers of microbiome assembly throughout shoot development in the phyllosphere of tea plants. This was complemented by tracking of beneficial microorganisms in all compartments of the plant. Synthetic assemblages (SynAss), bioassays and field surveys were implemented to verify functioning of the phyllosphere microbiota. RESULTS: Theophylline and epigallocatechin gallate, two prevalent metabolites at the early and late shoot development stage respectively, were identified as the main drivers of microbial community assembly. Flavobacterium and Myriangium were distinct microbial responders at the early stage, while Parabacteroides and Mortierella were more enriched at the late stage. Reconstructed, stage-specific SynAss suppressed various tree phytopathogens by 13.0%-69.3% in vitro and reduced disease incidence by 8.24%-41.3% in vivo. CONCLUSION: The findings indicate that a functional phyllosphere microbiota was assembled along with development-specific metabolites in tea plants, which continuously suppressed prevalent fungal pathogens. The insights gained into the temporally resolved metabolite response of the tea plant microbiota could provide novel solutions for disease management.

NUCLEOCYTOPLASMIC shuttling of ETHYLENE RESPONSE FACTOR 5 mediated by nitric oxide suppresses ethylene biosynthesis in apple fruit.

Nitric oxide (NO) is known to modulate the action of several phytohormones. This includes the gaseous hormone ethylene, but the molecular mechanisms underlying the effect of NO on ethylene biosynthesis are unclear. Here, we observed a decrease in endogenous NO abundance during apple (Malus domestica) fruit development and exogenous treatment of apple fruit with a NO donor suppressed ethylene production, suggesting that NO is a ripening suppressor. Expression of the transcription factor MdERF5 was activated by NO donor treatment. NO induced the nucleocytoplasmic shuttling of MdERF5 by modulating its interaction with the protein phosphatase, MdPP2C57. MdPP2C57-induced dephosphorylation of MdERF5 at Ser260 is sufficient to promote nuclear export of MdERF5. As a consequence of this export, MdERF5 proteins in the cytoplasm interacted with and suppressed the activity of MdACO1, an enzyme that converts 1-aminocyclopropane-1-carboxylic acid (ACC) to ethylene. The NO-activated MdERF5 was observed to increase in abundance in the nucleus and bind to the promoter of the ACC synthase gene MdACS1 and directly suppress its transcription. Together, these results suggest that NO-activated nucleocytoplasmic MdERF5 suppresses the action of ethylene biosynthetic genes, thereby suppressing ethylene biosynthesis and limiting fruit ripening.

Exogenous gibberellin treatment improves fruit quality in self-pollinated apple.

Although a few apple (Malus × ×domestica) varieties are self-compatible, little is known about the differences in fruit quality between self- and cross-pollinated apple. In our current study, we compared the fruit quality of self-pollinated apple plants (cultivar 'Hanfu') in self-pollination or cross-pollinated by another cultivar 'Qinguan'. Analysis of fruit quality revealed substantial differences in the external qualities between self- and cross-pollinated apple fruit, but not in the internal qualities. Fruits harvested from self-pollinated 'Hanfu' were smaller and more asymmetrical than those harvested from the cross-pollinated plants. We developed a mathematical model describing how seed number and distribution affect fruit growth. According to this model, the fewer the seeds, the greater the force released from the seeds and the more asymmetrical the fruit. Detection of endogenous hormone and the associated gene expression revealed that gibberellin (GA) levels and GA transporter gene expression on the seedless side were significantly lower than those on the seeded side. Analysis of fruit pectin methylesterase activity and demethylated pectin levels indicated that the lack of GA limits fruit cell wall extension. Additionally, spraying the self-pollinating plants with gibberellic acid increased the fruit weight and lowered the proportion of asymmetrical fruit, recovering the exterior fruit quality to that of the cross-pollinated fruit. Furthermore, exogenous GA treatment increased the wax layer thickness and reduced the fruit water loss rate, leading to a dramatic improvement in fruit storage capacity. Therefore, exogenous GA treatment could be used to ensure regular fruit production of self-pollinated 'Hanfu'.

Polyamines Involved in Regulating Self-Incompatibility in Apple.

Apple exhibits typical gametophytic self-incompatibility, in which self-S-RNase can arrest pollen tube growth, leading to failure of fertilization. To date, there have been few studies on how to resist the toxicity of self-S-RNase. In this study, pollen tube polyamines were found to respond to self-S-RNase and help pollen tubes defend against self-S-RNase. In particular, the contents of putrescine, spermidine, and spermine in the pollen tube treated with self-S-RNase were substantially lower than those treated with non-self-S-RNase. Further analysis of gene expression of key enzymes in the synthesis and degradation pathways of polyamines found that the expression of DIAMINE OXIDASE 4 (MdDAO4) as well as several polyamine oxidases such as POLYAMINE OXIDASES 3 (MdPAO3), POLYAMINE OXIDASES 4 (MdPAO4), and POLYAMINE OXIDASES 6 (MdPAO6) were significantly up-regulated under self-S-RNase treatment, resulting in the reduction of polyamines. Silencing MdPAO6 in pollen tubes alleviates the inhibitory effect of self-S-RNase on pollen tube growth. In addition, exogenous polyamines also enhance pollen tube resistance to self-S-RNase. Transcriptome sequencing data found that polyamines may communicate with S-RNase through the calcium signal pathway, thereby regulating the growth of the pollen tubes. To summarize, our results suggested that polyamines responded to the self-incompatibility reaction and could enhance pollen tube tolerance to S-RNase, thus providing a potential way to break self-incompatibility in apple.

Keystone taxa-mediated bacteriome response shapes the resilience of the paddy ecosystem to fungicide triadimefon contamination.

The increasing input of fungicides has emerged as a global concern for agroecosystem stability and sustainability. Agroecosystem resilience has been linked to microbiome response, however, is not well understood. Focusing on a widespread triazole-class fungicide triadimefon in the paddy ecosystem, we characterized that the soils and sediments were dominant triadimefon reservoirs with the peak level at 195 µg kg-1 and 31.3 µg kg-1, respectively, but essential for the resilience of paddy ecosystem to triadimefon. In paddy simulation models, the half-life of triadimefon in soil-sediment was 8.4-28.9 days, while it was prolonged to 86.6-115.5 days after elimination of resident microbial community. Phospholipid fatty acid profiling and high-throughput sequencing showed that the distinctive bacterial community responses contributed to variable degradation of triadimefon in paddy soils and sediments. Sphingomonas and Xanthomonas were identified as positive responders of the keystone taxa in the responsive bacteriome, whereas Enterobacter were negative responders that declined over time. Synthetic assemblages combined with quantitative polymerase chain reaction further validated that Sphingomonas and Xanthomonas were involved in sustaining soil-sediment resilience to triadimefon contamination. Collectively, our results revealed that the shaping of soil and sediment bacteriomes was responsible for the resilience of the paddy agroecosystem to fungicide contamination.

Screening and functional verification of the target protein of pedunsaponin A in the killing of Pomacea canaliculata.

Previous study found that pedunsaponin A (PA) influenced the cytoskeleton of Pomacea canaliculata hemocytes, leading to depolarization and haemocyte destruction and eventually to snail death. In this study, we analysed the changes in protein expression by iTRAQ-mediated proteomics and identified 51 downregulated proteins. Among these, we focused on proteins related to cytoskeletal function and identified neural Wiskott-Aldrich syndrome isoform X1 (PcnWAS). The full-length PcnWAS gene contains 9791 bp and includes an open reading frame of 1401 bp that encodes 735 amino acids with a predicted molecular mass of 49.83 kD. PcnWAS exhibited a relatively distant genetic relationship with known species; the closest homologue is Biomphalaria glabrata (57%). RNA interference (RNAi) was adopted to verify the function of PcnWAS after screening the siRNA sequence with an efficiency of 97%. Interference with the gene expression of PcnWAS did not lead to snail death, but the depolarization level increased, which demonstrated that PcnWAS is an important depolarization-related protein. The results of PA treatment of snails subjected to RNAi proved that interfering with PcnWAS gene expression decreased the molluscicidal activity of PA toward P. canaliculata; snail mortality after RNAi was significantly lower (40%) than that in PA-treated snails without RNAi (54%), while the survival rate and depolarization level in haemocytes were not significant, indicating that PcnWAS is only one of the important target proteins of PA in P. canaliculata. This study lays the foundation for further exploration of the molecular mechanism by which PA kills this harmful snail.

Mycoplasma ovipneumoniae-derived lipid-associated membrane proteins induce cytokine secretion in mouse peritoneal macrophages through TLR2 signalling.

BACKGROUND: Mycoplasma ovipneumoniae (M. ovi) is the causative agent of chronic non-progressive pneumonia in sheep, goats, bighorn, and wild small ruminants. However, the mechanism of infection and immune response to M. ovi remain unclear. Invading microbes express lipid-associated membrane proteins (LAMPs) on the cell surface that interact with host cells to facilitate infection, and are thus the major molecules recognised by the host immune system. Upon LAMP recognition, Toll-like receptor 2 (TLR2) and NLRP3 inflammasome sense the pathogens and signalling pathways for cytokine secretion. In this study, we investigated whether M. ovi and M. ovi-derived LAMPs are immuno-biologically active compounds capable of activating mouse peritoneal macrophages and explored the underlying mechanism. RESULTS: After infection of wild-type mice with M. ovi, the expression of TLR2 and NLRP3 at the transcriptional and translational levels was determined with reverse transcription-polymerase chain reaction and flow cytometry. In addition, the cytokine levels and associated pathways were detected in infected wild-type, Tlr2-/-, and Nlrp3-/- mice via enzyme-linked immunosorbent assays and western blotting. The nuclear factor (NF)-κB and mitogen-activated protein kinase (MAPK) signalling pathways were found to mediate the expression of inflammatory cytokines in M. ovi or M. ovi-derived LAMP-infected peritoneal macrophages, and cytokines were not induced in Tlr2-/- and/or Nlrp3-/- macrophages. CONCLUSION: Host cytokine production is activated in response to M. ovi-derived LAMPs through the NF-κB and MAPK signalling pathway via TLR2.

Auxin-activated MdARF5 induces the expression of ethylene biosynthetic genes to initiate apple fruit ripening.

The gaseous plant hormone ethylene induces the ripening of climacteric fruit, including apple (Malus domestica). Another phytohormone, auxin, is known to promote ethylene production in many horticultural crops, but the regulatory mechanism remains unclear. Here, we found that auxin application induces ethylene production in apple fruit before the stage of commercial harvest, when they are not otherwise capable of ripening naturally. The expression of MdARF5, a member of the auxin response factor transcription factor (TF) family involved in the auxin signaling pathway, was enhanced by treatment with the synthetic auxin naphthaleneacetic acid (NAA). Further studies revealed that MdARF5 binds to the promoter of MdERF2, encoding a TF in the ethylene signaling pathway, as well as the promoters of two 1-aminocyclopropane-1-carboxylic acid synthase (ACS) genes (MdACS3a and MdACS1) and an ACC oxidase (ACO) gene, MdACO1, all of which encode key steps in ethylene biosynthesis, thereby inducing their expression. We also observed that auxin-induced ethylene production was dependent on the methylation of the MdACS3a promoter. Our findings reveal that auxin induces ethylene biosynthesis in apple fruit through activation of MdARF5 expression.

Two new compounds from the roots of Pueraria peduncularis and their molluscicidal effects on Pomacea canaliculata.

Two oleanane-type triterpenoid saponins named pedunsaponin D (1) and pedunsaponin E (2) were isolated from the roots of Pueraria peduncularis. The structures of the new compounds were elucidated based on chemical and physicochemical evidence as follows: pedunsaponin D, 3-O-ß-glucopyranosyl-(1-3)-ß-glucuronopyranosyl-3ß,15α,23α-trihydroxy-11,13(18)-oleanadien-16-one (1); pedunsaponin E, 3-O-ß-glucopyranosyl-(1-2)-ß-glucopy ranosyl(1-2)[ß-glucopyranosyl(1-3)-ß-glucuronopyranosyl]-3ß-hydroxy-16-oxoolean-12-en-30-oic acid (2). The two compounds showed moderate molluscicidal activity.[Formula: see text].

iTRAQ-based proteomic analysis of Mycoplasma bovis NM-28 strain from two generations for vaccine screening.

Mycoplasma bovis is an important pathogenic bacterium affecting cows and cattle. Clinically, an inactivated vaccine of M. bovis is mainly used to prevent infection by this bacterium. The changes that occur in the antigen when M. bovis is continuously passaged in vitro remain unknown. Therefore, we performed an in vitro serial passage of the M. bovis NM-28 strain, which was isolated and identified in our laboratory. An isobaric tags for relative and absolute quantitation (iTRAQ)-based quantitative proteomics method was used to analyse the differences between generations 3 and 60. Many major membrane proteins or protective antigens reported in the literature did not exhibit changes between these generations. We found an imbalance between growth rate and nutrition in the 60th generation. The proteomics results were verified by western blotting and real-time PCR. Growth curves were also prepared based on colony-forming units (CFUs) between the 3rd and 60th generations. The number of colonies in the 60th generation in the stationary phase was 5 × 109 CFU mL-1, which was 10-fold higher than that in the 3rd generation. The 60th generation of the NM-28 strain can be used as an inactivated vaccine strain of M. bovis to lower production costs compared to use of the 3rd generation.

Study of the Differentially Expressed Genes in the Pomacea canaliculata Transcriptome after Treatment with Pedunsaponin A.

Transcriptomes, genomes, and proteomes have played important roles in the search for drug targets. To determine the molluscicidal mechanism of pedunsaponin A against Pomacea canaliculata, RNA-seq technology was adopted to analyze the differentially expressed genes (DEGs) in the P. canaliculata transcriptome after treatment with pedunsaponin A. As a result, 533 DEGs were identified, among which 255 genes were significantly upregulated and 278 genes were significantly downregulated. According to the analysis of Gene Ontology (GO) functions, we found that the DEGs were significantly enriched in the viral life cycle, UDP-glucose 4-epimerase activity, guanylate cyclase activity, the cyclic guanosine monophosphate (cGMP) biosynthetic process, and the cGMP metabolic process. The Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway results showed that the DEGs were mainly involved in the hedgehog signaling pathway, phagosome, cytosolic DNA-sensing pathway, retinoic acid-inducible gene I like (RIG-I-like) receptor signaling pathway, bacterial secretion system, and nuclear factor-kappa B (NF-kappa B) signaling pathway. The above results indicated that pedunsaponin A causes a metabolic disorder, anomalous opening of membrane ion channels, and an imbalance in osmotic pressure between the interior and exterior of cells, eventually resulting in the death of cells involved in immune defense and influencing the immune response of P. canaliculata.

The Damaging Effects of Pedunsaponin A on Pomacea canaliculata Hemocytes.

Pomacea canaliculata hemocytes are the main functional cells in the immune defense system, and hemocyte destruction disrupts the immune response mechanism of P. canaliculata, resulting in abnormal growth, development, reproduction, and even death. Our previous study found that Pedunsaponin A significantly affects P. canaliculata hemocyte structure. This study further investigated the damaging effects of Pedunsaponin A on P. canaliculata hemocytes. The cell mortality rate results showed that the hemocyte mortality was significantly increased after treatment with Pedunsaponin A, and the mortality rate exhibited a significant positive correlation with treatment time and dose. The membrane potential results showed that the cell membranes of P. canaliculata hemocytes exhibited time-dependent membrane depolarization after 40 mg/L Pedunsaponin A treatment. At 36 h, the cell depolarization rate in the Pedunsaponin A treatment group was 41.43%, which was significantly greater than the control group (6.24%). The cytoskeleton results showed that Pedunsaponin A led to disordered and dispersed arrangement of microfilaments and changes in the cytoskeletal structure. The apoptosis and cell cycle results showed that Pedunsaponin A induced apoptosis and influenced the cell cycle to some extent. These results showed that the cell membrane and cytoskeleton of P. canaliculata hemocytes were damaged after treatment with Pedunsaponin A, which led to an increase in cell mortality, dysfunction, cell cycle abnormalities and apoptosis. This study provides a foundation for further identification of the site of Pedunsaponin A activity on hemocytes.