Coriaceumins A-D, the First Nitrogen-Containing Crenulide Diterpenoids from the Brown Alga Dictyota coriacea Collected in the East China Sea.

A phytochemical investigation of an East China Sea collection of the brown alga Dictyota coriacea has led to the isolation of four novel nitrogen-containing crenulide diterpenoids, named coriaceumins A-D (1-4), two rare nitrogenous xenicane diterpenoids, dictyolactams C (5) and D (6), and one known crenulide diterpenoid, hydroxycrenulide (7). The structures of the new compounds were elucidated by detailed spectroscopic data analyses, including HRESIMS and 1D/2D NMR. The absolute configurations were determined by a comparison of the experimental ECD spectra with the spectra computed by DFT-based quantum chemical calculations. Coriaceumins A-D (1-4) represent the first examples of nitrogen-containing crenulide diterpenoids. In a bioassay, compounds 2, 3, 5, and 7 were found to exhibit different levels of inhibitory effects against protein tyrosine phosphatase 1B (PTP1B) with IC50 values ranging from 7.3 to 19 µM. In addition, the primary structure-activity relationships of all the isolates were summarized.

Lyonensinols A - C, 24-Norursane-Type Triterpenoids from the Twigs and Leaves of Lyonia doyonensis and Their Potential Anti-inflammatory and PTP1B Inhibitory Activities.

Lyonia doyonensis is a deciduous shrub native to high-altitude regions of Asia. So far, there is no report on any chemical and biological properties of L. doyonensis. An EtOH extract of L. doyonensis twigs and leaves showed inhibitory activities on protein tyrosine phosphatase 1B and lipopolysaccharide-induced inflammation in BV-2 microglial cells. A phytochemical investigation of this extract led to the isolation of a, so far only ambiguously described, 24-norursane-type triterpenoid, now named lyonensinol A (1: ), along with its two new derivatives, lyonensinols B and C (2: and 3: ), and six known triterpenoids (4 - 9: ). Their structures were elucidated by detailed analysis of spectroscopic data. A combination of chemical conversions, electronic circular dichroism, and Mo2(OAc)4-induced electronic circular dichroism was used to confirm their absolute configurations. Lyonensinols B (2: ) and C (3: ) represent the first examples of norursane-type triterpenoids acylated with a p-coumaroyl moiety. The potential anti-inflammatory and protein tyrosine phosphatase 1B inhibitory activities of all the isolates were evaluated. Compounds 3, 7: , and 8: at 10 µM showed potent inhibitory activities on lipopolysaccharide-induced nitric oxide production in BV-2 microglial cells, with nitric oxide levels decreasing to 31.5, 41.9, and 27.1%, respectively, while compounds 3, 4, 7: , and 8: exhibited notable inhibitory activities against protein tyrosine phosphatase 1B, with IC50 values ranging from 1.7 to 18.2 µM. Interestingly, compounds 3: and 8: , bearing a C-3 trans-p-coumaroyl group, showed not only more potent anti-inflammatory effects, but also exhibited stronger protein tyrosine phosphatase 1B inhibition than their respective stereoisomers (2: and 7: ) with a cis-p-coumaroyl group.

Nasal nitric oxide in healthy Chinese children aged 6-18 years.

Objectives: To obtain the normal values of fractional concentration of nasal nitric oxide in Chinese children aged 6-18 years, so as to provide reference for clinical diagnosis. Methods: 2,580 out of 3,200 children (1,359 males and 1,221 females), whom were included from 12 centers around China were taken tests, their height and weight were also recorded. Data were used to analyze the normal range and influencing factors of fractional concentration of nasal nitric oxide values. Measurements: Data was measured using the Nano Coulomb Breath Analyzer (Sunvou-CA2122, Wuxi, China), according to the American Thoracic Society/European Respiratory Society (ATS/ERS) recommendations. Main Results: We calculated the normal range and prediction equation of fractional concentration of nasal nitric oxide values in Chinese children aged 6-18 years. The mean FnNO values of Chinese aged 6-18 yrs was 454.5 ± 176.2 ppb, and 95% of them were in the range of 134.5-844.0 ppb. The prediction rule of FnNO values for Chinese children aged 6-11 yrs was: FnNO = 298.881 + 17.974 × age. And for children aged 12-18 yrs was: FnNO = 579.222-30.332 × (male = 0, female = 1)-5.503 × age. Conclusions: Sex and age were two significant predictors of FnNO values for Chinese children(aged 12-18 yrs). Hopefully this study can provide some reference value for clinical diagnosis in children.

Factors associated with reproductive concerns among young female patients with colorectal cancer: A cross-sectional study.

AIMS AND OBJECTIVES: The aim of this study was to describe the prevalence of reproductive concerns among young female patients with colorectal cancer and explore the associated factors. BACKGROUND: With the trend of longer survival and younger age at diagnosis of colorectal cancer patients, reproductive concerns have become increasingly prevalent among young female colorectal cancer patients. DESIGN: Cross-sectional design. METHODS: The study included 150 young female patients with colorectal cancer who completed cancer treatment at 2 hospitals in Guangzhou, China, between November 2020 and December 2021 completed an investigation comprising A general questionnaire, The Reproductive Concerns After Cancer scale, The Family Adaptation and Cohesion Evaluation Scale II and unmet fertility information needs questionnaire. Multivariable linear regression analysis was performed in order to identify factors that influence reproductive concerns. This study was prepared and is reported according to the STROBE checklist. RESULTS: The mean (SD) score on the Reproductive Concerns After Cancer scale was 54.78 ± 8.97. The highest score was for the children's health subscale (3.84 ± .92) and the lowest was for acceptance (2.24 ± .70). Multiple regression analysis showed that patients with fewer children, female children, lower education level (less than undergraduate degree), earlier disease stage, lower family function and higher unmet need for fertility information had more reproductive concerns, which explained 26.9% of the total variation of the model. CONCLUSIONS: The patients with fewer children, female children, low cultural degree (less than bachelor), early clinical patients, poorer family function and higher unmet fertility information needs had higher reproductive concerns. RELEVANCE TO CLINICAL PRACTICE: These findings can guide the development of interventions to mitigate reproductive concerns, including understand and meet their fertility information needs, improve the level of family function. PATIENT OR PUBLIC CONTRIBUTION: Survey questionnaires were completed by participants among young female with colorectal cancer in this study.

Coumaronochromones, flavanones, and isoflavones from the twigs and leaves of Erythrina subumbrans inhibit PTP1B and nitric oxide production.

A chemical investigation of the twigs and leaves of Erythrina subumbrans led to the isolation and structural elucidation of three coumaronochromones, erythrinasubumbrin A and (±)-erythrinasubumbrin B, five prenylated flavanones, (±)-erythrinasubumbrin C and erythrinasubumbrins D-F, and two prenylated isoflavones, (±)-5,4'-dihydroxy-[4,5-cis-4-ethoxy-5-hydroxy-6,6-dimethyl-4,5-dihydropyrano (2,3:7,6)]-isoflavone, in addition to 18 known analogues. Two extra cinnamylphenols previously only known as commercial synthetic products were also isolated and elucidated from a natural source for the first time, and assigned the trivial names erythrinasubumbrins G and H. Their structures were characterized by detailed analysis of spectroscopic data, including HRESIMS and 2D NMR. The absolute configurations of the previously undescribed isolates and the known coumaronochromone lupinol C were determined by specific rotation and electronic circular dichroism (ECD) data. All the isolates were evaluated for their inhibitory activities on protein tyrosine phosphatase 1 B (PTP1B) and nitric oxide (NO) production in lipopolysaccharide (LPS)-induced BV-2 microglial cells as well as their cytotoxicity against the HCT116 cell line. The pair of enantiomers, (+)-5,4'-dihydroxy-[4,5-cis-4-ethoxy-5-hydroxy-6,6-dimethyl-4,5-dihydropyrano (2,3:7,6)]-isoflavone and (-)-5,4'-dihydroxy-[4,5-cis-4-ethoxy-5-hydroxy-6,6-dimethyl-4,5-dihydropyrano (2,3:7,6)]-isoflavone, and the known compounds lupinol C, 4'-O-methyl-8-prenylnaringenin, glepidotin B, shuterin, parvisoflavones A, luteone, lupiwighteone, 2,3-dehydrokievitone, 6,8-diprenylgenistein, angustone A, and 2'-O-demethylbidwillol B exhibited different levels of PTP1B inhibitory activities with IC50 values ranging from 3.21 to 19.17 µM, while erythrinasubumbrin A, (-)-erythrinasubumbrin B, (+)-5,4'-dihydroxy-[4,5-cis-4-ethoxy-5-hydroxy-6,6-dimethyl-4,5-dihydropyrano (2,3:7,6)]-isoflavone, (-)-5,4'-dihydroxy-[4,5-cis-4-ethoxy-5-hydroxy-6,6-dimethyl-4,5-dihydropyrano (2,3:7,6)]-isoflavone, and the known compounds lupinol C, 8-prenylnaringenin, macatrichocarpin A, alpinumisoflavone, and 2'-O-demethylbidwillol B substantially inhibited NO production in BV-2 microglial cells. In addition, 8-prenylnaringenin showed weak cytotoxicity with an IC50 value of 9.13 µM. This is the first report of PTP1B inhibitory activity for a coumaronochromone.

Theileria annulata SVSP455 interacts with host HSP60.

BACKGROUND: Theileria annulata, a transforming parasite, invades bovine B cells, dendritic cells and macrophages, promoting the uncontrolled proliferation of these cells. This protozoan evolved intricate strategies to subvert host cell signaling pathways related to antiapoptotic signaling to enable survival and proliferation within the host cells. However, the molecular mechanisms of the cell transformation induced by T. annulata remain largely unclear. Although some studies have predicted that the subtelomere-encoded variable secreted protein (SVSP) family plays roles in host-parasite interactions, the evidence for this is limited. METHODS: In the present study, the SVSP455 (TA05545) gene, a member of the SVSP gene family, was used as the target molecule. The expression pattern of SVSP455 in different life-cycle stages of T. annulata infection was explored using a quantitative real-time PCR assay, and the subcellular distribution of SVSP455 was observed using confocal microscopy. The host cell proteins interacting with SVSP455 were screened using the Y2H system, and their interactions were verified in vivo and in vitro using both bimolecular fluorescence complementation and confocal microscopy, and co-immunoprecipitation assays. The role played by SVSP455 in cell transformation was further explored by using overexpression, RNA interference and drug treatment experiments. RESULTS: The highest level of the SVSP455 transcript was detected in the schizont stage of T. annulata, and the protein was located both on the surface of schizonts and in the host cell cytoplasm. In addition, the interaction between SVSP455 and heat shock protein 60 was shown in vitro, and their link may regulate host cell apoptosis in T. annulata-infected cells. CONCLUSION: Our findings are the first to reveal that T. annulata-secreted SVSP455 molecule directly interacts with both exogenous and endogenous bovine HSP60 protein, and that the interaction of SVSP455-HSP60 may manipulate the host cell apoptosis signaling pathway. These results provide insights into cancer-like phenotypes underlying Theilera transformation and therapeutics for protection against other pathogens.

Anti-inflammatory and PTP1B inhibitory sesquiterpenoids from the twigs and leaves of Aglaia lawii.

Twelve sesquiterpenoids with seven different carbon skeletons, including four isodaucanes (1-4), an aromadendrane (5), a guaiane (6), a cadalane (7), two eudesmanes (8 and 9), two bisabolanes (10 and 11), and a megastigmane (12), were isolated from the twigs and leaves of Aglaia lawii (Wight) C. J. Saldanha et Ramamorthy. Of these compounds, amouanglienoids A (1) and B (2) are new isodaucane sesquiterpenoids. This is the first report of isodaucanes from the genus Aglaia, and amouanglienoid A (1) represents the first isodaucane containing a Δ7(8) double bond. Their structures were discerned from extensive spectroscopic analyses, single-crystal X-ray diffraction, and comparison of the experimental and calculated ECD data. In in vitro bioassays, compounds 1, 10, and 11 showed potent inhibitory effects against lipopolysaccharide (LPS)-induced inflammation in BV-2 microglial cells, while compound 11 exhibited considerable inhibition of PTP1B with an IC50 value of 16.05 ± 1.09 µM.

Reference values of impulse oscillometry (IOS) for healthy Chinese children aged 4-17 years.

OBJECTIVE: To establish the predicted value of pulmonary function determined by impulse oscillometry (IOS) in children (4-17 years old) in China. METHODS: A total of 6270 healthy children aged 4-17 years in China were included. The Master Screen IOS pulmonary function device (Jaeger Co, Germany) was used to detect the respiratory impedance (Zrs), resonant frequency (Fres), respiratory system resistance (Rrs) and respiratory system reactance (Xrs) at various oscillation frequencies, and the indices above were analysed. Stepwise multivariate regression was used to establish the regression equation of related parameters of IOS in different sexes, ages, height, and weight. RESULTS: The differences in the main IOS parameters between different age stages were statistically significant regardless of sex (P < 0.05). The stepwise multivariate regression analysis showed that IOS parameters were related to height, age, and weight, and most IOS parameters were most closely related to height (the absolute value of the regression coefficient was the largest). With increasing age and height, the values of Z5, R5, R20, R5-R20, and Fres decreased, while the value of X5 increased. Through height, age, and weight, we obtained the normal predicted values equation of children's IOS parameters. Compared with the other reference equations, our reference equation is more suitable for Chinese children. CONCLUSIONS: The study revealed the reference values of IOS parameters in healthy Chinese children. In the evaluation of results for lung function measurements, this predicted value equation is more consistent with the characteristics of Chinese children than other reference equations. CLINICAL TRIAL: ChiCTR: 1800019029.

Identification and isolation of pathogenic Theileria orientalis Ikeda genotype from confined dairy cattle, in Hebei, China.

Theileria orientalis is known to be a group of benign cattle parasites with a cosmopolitan distribution, and has been classified into 11 genotypes through MPSP gene phylogenetic analysis. In China, T. orientalis is the most prevalent Theileria species, with several genotypes, but few fatal cases have been reported. In June 2020, dairy cattle in Zhangjiakou, Hebei Province, showed clinical symptoms of piroplasmosis, causing many animals to die. Blood smears and PCR detection results confirmed T. orientalis infection with a 66.7% positive rate of collected blood samples. The MPSP sequences analysis revealed parasite genotypes 1 (Chitose) and 2 (Ikeda). Aiming to isolate the pathogens, experimental animal was infected with T. orientalis via inoculation of the positive blood samples. The results has shown that only T. orientalis genotype 2 (Ikeda) was obtained that has confirmed by MPSP and 18S rRNA sequences analysis, indicating that the Ikeda type was predominant and responsible for the disease. Although many T. orientalis genotypes are present in China, the possibility of T. orientalis genotypes 1 and 2 infections in confined dairy cattle should be considered to avoid additional economic losses.

Analysis of Factors Associated With Depression in Community-Dwelling Older Adults in Wuhan, China.

Objectives: This study aimed to identify the independent factors associated with depression in community-dwelling older adults in Wuhan, China. Methods: Four hundred and seventy older adults (aged ≥65 years) from four communities dwelling on Junshan Street in Wuhan, China were included in this study. Participants completed a questionnaire that asked questions pertaining to age, gender, educational level, income, living situation, care situation, social support, and social engagement. The 30-item Geriatric Depression Scale (GDS-30), the Fried frailty phenotype scale, the activities of daily living (ADL) scale, the mini nutritional assessment scale-short form (MNA-SF), and the Mini-cog scale were used to assess depression, frailty, self-care ability, malnutritional risk, and cognitive dysfunction, respectively. Differences in age, gender, educational level, income, living situation, care situation, social support, social engagement, ADL score, risk of malnutrition, frailty, and cognitive dysfunction between the non-depression (GDS-30 score <10 points) and depression groups (GDS-30 score ≥10 points) were compared using a chi-square test. Moreover, correlations between factors and depression were analyzed using Pearson's correlation. Then, significant variables (p < 0.05) from the chi-square test were included in a multivariable logistic regression model to identify the independent factors associated with depression. Results: The incidence of depression among the participants was 14.04%. Age (p < 0.001), educational level (p < 0.001), living situation (p < 0.001), social support (p = 0.001), ADL score (p = 0.023), frailty (p < 0.001), and cognitive dysfunction (p < 0.001) were all significantly associated with depression, in which age, poor social support, frailty, and cognitive dysfunction were identified as independent factors. Conclusion: Improving social support and effective interventions for frailty and cognitive dysfunction may help relieve depression in community-dwelling older adults.

A universal ELISA assay for detecting six strains of ovine Babesia species in China.

Ovine babesiosis, caused by genus of Babesia, is a zoonotic disease and mainly transmitted by hard ticks. It has led to enormous economic losses to the sheep industry in China. In the present study, an ELISA assay for simultaneous detection six strains of Babesia spp., including B. motasi Lintan, B. motasi Tianzhu, B. motasi Hebei, B. motasi Ningxian, Babesia sp. Xinjiang and Babesia sp. Dunhuang, was developed using Apical Membrane Antigen 1 (AMA1) as candidate diagnostic antigen. The sensitivity and specificity of the established ELISA were 97.4 % and 98.0 %, respectively. Relatively high level of specific antibodies could be detected from 12th day to 126th day after sheep experimentally infected with Babesia spp.. A small scale of field sera was investigated using the developed ELISA assay, and the average positive rate was 51.98 %. This study provides an easy to operate, cost effective and time saving approach, which is suitable for both field and experimental samples, thus it could be a useful tool in epidemiological investigations and diagnoses of ovine babesiosis.

Systematic Comparison of the Performances of De Novo Genome Assemblers for Oxford Nanopore Technology Reads From Piroplasm.

Background: Emerging long reads sequencing technology has greatly changed the landscape of whole-genome sequencing, enabling scientists to contribute to decoding the genetic information of non-model species. The sequences generated by PacBio or Oxford Nanopore Technology (ONT) be assembled de novo before further analyses. Some genome de novo assemblers have been developed to assemble long reads generated by ONT. The performance of these assemblers has not been completely investigated. However, genome assembly is still a challenging task. Methods and Results: We systematically evaluated the performance of nine de novo assemblers for ONT on different coverage depth datasets. Several metrics were measured to determine the performance of these tools, including N50 length, sequence coverage, runtime, easy operation, accuracy of genome and genomic completeness in varying depths of coverage. Based on the results of our assessments, the performances of these tools are summarized as follows: 1) Coverage depth has a significant effect on genome quality; 2) The level of contiguity of the assembled genome varies dramatically among different de novo tools; 3) The correctness of an assembled genome is closely related to the completeness of the genome. More than 30× nanopore data can be assembled into a relatively complete genome, the quality of which is highly dependent on the polishing using next generation sequencing data. Conclusion: Considering the results of our investigation, the advantage and disadvantage of each tool are summarized and guidelines of selecting assembly tools are provided under specific conditions.

Stable transfection system for Babesia sp. Xinjiang.

BACKGROUND: Stable transfection systems have been described in many protozoan parasites, including Plasmodium falciparum, Cryptosporidium parvum, Babesia bovis, Babesia ovata, and Babesia gibsoni. For Babesia sp. Xinjiang (Bxj), which is the causative pathogen of ovine babesiosis and mainly prevails across China, the platform of those techniques remains absent. Genetic manipulation techniques are powerful tools to enhance our knowledge on parasite biology, which may provide potential drug targets and diagnostic markers. METHODS: We evaluated the inhibition efficiency of blasticidin (BSD) and WR99210 to Bxj. Then, a plasmid was constructed bearing selectable marker BSD, green fluorescent protein (GFP) gene, and rhoptry-associated protein-1 3' terminator region (rap 3' TR). The plasmid was integrated into the elongation factor-1 alpha (ef-1α) site of Bxj genome by cross-over homologous recombination technique. Twenty µg of plasmid was transfected into Bxj merozoites. Subsequently, drug selection was performed 24 h after transfection to generate transfected parasites. RESULTS: Transfected parasite lines, Bxj-c1, Bxj-c2, and Bxj-c3, were successfully obtained after transfection, drug selection, and colonization. Exogenous genes were integrated into the Bxj genome, which were confirmed by PCR amplification and sequencing. In addition, results of western blot (WB) and indirect immunofluorescence assay (IFA) revealed that GFP-BSD had expressed for 11 months. CONCLUSIONS: In our present study, stable transfection system for Bxj was successfully developed. We anticipate that this platform will greatly facilitate basic research of Bxj.

Assessment of probiotic adhesion and inhibitory effect on Escherichia coli and Salmonella adhesion.

In this study, we screened bacterial strains to identify specific probiotics to treat pig diarrhea caused by Escherichia coli or Salmonella. The potential probiotics were assayed for their survival in gastrointestinal solution, their antimicrobial activity, cell-surface properties, adhesion to Caco-2 cells, and inhibition of pathogen adhesion. Nine out of the 20 strains tested showed high tolerance of a simulated gastrointestinal environment and six strains exerted antagonistic effects against enterohemorrhagic E. coli (EHEC) O157:H7 and Salmonella Typhimurium MQ. Lactobacillus johnsonii pDX1e exhibited a higher potent antibacterial activity. Four strains (pDX1a, pDX1e, pDX3a, and pDX5a) displayed auto-aggregation, hydrophobicity, and adhesion to Caco-2 cells similar to those of the reference strain Lacticaseibacillus rhamnosus GG (LGG). Enterococcus durans pDX5a showed the highest adhesion capacity (13.86%), followed by the reference strain LGG (11.20%). All the tested strains competitively suppressed the attachment of pathogens to Caco-2 cells (by 30.73-55.18%); L. johnsonii pDX1e and Ent. durans pDX5a significantly inhibited the adhesion of pathogens by substitution and exclusion, respectively. Therefore, pDX1e and pDX5a were selected as probiotic strains for further investigation and application.

Application Value of Information-Based Health Education and Continuity of Care in Patients With Peptic Ulcer.

Objective: This study is to assess the application value of information-based health education and continuity of care in patients with PU (peptic ulcer). Methods: Patients (116) with PU who have been treated in the hospital between January 2019 and October 2020 were taken as research objects and equally assigned to a control group and an observation group in a random manner. In contrast to the routine care applied to the control group, the observation group received information-based health education and continuity of care intervention. The clinical efficacy, the mastery of health knowledge, self-care ability, medication compliance, quality of life, mental state, and nursing satisfaction of the two groups were compared. Results: After the intervention, the total effective rate, health knowledge adequate rate, Exercise of Self-Care Agency (ESCA) scores of all dimensions, the MOS 36-item short-form health survey (SF-36) scores of all dimensions, medication compliance rate, and total nursing satisfaction of the observation group all notably exceeded those of the control group, with a p < 0.05. Patients of the group with continuity of care intervention showed lower Self-Rating Anxiety Scale (SAS) and Self-Rating Depression Scale (SDS) scores, as compared to the group with conventional care (p < 0.05). Conclusion: Information-based health education and continuity of care elevates the medication adherence and nursing satisfaction of patients with PU, enhances disease-related knowledge of patients and their self-care ability, and eventually ameliorates the quality of life and psychological state. It is worthy of clinical application.

Nomogram for Predicting Survival in Advanced Gastric Cancer after Neoadjuvant Chemotherapy and Radical Surgery.

BACKGROUND: Neoadjuvant chemotherapy (NAC) with subsequent radical surgery has become a popular treatment modality for advanced gastric cancer (AGC) worldwide. However, the survival benefit is still controversial, and prognostic factors remain undetermined. AIM: To identify clinical parameters that are associated with the survival of AGC patients after NAC and radical surgery and to establish a nomogram integrating multiple factors to predict survival. METHODS: We reviewed the medical profiles of 215 AGC patients who received NAC and radical resection, and clinical parameters concerning NAC, surgery, pathological findings, and adjuvant chemotherapy were analyzed using a Cox regression model to determine their impact on survival. Based on these factors, a nomogram was developed and validated. RESULTS: The overall 1-year and 3-year survival rates were 85.8% and 55.6%, respectively. Younger age (<60 years old), increased examined lymph nodes (exLNs), successful R0 resection, the achievement of pathological complete response (pCR), and acceptance of adjuvant chemotherapy were positive predictors of survival. The C-index of the established nomogram was 0.785. The area under receiver operating curve (ROC) at 1/3 years of prediction was 0.694/0.736, respectively. The model showed an ideal calibration following internal bootstrap validation. CONCLUSION: A nomogram predicting survival after NAC and surgery was established. Since this nomogram exhibited satisfactory and stable predictive power, it can be inferred that this is a practical tool for predicting AGC patient survival after NAC and radical surgery.

Establishment of a transient transfection system for Babesia sp. Xinjiang using homologous promoters.

Babesia species, the agentic pathogens of human and animal babesiosis, are spread worldwide. Over the last decade, genetic manipulation approaches have been applied with many protozoan parasites, including Plasmodium falciparum, Trypanosoma cruzi, Cryptosporidium parvum, Theileria annulata, Theileria parva, Babesia bovis, Babesia bigemina, Babesia ovata, Babesia gibsoni, and Babesia ovis. For Babesia sp. Xinjiang (BspXJ), which is the causative pathogen of ovine babesiosis mainly in China, the efficiency of these techniques remains unclear. Firstly, a plasmid bearing the elongation factor-1 alpha promoter and the firefly luciferase reporter gene and rap stop region were transfected into BspXJ by electroporation and nucleoporation to determine the most suitable transfection solution. Then, six program settings were evaluated to confirm the best for BspXJ transient transfection, and a series of different amounts of plasmid DNA were transfected to generate relatively high luminescence values. Finally, the activities of four promoters derived from BspXJ were evaluated using the developed transient transfection system. After evaluating of various transfection parameters, the human T cell nucleofector solution, program V-024 and 20 µg of plasmid DNA were selected as the most favorable conditions for BspXJ transient transfection. These findings provide critical information for BspXJ genetic manipulation, an essential tool to identify virulence factors and to further elucidate the basic biology of this parasite.

Rearranged Diels-Alder Adducts and Prenylated Flavonoids as Potential PTP1B Inhibitors from Morus nigra.

Two novel rearranged Diels-Alder adducts, morunigrines A (1) and B (2), and four new prenylated flavonoids, morunigrols A-D (3-6), were isolated from the twigs of Morus nigra, together with four known prenylated phenolic compounds, including two flavonoids (7 and 8) and two 2-arylbenzofurans (9 and 10). Morunigrines A (1) and B (2) are a novel class of Diels-Alder adducts with unprecedented carbon skeletons featuring a rearranged chalcone-stilbene/2-arylbenzofuran core decorated with a unique methylbiphenyl moiety. The structures of the new compounds were assigned by analysis of spectroscopic data. The absolute configuration of compound 6 was determined by the measurement of specific rotation. A plausible biogenetic pathway for 1 and 2 is also proposed. Compounds 1 and 2 exhibited more potent protein tyrosine phosphatase 1B inhibitory activity with IC50 values of 1.8 ± 0.2 and 1.3 ± 0.3 µM, respectively, than that of the positive control oleanolic acid (IC50, 2.5 ± 0.1 µM).

Screening and identification of Theileria annulata subtelomere-encoded variable secreted protein-950454 (SVSP454) interacting proteins from bovine B cells.

BACKGROUND: Theileria annulata is a protozoan parasite that can infect and transform bovine B cells, macrophages, and dendritic cells. The mechanism of the transformation is still not well understood, and some parasite molecules have been identified, which contribute to cell proliferation by regulating host signaling pathways. Subtelomeric variable secreted proteins (SVSPs) of Theileria might affect the host cell phenotype, but its function is still not clear. Therefore, in the present study, we explored the interactions of SVSP454 with host cell proteins to investigate the molecular mechanism of T. annulata interaction with host cells. METHODS: The transcription level of an SVSP protein from T. annulata, SVSP454, was analyzed between different life stages and transformed cell passages using qRT-PCR. Then, SVSP454 was used as a bait to screen its interacting proteins from the bovine B cell cDNA library using a yeast two-hybrid (Y2H) system. The potential interacting proteins of host cells with SVSP454 were further identified by using a coimmunoprecipitation (Co-IP) and bimolecular fluorescence complementation (BiFC) assays. RESULTS: SVSP454 was transcribed in all three life stages of T. annulata but had the highest transcription during the schizont stage. However, the transcription level of SVSP454 continuously decreased as the cultures passaged. Two proteins, Bos Taurus coiled-coil domain 181 (CCDC181) and Bos Taurus mitochondrial ribosomal protein L30 (MRPL30), were screened. The proteins CCDC181 and MRPL30 of the host were further identified to directly interact with SVSP454. CONCLUSION: In the present study, SVSP454 was used as a bait plasmid, and its prey proteins CCDC181 and MRPL30 were screened out by using a Y2H system. Then, we demonstrated that SVSP454 directly interacted with both CCDC181 and MRPL30 by Co-IP and BiFC assays. Therefore, we speculate that SVSP454-CCDC181/SVSP454MRPL30 is an interacting axis that regulates the microtubule network and translation process of the host by some vital signaling molecules. Identification of the interaction of SVSP454 with CCDC181 and MRPL30 will help illustrate the transformation mechanisms induced by T. annulata.