[Discussion on the history of Pinelliae Rhizoma Praeparatum(Fa banxia) Processing].

Before the Song Dynasty, the main processing method of Pinelliae Rhizoma was soup washing. The "new method" in Taiping Huimin Heji JuFang is a processing method that concocted with Ginger,white alum and starter-making.The "Fa Banxia" in the Yuan Dynasty's Yuyuan Yaofang comes from the Taiping Huimin Heji JuFang, and the Fa Wen Banxia, Fa Bai Banxia, and Fa Hong Banxia are the processing methods of patent medicine with a variety of other herbs.Fa Banxia appeared in the Ming Dynasty, and its auxiliary materials were ginger and white alum, and medical formulary began to include formulas containing Fa Banxia. Bencao Gangmu abbreviates the Yuan Dynasty's Fabai Banxia as "Fa Banxia", and is elaborated under the item attached "prescription" item instead of the "treatment". In the literature of Materia Medica, it is recorded that the preparation of auxiliary materials in Fa Banxia increased, including lime, licorice, soap horn, and simple nitro.After Daoguang in the Qing Dynasty, the Fa Banxia in famous medical cases was more used, and at that time, Fa Banxia was Xian Banxia. There are two recipes for Xian Banxia: one is made with seven processes, and the other is soaked in alum licorice water. During the Republican period, Zhang Cigong also pointed out that Fa Banxia was sliced Xian Banxia after rinsing and boiling, while the preparation method of Xian Banxia was Banxia made of ginger and white alum.Ye Juquan pointed out that the so-called "fa" is neither an ancient method nor a new method, questioning the process of repeated immersion in Banxia. After 1949, the questioning of Fa Banxia continued unceasing.Influenced by this, the 1960 edition Beijing Traditional Chinese Medicine Slice Cutting Experience included the method of soaking alum, licorice ,lime water, and was included in the 1963 edition of the Pharmacopoeia of the People's Republic of China.The 1985 version reduced the soaking time and eliminated the soaking process of alum based on the 1963 version, and this method is still used today.

Association of Fear of Falling With Cognition and Physical Function in Community-Dwelling Older Adults.

BACKGROUND: Fear of falling (FOF) might be associated with physical and cognitive function, but there is a lack of understanding of the specific relationship between the three variables. OBJECTIVES: The aim of this study was to accurately investigate the association of FOF with cognitive and physical function in community-dwelling older adults. METHODS: Six hundred sixty-nine older adults (>60 years old) participated in this study. A self-report questionnaire collected information about demographic characteristics, lifestyle, and behavioral habits. FOF was evaluated through the Shortened Version of the Falls Efficacy Scale International. Global cognitive function and the subdomains of cognitive function (including memory, visual-spatial, language, attention, and executive function) were assessed using the Montreal Cognitive Assessment scale, the Auditory Verbal Learning Test, the Clock Drawing Test (CDT), the Verbal Fluency Test, and the Trail Making Test. Subjective memory complaints were assessed using the Subjective Memory Complaints Questionnaire. Physical function was evaluated by measuring muscle strength and balance ability, and muscle strength was indicated by hand grip strength. In contrast, balance was assessed using the Timed Up and Go (TUG) Test. RESULTS: After adjustment for potential confounding factors, the linear or ordinal regression analysis showed that the values of hand grip strength, Montreal Cognitive Assessment, Auditory Verbal Learning Test, and CDT were significantly and negatively correlated with the score of FOF. On the other hand, Subjective Memory Complaints Questionnaire and TUG Test values showed significant positive correlations with FOF scores. Moreover, compared with other cognitive or physical measures, the CDT and TUG Test values showed a greater association with the FOF scores. DISCUSSION: Low subjective or objective cognitive ability and low physical function, especially low visuospatial and balance ability, were positively associated with the risk of FOF in a community-dwelling older population.

Real-world circulating tumor DNA analysis depicts resistance mechanism and clonal evolution in ALK inhibitor-treated lung adenocarcinoma patients.

BACKGROUND: Sequential treatment with different generations of anaplastic lymphoma kinase (ALK) inhibitors have been widely applied to ALK-positive lung cancer; however, resistance mutations inevitably developed. Further characterization of ALK resistance mutations may provide key guidance to subsequent therapies. Here we explored the emergence of secondary ALK mutations during sequential ALK tyrosine kinase inhibitor (TKI) treatment in a real-world study of Chinese lung adenocarcinoma (ADC) patients. METHODS: A clinical-genomic database was queried for lung ADC patients with at least one ALK inhibitor treatment and at least one plasma sample collected following ALK inhibitor treatment. Targeted genome profiling was performed with a 139-gene panel in baseline tumor tissue and serial plasma samples of patients. RESULTS: A total of 116 patients met inclusion criteria. ALK G1202R was more common in patients with echinoderm microtubule-associated protein-like 4 (EML4)-ALK v3 fusion, whereas ALK L1196M was more common in v1. TP53 mutant patients were significantly associated with harboring multiple ALK resistance mutations (P = 0.03) and v3+/TP53 mutant patients had the highest rate of multiple ALK resistance mutations. The sequential use of ALK TKI led to an increased incidence of concurrent ALK mutations along the lines of therapies. Alectinib had a lower rate (9%) harboring ALK resistance mutation as first-line ALK TKI compared with crizotinib (36%). ALK compound mutations identified included ALK D1203N/L1196M, ALK G1202R/L1196M, and ALK G1202R/F1174C, which may be lorlatinib resistant. Using paired pretreatment and post-treatment samples, we identified several ALK-independent resistance-related genetic alterations, including PTPRD and CNKN2A/B loss, MYC, MYCN and KRAS amplification, and EGFR19del. CONCLUSIONS: Sequential postprogression plasma profiling revealed that increased lines of ALK inhibitors can accelerate the accumulation of ALK resistance mutations and may lead to treatment-refractory compound ALK mutations. The selection for optimal first-line TKI is very important to achieve a more efficacious long-term strategy and prevent the emergence of on-target resistance, which may provide guidance for clinical decision making.

Efficacy of intranasal ketamine for acute pain management in adults: a systematic review and meta-analysis.

OBJECTIVE: This study aimed to compare the efficacy of intranasal (IN) ketamine for pain control with placebo and other analgesics in an emergency setting. MATERIALS AND METHODS: Electronic databases of PubMed, Embase, and CENTRAL were searched for randomized controlled trials (RCTs) comparing IN ketamine with placebo or other analgesics up to 1st January 2021. Studies were to be conducted on adults and in an emergency setting. Pain outcomes and adverse events were compared. RESULTS: Seven RCTs were included. Three compared IN ketamine with placebo while others with opioids. Comparing IN ketamine with opioids, the pooled analysis demonstrated no significant difference in pain scores between the two groups at 15 minutes but better pain reduction with opioids at 30 minutes. Comparing IN ketamine with placebo, our analysis demonstrated a non-significant difference but a tendency for better pain relief with IN ketamine at 15 minutes and 60 minutes. Pain scores at 30 minutes were, however, significantly lower with IN ketamine as compared to placebo. The need for rescue analgesics was significantly lower with IN ketamine as compared to placebo. There was no significant difference in the incidence of dizziness and nausea/vomiting between IN ketamine and opioids. As compared to placebo, IN ketamine was associated with an increased incidence of dizziness but not nausea/vomiting. Emergence reactions were significantly increased with IN ketamine as compared to opioids and placebo. CONCLUSIONS: There may be a role of IN ketamine for acute pain management in adults in an emergency setting. There is a tendency for better pain control with IN ketamine as compared to control and the possibility of similar efficacy of IN ketamine as compared to opioids. However, the results are not unequivocal and are limited by the low number of studies in literature and limited pain indications studied. Further RCTs are required to strengthen the evidence.

Bioinformatics analysis of candidate genes for milk production traits in water buffalo (Bubalus bubalis).

Water buffalo (Bubalus bubalis) is an important livestock species in developing countries due to its contribution to meat, milk production, and a certain form of labor. However, the genetic potential of buffalo milk production traits has not been fully exploited. To date, 516 candidate genes associated with milk production traits of buffalo have been identified. The present study aimed to explore the possible molecular mechanisms underlying milk production traits of this species through functional genomics analysis of these candidate genes by using different bioinformatics tools. Gene ontology (GO) analysis indicated that these candidate genes were associated with complex biological processes, such as cell proliferation and mitotic nuclear division. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis demonstrated that these candidate genes were enriched in multiple signaling pathways, such as AMPK, ErbB, Toll-like receptor, and Jak-STAT. In addition, one function module consisting of 57 nodes and 139 edges were identified from the protein-protein interaction (PPI) network. GO analysis showed that the 57 candidate genes in this function module were enriched in three main biological processes, including homeostasis, metabolism, and cell response. These three distinct biological processes are well known for regulating mammary gland activities, which explained clearly the mechanism underlying milk production traits. This study provides a novel perspective for better understanding of the biological processes linked with milk production traits. This knowledge is conducive to the improvement of milk yield and composition of this species.

Systematic analyses for candidate genes of milk production traits in water buffalo (Bubalus Bubalis).

Water buffalo (Bubalus bubalis) is of great economic importance as a provider of milk and meat in many countries. However, the milk yield of buffalo is much lower than that of Holstein cows. Selection of candidate genes related to milk production traits can be applied to improve buffalo milk performance. A systematic review of studies of these candidate genes will be greatly beneficial for researchers to timely and efficiently understand the research development of molecular markers for buffalo milk production traits. Here, we identified and classified the candidate genes associated with buffalo milk production traits. A total of 517 candidate genes have been identified as being associated with milk performance in different buffalo breeds. Nineteen candidate genes containing 47 mutation sites have been identified using the candidate gene approach. In addition, 499 candidate genes have been identified in six genome-wide association studies (GWASes) including two studies performed with the bovine SNP chip and four studies with the buffalo SNP chip. Genes CTNND2 (catenin delta 2), APOB (apolipoprotein B), FHIT (fragile histidine triad) and ESRRG (estrogen related receptor gamma) were identified in at least two GWASes. These four genes, especially APOB, deserve further study to explore regulatory roles in buffalo milk production. With growth in the number of buffalo genomic studies, more candidate genes associated with buffalo milk production traits will be identified. Therefore, future studies, such as those investigating gene location and functional analyses, are necessary to facilitate the exploitation of genetic potential and the improvement of buffalo milk performance.

Integrating RNA-seq and GWAS reveals novel genetic mutations for buffalo reproductive traits.

Genome-wide association study (GWAS) has been applied in buffalo breeding programs and been used to identify a number of candidate genes associated with buffalo reproductive traits. The genetic code of specific genes underlying buffalo reproductive traits remains unclear. Association study that measures both genetic and transcriptional variation has been applied for the investigation of complex traits. To investigate genes involved in buffalo reproductive traits, integrated RNA-seq results were investigated of buffalo granulosa cells and candidate genes which were reported to be associated with buffalo reproductive traits in a previous GWAS. A large number of variants were detected by RNA-seq, and 214 variants were located within the buffalo reproductive candidate genes identified by GWAS. A further association study in 462 Italian Mediterranean buffalo indicated that 25 SNPs distributed in 13 genes were associated with reproductive traits. Of the 13 genes, 11 were expressed in granulosa cells of all antral follicle development stages, and significant difference was found in the expression of NDUFS2 between follicles of diameter <8 mm and > 8 mm. These findings extend the results of GWAS by expanding the knowledge about new and potentially functional single-nucleotide polymorphisms and provide useful information about regulatory genes affecting buffalo reproductive traits.

DAMP molecular IL-33 augments monocytic inflammatory storm in hepatitis B-precipitated acute-on-chronic liver failure.

BACKGROUND & AIMS: Patients with acute-on-chronic liver failure (ACLF) usually exhibit defective monocyte function and excessive systemic inflammatory response. Interleukin-33 (IL-33) acts as a danger-associated molecular pattern (DAMP) to modulate immune response. However, the role of IL-33 in regulating monocyte function during hepatitis B-precipitated ACLF (HB-ACLF) in response to lipopolysaccharide (LPS) has not been clear. METHODS: In this study, the levels of IL-33/ST2 in blood and liver samples collected from patients with HB-ACLF, chronic hepatitis B (CHB) and normal controls and the associated of those findings with disease severity were analysed. HLA-DR and CD80 expression, phagocytosis capacity, cytokine secretion and MAP kinase activation induced by LPS were detected to explore the role of IL-33/ST2 signal in regulating monocyte function in patients. RESULTS: The expression levels of IL-33/ST2 were significantly increased in peripheral blood and livers of patients with HB-ACLF, as compared with patients with CHB and controls. It was found that serum IL-33 level was associated with severity of liver disease. Treatment with IL-33 on monocytes significantly increased HLA-DR, CCR2 and CD80 expression, enhanced LPS-stimulated TNF-α, IL-6 and IL-1ß secretion, but did not affect the phagocytic capacity. Furthermore, IL-33 signalling enhanced the ERK1/2 activation of monocytes in response to LPS. CONCLUSIONS: DAMP molecular IL-33 augmented the 'storm' of monocytic inflammation in response to LPS through ERK1/2 activation during HB-ACLF.

The four and a half LIM domains 2 (FHL2) regulates ovarian granulosa cell tumor progression via controlling AKT1 transcription.

The four and a half LIM domains 2 (FHL2) has been shown to play important roles in the regulation of cell proliferation, survival, adhesion, motility and signal transduction in a cell type and tissue-dependent manner. However, the function of FHL2 in ovarian physiology and pathology is unclear. The aim of this study was to determine the role and functional mechanism of FHL2 in the progression of ovarian granulosa cell tumors (GCTs). Immunohistochemical analysis indicated that FHL2 was overexpressed in GCT tissues. Cellular localization of FHL2 in GCT cells was cell cycle dependent. Knockdown of FHL2 suppressed GCT cell growth, reduced cell viability and inhibited cell migration. Consistently, ectopic expression of FHL2 in GCT cells with very low endogenous FHL2 promoted cell growth, improved cell viability and enhance cell migration. Importantly, overexpression of FHL2 promoted GCT progression in vivo. Mechanistic studies indicated that FHL2 regulates AKT1 gene expression in vitro and in vivo. Knockdown of FHL2 or AKT1 in GCT cell lines induced very similar phenotypes. Ectopic expression of constitutively active AKT1 rescued FHL2 knockdown-induced arrest of GCT cell growth and reduction of GCT cell viability, suggesting that FHL2 regulates GCT cell growth and viability through controlling AKT1 expression. Finally, co-immunoprecipitation and chromatin immunoprecipitation analyses indicated that FHL2 functions as a co-activator of NFκB and AP-1 to regulate AKT1 gene transcription. In conclusion, results from the present study indicate that FHL2 exerts its oncogenic action in GCT cells via controlling AKT1 gene expression. FHL2 is a promising target for the development of novel drugs against ovarian granulosa cell tumor.

YAP induces high-grade serous carcinoma in fallopian tube secretory epithelial cells.

Accumulating evidence indicates that ovarian high-grade serous carcinoma (HGSC) originates from fallopian tube secretory epithelial cells (FTSECs). However, the molecular mechanisms underlying the initiation and progression of HGSC derived from FTSECs remains unclear. In this study, we found that the Hippo/Yes-associated protein (YAP) signaling pathway has a critical role in the initiation and progression of fallopian tube and ovarian HGSC. Importantly, YAP was overexpressed in inflammatory and cancerous fallopian tube tissues. Further, overexpression of wild-type YAP, or constitutively active YAP in immortalized FTSECs, induced cell proliferation, migration, colony formation and tumorigenesis. Moreover, the Hippo/YAP and the fibroblast growth factor (FGF) signaling pathways formed an autocrine/paracrine-positive feedback loop to drive the progression of the FTSEC-derived HGSC. Evidence in this study strongly suggests that combined therapy with inhibitors of YAP (such as verteporfin) and FGF receptors (such as BGJ398) can provide a novel therapeutic strategy to treat fallopian tube and ovarian HGSC.

Runoff characteristics and non-point source pollution analysis in the Taihu Lake Basin: a case study of the town of Xueyan, China.

Non-point source pollution is a significant environmental issue in small watersheds in China. To study the effects of rainfall on pollutants transported by runoff, rainfall was monitored in Xueyan town in the Taihu Lake Basin (TLB) for over 12 consecutive months. The concentrations of different forms of nitrogen (N) and phosphorus (P), and chemical oxygen demand, were monitored in runoff and river water across different land use types. The results indicated that pollutant loads were highly variable. Most N losses due to runoff were found around industrial areas (printing factories), while residential areas exhibited the lowest nitrogen losses through runoff. Nitrate nitrogen (NO3-N) and ammonia nitrogen (NH4-N) were the dominant forms of soluble N around printing factories and hotels, respectively. The levels of N in river water were stable prior to the generation of runoff from a rainfall event, after which they were positively correlated to rainfall intensity. In addition, three sites with different areas were selected for a case study to analyze trends in pollutant levels during two rainfall events, using the AnnAGNPS model. The modeled results generally agreed with the observed data, which suggests that AnnAGNPS can be used successfully for modeling runoff nutrient loading in this region. The conclusions of this study provide important information on controlling non-point source pollution in TLB.

YAP forms autocrine loops with the ERBB pathway to regulate ovarian cancer initiation and progression.

Mechanisms underlying ovarian cancer initiation and progression are unclear. Herein, we report that the Yes-associated protein (YAP), a major effector of the Hippo tumor suppressor pathway, interacts with ERBB signaling pathways to regulate the initiation and progression of ovarian cancer. Immunohistochemistry studies indicate that YAP expression is associated with poor clinical outcomes in patients. Overexpression or constitutive activation of YAP leads to transformation and tumorigenesis in human ovarian surface epithelial cells, and promotes growth of cancer cells in vivo and in vitro. YAP induces the expression of epidermal growth factor (EGF) receptors (EGFR, ERBB3) and production of EGF-like ligands (HBEGF, NRG1 and NRG2). HBEGF or NRG1, in turn, activates YAP and stimulates cancer cell growth. Knockdown of ERBB3 or HBEGF eliminates YAP effects on cell growth and transformation, whereas knockdown of YAP abrogates NRG1- and HBEGF-stimulated cell proliferation. Collectively, our study demonstrates the existence of HBEGF & NRGs/ERBBs/YAP/HBEGF & NRGs autocrine loop that controls ovarian cell tumorigenesis and cancer progression.

Effects of plant roots on the hydraulic performance during the clogging process in mesocosm vertical flow constructed wetlands.

The aim of this study was to evaluate the effects of plant roots (Typha angustifolia roots) on the hydraulic performance during the clogging process from the perspective of time and space distributions in mesocosm vertical flow-constructed wetlands with coarse sand matrix. For this purpose, a pair of lab-scale experiments was conducted to compare planted and unplanted systems by measuring the effective porosity and hydraulic conductivity of the substrate within different operation periods. Furthermore, the flow pattern of the clogging process in the planted and unplanted wetland systems were evaluated by their hydraulic performance (e.g., mean residence time, short circuiting, volumetric efficiency, number of continuously stirred tank reactors, and hydraulic efficiency factor) in salt tracer experiments. The results showed that the flow conditions would change in different clogging stages, which indicated that plants played different roles related to time and space. In the early clogging stages, plant roots restricted the flow of water, while in the middle and later clogging stages, especially the later stage, growing roots opened new pore spaces in the substrate. The roots played an important role in affecting the hydraulic performance in the upper layer (0-30 cm) where the sand matrix had a larger root volume fraction. Finally, the causes of the controversy over plant roots' effects on clogging were discussed. The results helped further understand the effects of plant roots on hydraulic performance during the clogging process.

Polymorphisms in the bone morphogenetic protein 15 gene and their effect on sperm quality traits in Chinese Holstein bulls.

Bone morphogenetic protein 15 (BMP-15) expression has been detected in the testis, but its roles in this organ has not been well elucidated. We evaluated polymorphisms of the BMP-15 gene by PCR-SSCP and PCR-RFLP in 212 Chinese Holstein bulls, and investigated possible associations with sperm quality traits, including semen volume per ejaculate, sperm density, fresh sperm motility, thawed sperm motility, acrosome integrity rate, and abnormal sperm rate. A single nucleotide polymorphism (C5697T) in intron 1 of the BMP-15 gene was identified in these bulls. Age was found to have significant effects on both fresh sperm motility and abnormal sperm rate. A significant effect of genotype on fresh sperm motility was also observed. Least square analysis showed that CT genotype bulls had significantly lower fresh sperm motility than CC or TT genotype bulls. In conclusion, BMP-15 should be considered as a potential genetic marker for sperm quality, based on its association with fresh sperm motility.

The G-protein-coupled estrogen receptor agonist G-1 suppresses proliferation of ovarian cancer cells by blocking tubulin polymerization.

The G-protein-coupled estrogen receptor 1 (GPER) has recently been reported to mediate the non-genomic action of estrogen in different types of cells and tissues. G-1 (1-[4-(6-bromobenzo[1,3] dioxol-5yl)-3a,4,5,9b-tetrahydro-3H-cyclopenta[c]quinolin-8-yl]-ethanone) was developed as a potent and selective agonist for GPER. G-1 has been shown to induce the expression of genes and activate pathways that facilitate cancer cell proliferation by activating GPER. Here we demonstrate that G-1 has an anticancer potential with a mechanism similar to vinca alkaloids, the commonly used chemotherapy drugs. We found that G-1 blocks tubulin polymerization and thereby interrupts microtubule assembly in ovarian cancer cells leading to the arrest of cell cycle in the prophase of mitosis and the suppression of ovarian cancer cell proliferation. G-1 treatment also induces apoptosis of ovarian cancer cells. The ability of G-1 to target microtubules to suppress ovarian cancer cell proliferation makes it a promising candidate drug for treatment of ovarian cancer.

An integrated model of substrate clogging in vertical flow constructed wetlands.

This paper presents an integrated model of substrate clogging in a vertical flow constructed wetland (VFCW). The model simulates the reduction of pore space in the wetland substrate due to combined influences of various physical, biogeochemical and plant-related processes. A series of experiments based on laboratory-scale VFCWs were conducted to examine and measure key parameters related to clogging of the wetland substrate during operation under different conditions. The model was then validated using data collected from the experiments. The results showed that the model was able to replicate the clogging phenomenon as observed in the experiments, in particular, the characteristic clogging time. The model also predicted well individual contributions to clogging by accumulated inert suspended solids, microbial biomass and plant root materials during the wetland operation. Although the validation was based on the laboratory data, the results indicated that the model describes well the processes underlying the clogging and has the potential to become a tool for assessing the performance of prototype CWs in relation to clogging at both the design and operation stages.

Effects of the polymorphisms of Mx1, BAT2 and CXCL12 genes on immunological traits in pigs.

It is necessary that genetic markers or biomarkers can be used to predict resistance towards a wide range of infectious diseases. In the present study, we estimated the potential markers and measured their relationship with heritabilities of a wide range of immune traits. Polymorphisms in exon 13 of Mx1, intron 25 of BAT2 and intron 3 of CXCL12 were identified by sequencing, and the genotypes were analyzed by PCR-RFLP in a resource population composed of 352 pure breed Landrace piglets at days 0, 17 and 32 after birth. Associations of single-nucleotide polymorphisms (SNPs) in these genes with a variety of immunological traits and antibody levels for pig reproduction and porcine respiratory syndrome virus (PRRSV), pseudorabies virus (PRV) and classical swine fever virus (CSFV) were performed. The performance of GG genotype of BAT2 on hemoglobin concentration (HBG) and hematocrit (HCT) of piglets at day 0 was significantly higher than that of the AA and AG individuals. For Mx1, compared with CT genotype, the pigs with TT or CC generated more PRRS antibody at day 0. The piglets with CT genotype had highly significant difference of PRV antibody from those with CC and TT genotypes at day 0. And the piglets with CC genotype had higher level red blood cell count (RBC), hemoglobin concentration (HBG) and hematocrit (HCT) than those with CT and TT genotypes at day 17. For the C7462G SNP in the intron 3 of CXCL12, the PRV antibody level of piglets with the CG genotype were higher than that of piglets with CC and GG genotypes at day 17, and the mean corpuscular volume (MCV) of GG piglets were larger than that of CC and CG individuals at day 0. At the locus 7331 bp in the intron 3 of CXCL12, there were significantly differences of mean corpuscular hemoglobin concentrations (MCHC) at day 0 and white blood cell count (WBC) at day 32, which showed the trend GG or AG>AA, AA>AG or GG, respectively. The pigs with AA or GG genotype had more platelet distribution width (PDW), mean platelet volume (MPV) and platelet-large cell ratio (PLR) at day 17 than those with AG. The results of this study indicated that polymorphisms in Mx1, BAT2 and CXCL12 genes were significantly associated with the immunological traits in Landrace piglets and had potential application value for marker-assisted selection of pig breeding with disease resistance.

Identification of complex vertebral malformation carriers in Holstein cattle in south China.

Complex vertebral malformation (CVM) is a recently described monogenic autosomal recessive hereditary defect of Holstein dairy cattle that causes premature birth, aborted fetuses and stillborn calves. Guanine is substituted by thymine (G>T) in the solute carrier family 35 member A3 gene (SLC35A3). A valine is changed to a phenylalanine at position 180 of uridine 5'-diphosphate-N-acetyl-glucosamine transporter protein. CVM is expected to occur in many countries due to the widespread use of sire semen. We developed a created restriction site PCR (CRS-PCR) method to diagnose CVM in dairy cows. This was tested on 217 cows and 125 bulls selected randomly from a Holstein cattle population in south China. Five Holstein cows and five Holstein bulls were identified to be CVM carriers; the percentages of CVM carriers were estimated to be 2.3, 4.0 and 2.9% in the cows, bulls and entire Holstein cattle sample, respectively.

An immunological method to screen sex-specific proteins of bovine sperm.

This study was designed to identify sex-specific antibodies (SSAb) in rabbit antisera against bovine sex-sorted sperm, and capture sex-specific proteins of bovine X- or Y- proteins by SSAb. The rabbit antisera against bovine X- or Y-sperm were first produced by a series of immunological approaches, and further purified through immuno-neutralization with excess sex-sorted Y- or X-sperm, respectively, to remove non-sex specific antibodies and enrich sex-specific antibodies. After removal of non-sex specific antibodies, the purified rabbit sera with enriched sex-specific antibodies were screened for sex-specific antibodies by immunofluorescence staining and flow cytometry. The results showed that 3.0, 2.2, and 4.2% of unsorted sperm, sex-sorted X-sperm, and sex-sorted Y-sperm were recognized by the purified rabbit antisera against Y-sperm, respectively, whereas 29.2, 19.7, and 3.9% of unsorted sperm, sex-sorted X-sperm, and sex-sorted Y-sperm were recognized by the purified rabbit antisera against X-sperm. These results suggested that the purified rabbit antisera against X-sperm contained SSAb that preferentially bound to sex-sorted X-sperm. Subsequently, the purified rabbit antisera against X- or Y-sperm were used to immunoprecipitate sex-specific proteins in bovine sperm proteins, and a 30-kDa protein was specifically captured by the rabbit antisera against X-sperm. In conclusion, our results implied that this 30-kDa protein might be a sex-specific protein in bovine X-sperm, which has the potential to be used in immunological procedures for sexing sperm.

N-Acylhomoserine lactone quorum-sensing signalling in antagonistic phenazine-producing Pseudomonas isolates from the red cocoyam rhizosphere.

Forty fluorescent Pseudomonas strains isolated from white and red cocoyam roots were tested for their ability to synthesize N-acyl-l-homoserine lactones (acyl-HSLs). Remarkably, only isolates from the red cocoyam rhizosphere that were antagonistic against the cocoyam root rot pathogen Pythium myriotylum and synthesized phenazine antibiotics produced acyl-HSLs. This supports the assumption that acyl-HSL production is related to the antagonistic activity of the strains. After detection, the signal molecules were identified through TLC-overlay and liquid chromatography-multiple MS (LC-MS/MS) analysis. In our representative strain, Pseudomonas CMR12a, production of the signal molecules could be assigned to two quorum-sensing (QS) systems. The first one is the QS system for phenazine production, PhzI/PhzR, which seemed to be well conserved, since it was genetically organized in the same way as in the well-described phenazine-producing Pseudomonas strains Pseudomonas fluorescens 2-79, Pseudomonas chlororaphis PCL1391 and Pseudomonas aureofaciens 30-84. The newly characterized genes cmrI and cmrR make up the second QS system of CMR12a, under the control of the uncommon N-3-hydroxy-dodecanoyl-homoserine lactone (3-OH-C12-HSL) and with low similarity to other Pseudomonas QS systems. No clear function could yet be assigned to the CmrI/CmrR system, although it contributes to the biocontrol capability of CMR12a. Both the PhzI/PhzR and CmrI/CmrR systems are controlled by the GacS/GacA two-component regulatory system.