Toxicity and enzymatic mechanism of Citrus spp. essential oils and major constituents on Haemaphysalis longicornis (Acari: Ixodidae) and non-target Harmonia axyridis (Coleoptera: Coccinellidae).

Plant essential oils (EOs)-based acaricides have been recognized as environmentally-friendly alternatives to synthetic acaricides because of their low toxicity against non-target species. Despite this, there are knowledge gaps regarding the toxicity mechanisms of plant EOs against non-target species. Here, the toxicology and enzymatic mechanism of Citrus reticulata and Citrus lemon EOs were evaluated against the vector pest, Haemaphysalis longicornis, and non-target ladybird beetle, Harmonia axyridis. Both EOs were mainly composed of d-Limonene, followed by ß-Myrcene and γ-Terpinene in C. reticulata, and (-)-ß-Pinene and γ-Terpinene in C. lemon. Citrus reticulata and C. lemon EOs were toxic to Hae. longicornis, with 50 % lethal concentration (LC50) values estimated at 0.43 and 0.98 µL/mL via nymphal immersion test, and 42.52 and 46.38 µL/mL via spray application, respectively. Among the constituents tested, ß-Myrcene was the most effective, with LC50 values of 0.17 and 47.87 µL/mL via immersion and spray treatment, respectively. A significant mortality of non-target Har. axyridis was found when treated by the EOs at concentrations two times greater than LC50 estimated against H. longicornis. The biochemical assay revealed that the EOs induced changes in the antioxidant enzyme activity of superoxide dismutases, catalase, and glutathione peroxidase in Hae. longicornis and Har. axyridis. The results demonstrated the acaricidal potential of citrus EOs and their major constituents for tick control, revealed the risk of the EOs to non-target species, and provided relevant insights into the mechanisms underlying their toxicity.

Stochastic processes drive the dynamic assembly of bacterial communities in Salix matsudana afforested soils.

Introduction: This study investigates the dynamic shifts in soil bacterial communities within a Salix matsudana afforested ecosystem transitioning from agricultural land. Understanding the temporal variability in bacterial diversity and community structures is crucial for informing forest management and conservation strategies, particularly in regions undergoing afforestation. Methods: We employed high-throughput sequencing across three distinct months (August, September, and October) to analyze the temporal variability in bacterial community composition and diversity. Network analysis was utilized to identify keystone species and assess community stability under varying environmental conditions, including fluctuations in temperature and precipitation. Results: We uncover significant temporal variability in bacterial diversity and community structures, which are closely tied to fluctuations in temperature and precipitation. Our findings reveal the abundance of the dominant bacterial phyla, such as Actinobacteria and Proteobacteria, which did not change overall, highlighting the stability and resilience of the microbial community across seasonal transitions. Notably, the increasing similarity in community composition from August to October indicates a reduction in species turnover, likely driven by more homogeneous environmental conditions. Through comprehensive network analysis, we identify the pivotal role of keystone species, particularly the human pathogen Nocardia, in maintaining community stability under reduced soil moisture. The observed variations in community connectivity underscore the microbial community's resilience and adaptability to seasonal shifts, with higher stability in August and October contrasting with the instability observed in September. Discussion: These results underscore the complex interplay between stochastic and deterministic processes in bacterial community assembly, significantly shaped by prevailing environmental conditions. The insights gained from this research have far-reaching implications for forestry management and conservation strategies, particularly in regions undergoing similar afforestation efforts.

Seasonal variations affect the ecosystem functioning and microbial assembly processes in plantation forest soils.

While afforestation mitigates climate concerns, the impact of afforestation on ecological assembly processes and multiple soil functions (multifunctionality) in afforested areas remains unclear. The Xiong'an New Area plantation forests (Pinus and Sophora forests) in North China were selected to examine the effects of plantation types across four distinct seasons on soil microbiomes. Three functional categories (nutrient stocks, organic matter decomposition, and microbial functional genes) of multifunctionality and the average (net) multifunctionality were quantified. All these categories are directly related to soil functions. The results showed that net soil multifunctionality as a broad function did not change seasonally, unlike other narrow functional categories. Bacterial communities were deterministically (variable selection and homogenous selection) structured, whereas the stochastic process of dispersal limitation was mainly responsible for the assembly and turnover of fungal and protist communities. In Pinus forests, winter initiates a sudden shift from deterministic to stochastic processes in bacterial community assembly, accompanied by decreased Shannon diversity and heightened nutrient cycling (nutrient stocks and organic matter decomposition). This indicates the potential vulnerability of deterministic assembly to seasonal fluctuations, particularly in environments rich in nutrients. The results predicted that protist community composition was uniquely structured with C-related functional activities relative to bacterial and fungal ß-diversity variations, which were mostly explained by seasonal variations. Our study highlighted the importance of the protist phagocytosis process on soil microbial interactions through the predicted impact of protist α-diversity on microbial cooccurrence network parameters. This association might be driven by the high abundance of protist consumers as the main predators of bacterial and fungal lineages in our sampling plots. Our findings reveal that the complexity of microbial co-occurrence interactions was considerably higher in spring, perhaps attributing thermal variability and increased resource availability within spring that foster microbial diversity and network complexity. This study contributes to local ecosystem prospects to model the behavior of soil biota seasonally and their implied effects on soil functioning and microbial assembly processes, which will benefit global-scale afforestation programs by promoting novel, precise, and rational plantation forests for future environmental sustainability and self-sufficiency.

Cross-Lagged Panel Networks of Sleep Inertia Across Its Distinct Change Patterns Among Intern Nurses with Shift Work in China.

Purpose: Although experimental psychopathology using PET, EEG, and fMRI is at the forefront of understanding the underlying mechanisms of sleep inertia, many questions concerning causality remain unanswerable due to ethical constraints and the use of small and heterogeneous samples in experimental methods. There is a pressing need for a novel perspective in a large and relatively homogeneous population to fully capture and elucidate longitudinal processes and dynamic causality that culminate in episodes of sleep inertia over time. Therefore, this study aimed to reveal the causal relationships between symptoms of sleep inertia across its distinct patterns. Patients and Methods: A total of 1636 intern nurses participated in the first survey (94.1% validity rate), then 1277 intern nurses were followed up (82.9% tracing rate). Symptoms of sleep inertia were self-reported using the Sleep Inertia Questionnaire. The cross-lagged panel network models were used to examine unique longitudinal relationships between symptoms of sleep inertia across distinct trajectories. Results: Four distinct trajectories of sleep inertia were established. Additionally, we found differences in those symptoms with the highest influence on other symptoms at the subsequent point across the networks of four trajectories, particularly, "Difficulty in concentrating" in the persistent-high group and "Feeling tense" in the deteriorating groups. Conclusion: The current study highlights changes in sleep inertia based on the long-term course over time. Notably, symptoms of "Difficulty in concentrating" and "Feeling tense" are imperative to address these specific symptoms within subpopulations.

Lysine Methylation and Histone Modifications during Cold Stress of Insects: Freeze-Tolerant Eurosta solidaginis and Freeze-Avoiding Epiblema scudderiana.

Overwintering survival by insects, whether of the freeze-tolerant or freeze-avoiding types, is typically associated with a strong suppression of metabolic rate (e.g., entry into diapause) that involves the differential expression of many genes with regulation at the transcriptional, translational or post-translational levels. Epigenetic modifications have been suggested to play a vital role in regulating cold responses of insects. However, knowledge of the roles of epigenetic mechanisms in modulating gene expression for winter survival of the larvae of two goldenrod gall formers, the freeze-tolerant dipteran Eurosta solidaginis and the freeze-avoiding lepidopteran Epiblema scudderiana, remain unknown. The current study evaluates the role of cold-induced lysine methylation and histone modifications, with enzymes of lysine methylation (SETD8, SETD7, SUV39H1, SMYD2 and ASH2L), as well as relative levels of histone H3 acetylation (H3K9ac, H3K18ac, H3K27ac, H3K56ac) and methylation (H3K4me1, H3K9me3, H3K36me2) examined in two insects. Significant (p < 0.05) reductions were observed in most of the targets of histone methylation/acetylation for decreasing temperatures of Ep. scudderiana larvae, whereas selected histone methylation/acetylation targets were conversely elevated (p < 0.05) in E. solidaginis, particularly under conditions of 5 °C for 4 h. Histone H3 expression was found to be variable without statistical differences in larval goldenrod gall moths and gall flies. These results provide basic information on the patterns of epigenetic regulation involved in insect cold hardiness.

Redox-modulated SNX25 as a novel regulator of GPCR-G protein signaling from endosomes.

GPCR-G protein signaling from endosomes plays a crucial role in various physiological and pathological processes. However, the mechanism by which endosomal G protein signaling is terminated remains largely unknown. In this study, we aimed to investigate the regulatory mechanisms involved in terminating the signaling of Gα subunits from endosomes. Through structural analysis and cell-based assays, we have discovered that SNX25, a protein that targets endosomes via its PXA or PXC domain, interacts with regulator of G protein signaling (RGS) proteins (including RGS2, RGS4, RGS8, and RGS17) in a redox-regulated manner. The interaction between SNX25 and these RGS proteins enhances their GTPase-accelerating activity towards Gαi/q and their ability to bind GDP-bound (inactive form) Gαi/q. As a result, SNX25 recruits these RGS proteins to endosomes, leading to the termination of endosomal Gαi/q signaling. Furthermore, we have found that the SNX25/RGS complex also exerts a negative regulatory effect on Gαi/q signaling from the plasma membrane. This is achieved by recruiting Gαi/q to endosomes and preventing its activation on the plasma membrane. Our findings shed light on the previously unknown role of redox-modulated SNX25 in inhibiting Gαi/q signaling, thereby uncovering a novel mechanism for terminating Gαi/q signaling from endosomes. Importantly, this study expands our understanding of the regulation of GPCR-Gαi/q signaling beyond the plasma membrane.

Perceived school bullying and psychotic-like experiences in sexual minority adolescents: the mediating and moderating roles of resilience.

AIMS: This two-wave, longitudinal study aimed to examine the potential moderating and mediating effects of resilience on the association between perceived school bullying and psychotic-like experiences among Chinese sexual minority adolescents. METHODS: A total of 4192 senior high students were included and 984 (23.5%) of them were identified as a sexual minority (mean age = 16.68 years, SD = 0.71). Participants completed two online surveys during April 21 to May 12, 2021 and December 17 to 26, 2021, respectively, as well as completed self-report measures of sample characteristics, perceived school bullying, resilience, and psychotic-like experiences (including two dimensions: delusional experiences and hallucinatory experiences). RESULTS: Perceived school bullying and resilience were associated with psychotic-like experiences in sexual minority adolescents. Resilience mediated the relationship between perceived school bullying and subsequent psychotic-like experiences (b = 0.03, 95% CI = 0.01 ~ 0.04)/ delusional experiences (b = 0.03, 95% CI = 0.01 ~ 0.04)/ hallucinatory experiences (b = 0.02, 95% CI = 0.01 ~ 0.03). Additionally, resilience only moderated the associations of perceived school bullying with hallucinatory experiences (b = -0.06, 95% CI = -0.12 ~ -0.01). CONCLUSIONS: These findings indicated that resilience plays a crucial role in mediating or moderating the relationship between perceived school bullying and psychotic-like experiences. Assessing and reducing school bullying, as well as promoting resilience, may have important clinical implications for reducing the risk of psychotic-like experiences in sexual minority adolescents.

Molecular characterization and induced changes of histone acetyltransferases in the tick Haemaphysalis longicornis in response to cold stress.

BACKGROUND: Epigenetic modifications of histones play important roles in the response of eukaryotic organisms to environmental stress. However, many histone acetyltransferases (HATs), which are responsible for histone acetylation, and their roles in mediating the tick response to cold stress have yet to be identified. In the present study, HATs were molecularly characterized and their associations with the cold response of the tick Haemaphysalis longicornis explored. METHODS: HATs were characterized by using polymerase chain reaction (PCR) based on published genome sequences, followed by multiple bioinformatic analyses. The differential expression of genes in H. longicornis under different cold treatment conditions was evaluated using reverse transcription quantitative PCR (RT-qPCR). RNA interference was used to explore the association of HATs with the cold response of H. longicornis. RESULTS: Two HAT genes were identified in H. longicornis (Hl), a GCN5-related N-acetyltransferase (henceforth HlGNAT) and a type B histone acetyltransferase (henceforth HlHAT-B), which are respectively 960 base pairs (bp) and 1239 bp in length. Bioinformatics analysis revealed that HlGNAT and HlHAT-B are unstable hydrophilic proteins characterized by the presence of the acetyltransferase 16 domain and Hat1_N domain, respectively. RT-qPCR revealed that the expression of HlGNAT and HlHAT-B decreased after 3 days of cold treatment, but gradually increased with a longer period of cold treatment. The mortality rate following knockdown of HlGNAT or HlHAT-B by RNA interference, which was confirmed by RT-qPCR, significantly increased (P < 0.05) when H. longicornis was treated at the lowest lethal temperature (- 14 °C) for 2 h. CONCLUSIONS: The findings demonstrate that HATs may play a crucial role in the cold response of H. longicornis. Thus further research is warranted to explore the mechanisms underlying the epigenetic regulation of the cold response in ticks.

Coronary Artery Fistula and Severe Coronary Artery Stenosis: A Case Report and an Insight for Potential Pathogenesis of Coronary Artery Atherosclerosis.

Coronary artery fistulae (CAF) are a rare anomaly characterized by abnormal connections between a coronary artery and a cardiac chamber or a great vessel, with most patients remaining asymptomatic. Despite being predisposed to severe complications like heart failure, patients with CAF infrequently experience severe stenosis in the coronary artery. This study delineates a case involving a 46-year-old male presenting with a fistula bridging the right coronary artery (RCA) and right atrium (RA), manifesting a pronounced 99% stenosis at the right extremity of the coronary artery proximal to the fistula. Concurrently, the individual exhibits six conventional risk factors: age over 40, male gender, hypertension, diabetes, smoking, and hypertriglyceridemia. Following pharmaceutical intervention, the patient was discharged and subjected to extended follow-up. This case highlights the dual processes of "accelerating damage" and "retarding renewal" in the progression of atherosclerosis. Factors such as shear stress, smoking, and hypertension are posited to expedite endothelial cell damage, while aging and diabetes may impede the renewal and repair of these cells. Together with the concept of secondary atherosclerotic plaque healing, this case prompts the introduction of a "Double Endothelial Healings" hypothesis, proposing a potential pathogenetic mechanism for coronary artery atherosclerosis.

Efficacy and safety of seven Chinese patent medicines combined with conventional triple/quadruple therapy for Helicobacter pylori-positive peptic ulcers: a systematic review and network meta-analysis.

OBJECTIVES: To compare the efficacy and safety of seven Chinese patent medicines (CPMs) combined with conventional triple/quadruple therapy (T/Q) for Helicobacter pylori-positive peptic ulcers. DESIGN: A systematic review and network meta-analysis. DATA SOURCES: China National Knowledge Infrastructure, VIP database, Wanfang database, ScienceDirect, EBSCO, EMBASE, Web of Science, Cochrane Library and PubMed were searched through 1 June 2022. ELIGIBILITY CRITERIA: Randomised controlled trials (RCTs) testing CPMs combined with T/Q for H. pylori-positive peptic ulcers were included. The CPMs included Anweiyang capsule, Jianweiyuyang tablets/capsule/granule, Jinghuaweikang capsule, Kangfuxin liquid, Puyuanhewei capsule, Weifuchun tablets/capsule and Weisu granule. At least one of the following outcome indicators was recorded: complete ulcer healing rate (CUHR), effective rate (ER), H. pylori eradication rate (HPER), rate of peptic ulcer recurrence (RPUR) and incidence of adverse reactions (IAR). DATA EXTRACTION AND SYNTHESIS: Two researchers independently conducted the study selection and extracted data for included studies. The risk of bias was assessed using the Cochrane risk of bias tool. A pairwise meta-analysis was performed using RevMan V.5.3. Network meta-analysis was performed using STATA/MP V.15.0. Confidence in the evidence was assessed using Grading of Recommendations, Assessment, Development and Evaluation. RESULTS: A total of 36 RCTs involving 3620 patients were included. Compared with T/Q alone, Weisu+T/Q, Weifuchun+T/Q and Puyuanhewei+T/Q had the highest CUHR, ER and HPER, respectively. Weisu+T/Q and Jianweiyuyang+T/Q had the lowest RPUR and IAR, respectively. The cluster analysis results showed Jianweiyuyang+T/Q might be the best choice concerning efficacy and safety simultaneously, followed by Kangfuxin+T/Q. CONCLUSION: Among the combination therapies with the CPMs, Jianweiyuyang+T/Q might be the most favourable option for H. pylori-positive peptic ulcers, followed by Kangfuxin+T/Q. Considering the limited quantity and quality of the included RCTs, the results should be interpreted with caution. PROSPERO REGISTRATION NUMBER: CRD42022327687.

Iron regulatory protein from the hard tick Haemaphysalis longicornis: characterization, function and assessment as a protective antigen.

BACKGROUND: Ticks are blood-feeding ectoparasites with different host specificities and are capable of pathogen transmission. Iron regulatory proteins (IRPs) play crucial roles in iron homeostasis in vertebrates. However, their functions in ticks remain poorly understood. The aim of the present study was to investigate the characteristics, functions, molecular mechanisms, and the vaccine efficacy of IRP in the hard tick Haemaphysalis longicornis. RESULTS: The full-length complementary DNA of IRP from Haemaphysalis longicornis (HlIRP) was 2973 bp, including a 2772 bp open reading frame. It is expressed throughout three developmental stages (larvae, nymphs, and adult females) and in various tissues (salivary glands, ovaries, midgut, and Malpighian tubules). Recombinant Haemaphysalis longicornis IRP (rHlIRP) was obtained via a prokaryotic expression system and exhibited aconitase, iron chelation, radical-scavenging, and hemolytic activities in vitro. RNA interference-mediated IRP knockdown reduced tick engorgement weight, ovary weight, egg mass weight, egg hatching rate, and ovary vitellin content, as well as prolonging the egg incubation period. Proteomics revealed that IRP may affect tick reproduction and development through proteasome pathway-associated, ribosomal, reproduction-related, and iron metabolism-related proteins. A trial on rabbits against adult Haemaphysalis longicornis infestation demonstrated that rHlIRP vaccine could significantly decrease engorged weight (by 10%), egg mass weight (by 16%) and eggs hatching rate (by 22%) of ticks. The overall immunization efficacy using rHlIRP against adult females was 41%. CONCLUSION: IRP could limit reproduction and development in Haemaphysalis longicornis, and HlIRP was confirmed as a candidate vaccine antigen to impair tick iron metabolism and protect the host against tick infestation. © 2024 Society of Chemical Industry.

Morphology, genetic characterization and phylogeny of Moniliformis tupaia n. sp. (Acanthocephala: Moniliformidae) from the northern tree shrew Tupaia belangeri chinensis Anderson (Mammalia: Scandentia).

A new species of Moniliformis, M. tupaia n. sp. is described using integrated morphological methods (light and scanning electron microscopy) and molecular techniques (sequencing and analysing the nuclear 18S, ITS, 28S regions and mitochondrial cox1 and cox2 genes), based on specimens collected from the intestine of the northern tree shrew Tupaia belangeri chinensis Anderson (Scandentia: Tupaiidae) in China. Phylogenetic analyses show that M. tupaia n. sp. is a sister to M. moniliformis in the genus Moniliformis, and also challenge the systematic status of Nephridiacanthus major. Moniliformis tupaia n. sp. represents the third Moniliformis species reported from China.

The diverse functions of Mu-class Glutathione S-transferase HrGSTm1 during the development of Hyalomma rufipes with a focus on the detoxification metabolism of cyhalothrin.

BACKGROUND: Glutathione S-transferases (GSTs) are a superfamily of multifunctional enzymes in living organisms with metabolic and detoxification functions, which can detoxify exogenous and endogenous compounds and thereby reduce the damage caused by toxic substances to the body. Ticks are obligate blood-sucking ectoparasites that can transmit various pathogens, and the characterization of tick-derived GSTs may help improve current understanding of the molecular mechanism of tick resistance to insecticides. In this study, a novel GST gene, named HrGSTm1, was identified from Hyalomma rufipes. METHODS: Sequence analysis was performed by using bioinformatics techniques. A prokaryotic expression system was used to obtain the recombinant expression protein rHrGSTm1. Detection of spatiotemporal expression patterns of target genes and their response to the toxicity of cyhalothrin on female H. rufipes was performed by using a quantitative PCR platform. The optimal enzymological parameters of rHrGSTm1 using glutathione as substrate were calculated. The antioxidant capacity of the recombinant protein was evaluated by DPPH• (1,1-Diphenyl-2-picrylhydrazyl radical 2,2-Diphenyl-1-(2,4,6-trinitrophenyl) hydrazyl). Knockdown of the HrGSTm1 genes through RNA interference was used to analyze their effects on the physiological parameters of ticks. The changes in HrGSTm1 messenger RNA expression patterns under cypermethrin stress were analyzed. RESULTS: The complementary DNA sequence of HrGSTm1 contained a 672-bp open reading frame, which potentially encoded 223 amino acids. The predicted molecular weight was 25.62 kDa, and the isoelectric point 8.22. HrGSTm1 is a Mu-class GST, belonging to the cytoplasmic GSTs with no signal peptide observed. The Vmax and Km of rHrGSTm1 were 3.367 ± 0.81 uM and 2.208 ± 0.76 uM, respectively, and its activities were dependent on different temperatures and pH conditions; the scavenging rate of rHrGSTm1 to DPPH• reached 76.4% at 1.25 mg/ml. Variable expressions of HrGSTm1 were observed under various treatment periods and in different tissues, with the highest appearing in eggs (analysis of variance [ANOVA], F(2, 9) = 279.9, P < 0.0001) and Malpighian tubules (ANOVA, F(3, 12) = 290.5, P < 0.0001). After knockdown of HrGSTm1, compared with the control group, the mortality in the treatment group was increased by 16.7%, the average oviposition rate decreased by 33.9%, the average engorged body weight decreased by 287.38 mg and egg weight decreased by 127.46 mg, although only the engorged body weight was significantly different (t-test, t(44) = 2.886, P = 0.006). After exposure to three sublethal concentrations (LC05, LC10, LC50) of cyhalothrin, the expression level of HrGSTm1 in the midgut, ovary and salivary gland was upregulated, whereas in Malpighian tubules, it showed a trend of upregulation at first and then downregulation, implying different functions during the detoxification in different tissues. CONCLUSIONS: In this study, a novel GST of the Mu-class was successfully isolated from H. rufipes and systematically subjected to bioinformatic analysis and recombination identification. The variation trend of HrGSTm1 expression level in different tissues suggests that the gene has different detoxification functions in different tissues. The potential function of this gene was analyzed to provide basic research for further investigation of its detoxification mechanism.

Impacts of net cages on pollutant accumulation and its consequence on antibiotic resistance genes (ARGs) dissemination in freshwater ecosystems: Insights for sustainable urban water management.

There has been increasing interest in the role of human activities in disseminating antibiotic-resistance genes (ARGs) in aquatic ecosystems. However, the influence of pollutant accumulation on anthropogenic pollutant-ARG synergistic actions is limited. This study explored the association of net cages with the propagation of anthropogenic pollutants and their consequences for influencing the enrichment of ARGs using high-throughput metagenomic sequencing. We showed that net cages could substantially impact the ecology of freshwater systems by enhancing i) ARG diversity and the tendency for ARG-horizontal gene transfer and ii) the overlap of mobile genetic elements (MGEs) with biocide-metal resistance genes (BMRGs) and ARGs. These findings suggested that the cotransfer of these three genetic determinants would be favored in net cage plots and that nonantibiotic factors such as metal(loid)s, particularly iron (Fe), displayed robust selective pressures on ARGs exerted by the net cage. The resistome risk scores of net cage sediments and biofilms were higher than those from off-net cage plots, indicating that the net cage-origin antibiotic resistome should be of great concern. The combination of deterministic and stochastic processes acting on bacterial communities could explain the higher ARG variations in cage plots (8.2%) than in off-cage plots (3.4%). Moreover, MGEs and pollutants together explained 43.3% of the total variation in ARG communities, which was higher than that of off-cage plots (8.8%), considering pollutants, environmental variables, MGEs, and assembly processes. These findings will inform the development of policies and guidelines to more effectively limit the spread of antimicrobial resistance and achieve the goal of sustainability in freshwater systems in urban areas.

Sequential expression of small heat shock proteins contributing to the cold response of Haemaphysalis longicornis (Acari: Ixodidae).

BACKGROUND: Haemaphysalis longicornis is an important livestock pest and a serious threat to public health. Cold is a common form of stress affecting its survival and distribution. However, H. longicornis exhibits different physiological responses to cold stress. In this study, we systematically explored the regulation and functions of small heat shock proteins (sHsps) in H. longicornis during cold stress. RESULTS: Seven sHsp genes (HlsHsp14.9, HlsHsp19.9, HlsHsp20.3, HlsHsp21.4, HlsHsp23.7, HlsHsp24.0, and HlsHsp26.1) with open reading frame lengths ranging from 408 bp (HlsHsp14.9) to 673 bp (HlsHsp26.1) were cloned from H. longicornis, and featured the typical α-crystallin domain. Phylogenetic analysis revealed high similarity with the sHsps of arachnid species. Quantitative polymerase chain reaction analysis revealed that the regulation of sHsp genes depended on the severity and duration of cold treatment. Moreover, the relative expression of each gene was largely dependent on the treatment period (P < 0.01; 3, 6, and 9 days of treatment at 8, 4, 0, and -4 °C). Among all genes, HlsHsp14.9, HlsHsp19.9, HlsHsp20.3, and HlsHsp24.0 were most sensitive to rapid cold treatment. After RNA interference, the mortality of H. longicornis was significantly increased at -14 °C (P < 0.05), suggesting that the expression of sHsp genes is closely related to cold tolerance in H. longicornis. CONCLUSION: Our results indicate that sHsps play an important role in the cold stress response of H. longicornis, which may enhance our understanding of the cold adaptation mechanisms in ticks. © 2023 Society of Chemical Industry.

Molecular characterization and modulated expression of histone acetyltransferases during cold response of the tick Dermacentor silvarum (Acari: Ixodidae).

BACKGROUND: Histone acetylation is involved in the regulation of stress responses in multiple organisms. Dermacentor silvarum is an important vector tick species widely distributed in China, and low temperature is a crucial factor restricting the development of its population. However, knowledge of the histone acetyltransferases and epigenetic mechanisms underlying cold-stress responses in this tick species is limited. METHODS: Histone acetyltransferase genes were characterized in D. silvarum, and their relative expressions were determined using qPCR during cold stress. The association and modulation of histone acetyltransferase genes were further explored using RNA interference, and both the H3K9 acetylation level and relative expression of KAT5 protein were evaluated using western blotting. RESULTS: Three histone acetyltransferase genes were identified and named as DsCREBBP, DsKAT6B, and DsKAT5. Bioinformatics analysis showed that they were unstable hydrophilic proteins, characterized by the conserved structures of CBP (ZnF_TAZ), PHA03247 super family, Creb_binding, and MYST(PLN00104) super family. Fluorescence quantitative PCR showed that the expression of DsCREBBP, DsKAT6B, and DsKAT5 increased after 3 days of cold treatment, with subsequent gradual decreases, and was lowest on day 9. Western blotting showed that both the H3K9 acetylation level and relative expression of KAT5 in D. silvarum increased after treatment at - 4, 4, and 8 °C for 3 and 6 days, whereas they decreased significantly after a 9-day treatment. RNA interference induced significant gene silencing, and the mortality rate of D. silvarum significantly increased at the respective semi-lethal temperatures. CONCLUSION: These results imply that histone acetyltransferases play an important role in tick adaptation to low temperatures and lay a foundation for further understanding of the epigenetic regulation of histone acetylation in cold-stressed ticks. Further research is needed to elucidate the mechanisms underlying histone acetylation during cold stress in ticks.

The genome-wide characterization and associated cold-tolerance function of the superoxide dismutase in the cold response of the tick Haemaphysalis longicornis.

Accumulating evidence suggests that superoxide dismutase (SOD) is the first line of antioxidant defense in organisms and plays an important role in scavenging reactive oxygen species produced during environmental stress. However, limited information is available regarding the response of SOD genes to cold stress in ticks. Therefore, in the present study, SOD genes were cloned and identified from the genome of Haemaphysalis longicornis, and the function of SOD during the cold response was further explored. Seven SOD genes were characterized: HlCCS1, HlCCS2, HlMSD, HlCSD1, HlCSD2, HlCSD3, and HlCSD4. Bioinformatics analysis showed that HlCCS1 and HlCCS2 are copper chaperones of SODs. HlCSD1-HlCSD4 belong to the Cu/Zn SOD, whereas HlMSD belongs to the Mn SOD gene family. Fluorescence quantitative PCR showed that the expression of HlCCS2, HlMSD, and HlCSD1-3 was upregulated, whereas HlCCS1 and HlCSD4 were downregulated during the cold response of H. longicornis. Western blotting confirmed changes in the relative expression of HlCSD3 and HlMSD in H. longicornis after cold treatment. Mortality of H. longicornis increased significantly after dsRNA injection of HlCCS2, HlMSD, HlCSD1, and HlCSD3. The above results show that SODs have different regulatory functions during the cold response in H. longicornis, and there might be an interaction between treatment temperature and duration. Furthermore, the results lay a foundation for subsequent research on the molecular mechanism of cold tolerance in H. longicornis and shed light on the population distribution and diffusion limit of ticks.

Phylogenetic analysis and molecular characterization of the co-infection of the new variant of the porcine epidemic diarrhea virus and the novel porcine kobuvirus isolated from piglets with diarrhea.

Porcine epidemic diarrhea virus (PEDV) is a virus that can cause diarrhea in pigs, resulting in significant economic losses to the pig industry. The mutation of the virus and its co-infection with other enteroviruses leads to poor control of PEDV infection. In this study, we found that the diarrhea outbreak in a pig farm in Shandong Province was mainly caused by PEDV infection. Through high-throughput sequencing, we also detected one other diarrhea-related virus (porcine kobuvirus). In the phylogenetic analysis and molecular characterization of the detected PEDV S gene and PKV, it was found that the S gene of the PEDV strain detected in this study (named SD22-2) had more mutations than the CV777 strain. The highest homology between PKV (named SD/2022/China) detected in this study and other strains was only 89.66%. Based on polyprotein, we divided SD/2022/China strains into a new grouping (designated group 4) and detected recombination signals. In summary, SD22-2 detected in this study is a new PEDV variant strain, and SD/2022/China strain might be a novel PKV strain. We also found the co-infection of the new PEDV variant and the novel PKV isolated from piglets with diarrhea. Our data suggested the importance of continuous surveillance of PEDV and PKV.

A novel recombinant porcine sapovirus infection in piglets with diarrhea in Shandong Province, China, 2022.

Sapporo virus (SaV) is an emerging enteric virus causing acute gastroenteritis in animals. Here, we found a novel porcine SaV (PoSaV) strain (named SD2202) from the piglets with diarrhea in China in 2022. The highest nucleotide homology of SD2202 with other PoSaV strains is only 90.67%, and there are four amino acids insertion in the viral capsid protein and minor structural protein compared to other PoSaV; furthermore, we found that SD2202 belongs to a new GIII genogroup clade (GIII-6 clade). Interestingly, we found that SD2202 may be an intra-genogroup recombinant strain. Taken together, we found a novel PoSaV implicated in the piglet diarrhea epidemic and emphasized the importance of continuous surveillance of PoSaV.

CircRNA DICAR as a novel endogenous regulator for diabetic cardiomyopathy and diabetic pyroptosis of cardiomyocytes.

In this study, we identified that a conserved circular RNA (circRNA) DICAR, which was downregulated in diabetic mouse hearts. DICAR had an inhibitory effect on diabetic cardiomyopathy (DCM), as the spontaneous cardiac dysfunction, cardiac cell hypertrophy, and cardiac fibrosis occurred in DICAR deficiency (DICAR+/-) mice, whereas the DCM was alleviated in DICAR-overexpressed DICARTg mice. At the cellular level, we found that overexpression of DICAR inhibited, but knockdown of DICAR enhanced the diabetic cardiomyocyte pyroptosis. At the molecular level, we identified that DICAR-VCP-Med12 degradation could be the underlying molecular mechanism in DICAR-mediated effects. The synthesized DICAR junction part (DICAR-JP) exhibited a similar effect to the entire DICAR. In addition, the expression of DICAR in circulating blood cells and plasma from diabetic patients was lower than that from health controls, which was consistent with the decreased DICAR expression in diabetic hearts. DICAR and the synthesized DICAR-JP may be drug candidates for DCM.