Genomic analysis and characterization of bacteriophage vB_SpuS_NX263 infecting Salmonella enterica subsp. enterica serovar Pullorum.

In this study, a new Salmonella phage, NX263, was isolated from sewage. This phage could lyse 90.57% (48/53) of the bacterial strains tested and showed good activity over a wide range of temperature (up to 60°C) and pH (5-10). Phylogenetic analysis showed that it should be classified as a member of the genus Skatevirus. The genome of phage NX263 is 46,574 bp in length with a GC content of 45.52%. It contains 89 open reading frames and two tRNA genes. No lysogeny, drug resistance, or virulence-associated genes were identified in the genome sequence, suggesting that this phage could potentially be used to treat Salmonella Pullorum infections.

[Genetic study of an X-linked agammaglobulinemia pedigree caused by an BTK mutation].

OBJECTIVE: To explore the genetic pathogenesis of X-linked agammaglobulinemia in two patients for clinical diagnosis and family counseling. METHODS: Data was collected from the patients' family including clinical information, blood immunoglobulin level, as well as classification and subgrouping of B lymphocytes. Gene mutations were screened by whole exome sequencing (WES) through next-generation sequencing (NGS), the result was verified with Sanger sequencing. RESULTS: A BTK c.1627T>C (p.Ser543Pro) variant was found in the pedigree. The phenotype and variant have co-segregated in the pedigree. The variant was not found in population database. The variant has affected in the kinase domain which contained no benign variants and is harmful as predicted through bioinformatic analysis. CONCLUSION: BTK c.1627T>C (p.Ser543Pro) is a pathogenic variant contributing to X-linked agammaglobulinemia in this pedigree. Above finding has provided reproduction guidance for this family.

Epidemiology, molecular characterization, and recombination analysis of chicken anemia virus in Guangdong province, China.

Chicken anemia virus (CAV) causes severe anemia and immunosuppression in young chickens and a compromised immune response in older birds, resulting in great economic losses to the poultry industry worldwide. Here, we report the molecular epidemiology and characterization of CAV circulating in poultry in Guangdong province, China. Ninety-one of 277 chickens collected from 2016 to 2017 were CAV positive. Full-genome sequencing revealed the presence of eight separate strains. Phylogenetic analysis based on the genome sequences obtained in this study and related sequences available in the GenBank database showed that all of the CAV isolates exhibit a close relationship to each other and belong to the same genotypic group. Putative recombination events were also detected in the genomes of the newly isolated CAVs. Collectively, our findings underscore the importance of CAV surveillance and provide information that will lead to a better understanding of the evolution of CAV.

Characterization of Neoschoengastia gallinarum from subtropical China by rDNA and identification of two genotypes based on mitochondrial cox1.

Genetic variations in the 18S ribosomal DNA (18S), 28S ribosomal DNA (28S), second internal transcribed spacer of ribosomal DNA (ITS2), and mitochondrial cytochrome c oxidase subunit 1 (cox1) of Neoschoengastia gallinarum collected from subtropical China were examined. First, a portion of the 18S (p18S), a portion of the 28S (p28S), and the complete ITS2 were separately amplified from individual mites and sequenced. The lengths of the sequences of p18S, p28S, and ITS2 were found to be 1379 bp, 3465~3468 bp, and 200 bp, respectively. The intraspecific sequence variation was 0~0.1% for p28S and 0~1.6% for ITS2, though no variation was observed for p18S, suggesting conservation of rDNA sequences. Second, a portion of the mitochondrial cox1 gene (pcox1) of N. gallinarum was analyzed. The length of the pcox1 sequence is 460 bp, and two distinct groups were observed in N. gallinarum. All pcox1 sequences in group I were identical, and there was only one nucleotide transition observed in group II; however, 7.0~7.2% variations between the two groups were observed, suggesting that two genotypes of N. gallinarum: genotype I and genotype II. Phylogenetic analyses based on pcox1 sequences indicated that N. gallinarum isolates (genotype I or genotype II) clustered into one branch; according to cox1 sequence analysis of Trombiculidae, Walchia hayashii is the closest species. The present study shows that ITS2 rDNA sequence can act as marker for the identification of N. gallinarum samples. Furthermore, analysis of the mitochondrial pcox1 sequence suggests the existence of two genotypes, which has implications for further studies of the ecology and population genetic structures of N. gallinarum.

[Determination of flavonoids constituents in epimedium total flavone capsule and research on its anti-osteoporosis activity].

To develop a rapid resolution liquid chromatography (RRLC) method for the simultaneous determination of epimedoside A, epimedin A1, epimedin A, epimedin B, epimedin C, icariin, baohuosideⅡ, icarisideⅠ, sagittatoside B, 2"-O-rhamnosyl icarisideⅡ, and baohuosideⅠin epimedium total flavone capsule. At the same time, the effects of the above 11 compounds on osteogenic differentiation of MC3T3-E1 cells were investigated by detecting the content of alkaline phosphatase (AKP). The results showed that baohuoside Ⅱ had the highest activities, and both the activities of baohuoside Ⅱ and icariside Ⅰ were stronger than those of icariin.In this study, the content determination method of flavonoid glycosides was established, and the anti-osteoporosis effect of monomers was compared, providing technical support for the study of the pharmacodynamic and mechanism of Epimedium total flavone capsule.

Epidemiological investigation of Mycoplasma Synoviae in native chicken breeds in China.

BACKGROUD: Mycoplasma synoviae (M. synoviae) is widely distributed around the world, and leads to serious economic losses in the world every year. Nevertheless, the incidence and epidemiology of M. synoviae infection in China have remained unclear. RESULTS: In this study we demonstrate that over 9773 broiler chicken flocks in 16 Chinese provinces were affected by M. synoviae between 2010 and 2015. Our epidemiological study revealed that M. synoviae was widely prevalent in multi-aged Chinese native breeder chickens, and the prevalence of M. synoviae in embryos of breeders reached up to 16.29%. In addition, our data showed that chickens aged 14 days or younger carried simultaneously high levels of maternal antibody against M. synoviae and high M. synoviae infection (10%), and low M. synoviae antibody levels in breeders and high proportion of M. synoviae infection in embryos could increase the chances of incidence in the offspring. Finally, our results also indicated that 3- to 7-week-old chickens might be most the susceptible to M. synoviae and, therefore, might play a key role in the horizontal transmission of M. synoviae. CONCLUSION: Our findings suggest that M. synoviae is widely circulating in Chinese native chickens, accordingly, effective control measures are urgently needed to control the spread.

Simultaneous determination of 35 ultra-trace level organophosphorus pesticide residues in Sanjie Zhentong capsules of traditional Chinese medicine using ultra high performance liquid chromatography with tandem mass spectrometry.

The aim of this study was to develop an analytical method for the simultaneous determination of ultra-trace levels of 35 organophosphorus pesticide residues in Sanjie Zhentong capsules, a traditional Chinese medicine prescription. A method based on multiclass and multiresidue sample preparation was developed. First, samples were hydrated with water at 4°C. A ratio of 1:3 sample/water was used for each of the sample amounts. Then, different extraction solvents were screened. This step was followed by a dispersive solid-phase extraction clean-up procedure using both primary secondary amine and polyamide. A comprehensive sensitive multiresidue liquid chromatography tandem mass spectrometry method was investigated and validated. Good linearity was achieved in the range of 10-500 µg/kg for each analyte. The average recovery ranged from 70 to 120%, except for methamidophos, fonophos, diazinon, and chlorpyriphos-ethyl, the recoveries of which ranged from 60-70% at the lower concentration level. The precision values were lower than 10% for all the compounds in three concentration levels. The limits of detection and limits of quantification values were 0.01-2.1 and 0.05-3.4 µg/kg, respectively. The matrix effects were determined, and most of the compounds showed signal suppression. Finally, this optimized procedure was applied for the analysis of real samples.

[Rapid analysis on chemical constituents in Yinyanghuo Zonghuangtong capsule by UPLC/Q-TOF-MS/MS].

To analyze and identify the chemical constituents from Yinyanghuo Zonghuangtong capsule by using UPLC/Q-TOF-MS/MS. Chromatographic separation was carried out on an Agilent SB-C18 column (2.1 mm×100 mm, 1.8 µm) at the temperature of 30 ℃. The mobile phase was 0.1% formic acid and acetonitrile by gradient elution, with a flow rate at 0.30 mL•min⁻¹, and the injection volume of 2 µL. The MS spectrum was acquired in both negative and positive ion modes using ESI ion source. Based on relative accurate molecular weight, secondary mass spectrometry pyrolysis fragments and chromatographic peak retention time, as well as fragmentation regularity summarized from reference substance and the literature, we could effectively identify the chemical structure of the components under test. As a result, a total of 46 compounds, including flavonoid glycosides, phenolic acids and alkaloids, were successfully identified or tentatively predicted. The results provide technical support for quality control and late-stage clinical application of Yinyanghuo Zonghuangtong capsule, and reference for further expound the pharmacodynamic material basis.

[Rapid identification of chemical components in Congrong Zonggan capsule by UPLC-Q-TOF-MS/MS].

This study was aimed to qualitatively analyze the chemical components in Congrong Zonggan capsule by using ultra-performance liquid chromatography tandem quadrupole time-of-flight mass spectrometry method (UPLC-Q-TOF-MS/MS). An Agilent SB-C18 Rapid Resolution HD (3.0 mm×100 mm,1.8 µm) was used with acetonitrile (A) - 0.1% formic acid solution (B) as the mobile phase for gradient elution. The flow rate was 0.2 mL•min⁻¹; the detection wavelength was set at 330 nm and the column temperature was maintained at 30 ℃. Electrospray ion (ESI) source was applied for the qualitative analysis under the negative ion mode. Finally, based on comparison with standard samples, database matching analysis and reviewing relevant literature, 41 compounds were identified from Congrong Zonggan capsule. This method could be used to rapidly detect the chemical components in Congrong Zonggan capsule, providing reference for the quality control of Congrong Zonggan capsule and laying a foundation for the further study on active components mechanism.

[The agreement and clinical value of hepatic vein pressure gradient and portal vein pressure in patients with portal hypertension].

OBJECTIVE: To evaluate the agreement and correlation between hepatic vein pressure gradient (HVPG) and portal vein pressure (PVP) in patients with portal hypertension,and explore their clinical value. METHODS: A total of 46 patients with portal hypertension were directly measured the free hepatic pressure, wedged hepatic pressure, portal vein pressure before and after TIPS therapy. The agreement and correlation of HVPG and PVP were analyzed, and explore their clinical value. RESULTS: There is no significant agreement or correlation between HVPG and PVP in 5 patients, whose third hilar have large communicating branches between portal vein and Inferior vena cava, or with obvious umbilical vein opened. The HVPGs were significantly agreed with portal vein pressure in other 41 patients. There is no significant difference of HVPG or PVP between earlyTIPS and not early-TIPS groups. In addition, the portal vein pressures after TIPS were significantly decreased compared with that before TIPS. CONCLUSION: The HVPG can well show the PVP except these with obvious communicating branches between portal vein and Inferior vena cava in third hilar, and TIPS can effectively decrease the portal vein pressure in patients with portal hypertension.

[Research on ultrafine grinding technology of improving dissolution rates of effective components in Sanjie Zhentong capsule].

The effects of ultrafine grinding on the dissolution rates of the effective components in Sanjie Zhentong capsule (SZC) were studied in this experiment. Fine and ultrafine powder of SZC intermediates were made by ordinary grinding and ultrafine grinding technology, and then granulated by wet granulation. SZC were prepared by fine powder, ultrafine powder and ultrafine granules, respectively. With resveratrol and loureirin B as investigated indexes, dissolution rates of the four intermediates in SZC were determined by cup method and HPLC. The dissolution rates of resveratrol in SZC prepared by fine powder, ultrafine powder and ultrafine granules were 26.11%, 63.27%, 67.49%, respectively; and the dissolution rates of loureirin B were 7.160%, 20.29%, 23.05%, respectively. The dissolution rate of resveratrol and loureirin B in SZC prepared by ultrafine granules was the best. D90 size of ultrafine grinding was 13.221 µm and could improve the dissolution rates of resveratrol and loureirin B in SZC.

[Fingerprint analysis of Resina Draconis from different manufactuers by UPLC coupled with chemometrics].

This study is to establish an UPLC fingerprint of Resina Draconis from different manufacturers, which can provide a comprehensive evaluation for its quality control. The analysis was performed on a Phenomenex Kinetex 2.6 µ C18 100A column by agradientelution program with acetonitrile-water as mobile phase at a flow rate of 1.7 mL x min(-1). The column temperature was 40 degrees C and the detection wavelengthwas 280 nm. The fingerprints of 18 batches of Draconis Resina were further evaluated by chemometrics methods including similarity analysis (SA), hierarchical clustering analysis (HCA) and principal component analysis (PCA). As a result, there were 15 common peaks, 13 of which had been identified by LC-Q-TOF MS, and the similarity degrees of 15 batches of the samples was more than 0.9, and the samples were divided into 4 clusters by their quality difference. The method is reproducible, simple and reliablethat it can be used for quality control and evaluation of Resina Draconis from different manufacturers.

Isolation and characterization of a subtype C avian metapneumovirus circulating in Muscovy ducks in China.

Subtype C avian metapneumovirus (aMPV-C), is an important pathogen that can cause egg-drop and acute respiratory diseases in poultry. To date, aMPV-C infection has not been documented in Muscovy ducks in China. Here, we isolated and characterized an aMPV-C, designated S-01, which has caused severe respiratory disease and noticeable egg drop in Muscovy duck flocks in south China since 2010. Electron microscopy showed that the isolate was an enveloped virus exhibiting multiple morphologies with a diameter of 20-500 nm. The S-01 strain was able to produce a typical cytopathic effect (CPE) on Vero cells and cause death in 10- to 11-day-old Muscovy duck embryos. In vivo infection of layer Muscovy ducks with the isolate resulted in typical clinical signs and pathological lesions similar to those seen in the original infected cases. We report the first complete genomic sequence of aMPV-C from Muscovy ducks. A phylogenetic analysis strongly suggested that the S-01 virus belongs to the aMPV-C family, sharing 92.3%-94.3% of nucleotide identity with that of aMPV-C, and was most closely related to the aMPV-C strains isolated from Muscovy ducks in France. The deduced eight main proteins (N, P, M, F, M2, SH, G and L) of the novel isolate shared higher identity with hMPV than with other aMPV (subtypes A, B and D). S-01 could bind a monoclonal antibody against the F protein of hMPV. Together, our results indicate that subtype-C aMPV has been circulating in Muscovy duck flocks in South China, and it is urgent for companies to develop new vaccines to control the spread of the virus in China.

Sequence and phylogenetic analysis of surface protein genes of emerging H9N2 influenza viruses isolated from poultry in two geographical regions of China.

Subtype H9N2 avian influenza viruses (AIVs) circulating in China have aroused increasing concerns for their impact on poultry and risk to public health. The present study was an attempt to elucidate the phylogenetic relationship of H9N2 AIVs in two geographically distinct regions of China where vaccination is routinely practiced. A total of 18 emerging H9N2 isolates were identified and genetically characterized. Phylogenetic analysis of hemagglutinin (HA) and neuraminidase (NA) genes confirmed that the isolates belonged to the Y280 lineage. Based on the HA genes, the isolates were subdivided into two subgroups. The viruses from Zhejiang Province were clustered together in Group I, while the isolates from Guangdong Province were clustered together in Group II. Antigenic characterization showed that the tested viruses were antigenically different when compared to the current used vaccine strain. It was notable that 14 out of total 18 isolates had an amino acid exchange (Q→L) at position 216 (226 by H3 Numbering) in the receptor-binding site, which indicated that the virus had potential affinity of binding to human like receptor. These results suggest that the emerging viruses have potential risk to public health than previously thought. Therefore, continuous surveillance studies of H9N2 influenza virus are very important to the prognosis and control of future influenza pandemics.

Retrospective analysis of venograms of hepatic and portal veins: clinical implications for transjugular intrahepatic portosystemic shunt placement.

AIMS: This study was aimed to provide safety guidance of needle passes into the portal vein during transjugular intrahepatic portosystemic shunt (TIPS) placement. METHODOLOGY: On anteroposterior venograms, the orifice of right hepatic trunk (RHT), furcation position, and branching of portal trunk in 128 patients underwent TIPS were mapped in relation to the vertebrae and intervertebral space. Impact of clinical factors on these parameters was determined. RESULTS: The orifices of RHTs were all above the furcation position of portal trunk, RHTs were posterior and superior to portal branch. Of the 128 patients, 84.4% had the orifice positioned between 9th and 10th thoracic (T) vertebrae, 58.6% positioned to T10. Portal trunks were furcated between T11 and T12 in 80.5% patients. Portal trunks were bifurcated at right hepatic portal in 91.4% patients into left and right veins, and trifurcated into right posterior, right anterior and left branches in 8.6% patients. Statistical analysis indicated that these parameters were not affected by cause of disease, gender, age and Child-Pugh score. CONCLUSIONS: The puncture site for portal vein located at or beyond imaged trunk furcation would be safe for most of the patients.

Abdominal pain caused by Tibetan medicine: A case report of lead poisoning.

Nowadays, lead poisoning in children commonly occurs, but lead poisoning caused by the administration of Tibetan medicine is rarely reported. This report describes the diagnosis and management of lead poisoning in a 16-year-old girl presented with abdominal pain, vomiting, and anemia with limb numbness, who had a childhood history of epilepsy and took Tibetan medicine intermittently to control the symptoms. After admission into hospital, Computed tomography showed high-density shadows in the gastrointestinal tract. Video-Electroencephalography showed no signs of seizure. Reflux esophagitis was observed in gastroscopy. And no obvious abnormalities in the colonic mucosa through colonoscopy. Bone marrow smear test showed basophilic stippling in the erythrocytes. The blood and urine lead levels of 626 and 75.9 µg/L, respectively. We therefore considered lead poisoning, and the patient improved after chelation therapy. Due to its atypical clinical manifestations, lead poisoning is easily misdiagnosed. Thus, clinicians should pay more attention to this disease. When abdominal pain, anemia, and neurological symptoms are present, the possibility of lead poisoning should be considered.

Dietary Supplementation with Bupleuri Radix Reduces Oxidative Stress Occurring during Growth by Regulating Rumen Microbes and Metabolites.

Oxidative stress (OS) in ruminants is closely associated with disease; thus, improving antioxidant capacity is an important strategy for maintaining host health. Bupleuri Radix (BR) could significantly improve host health and stress levels. However, the clear antioxidant mechanism of the function of BR remains unknown. In the current study, LC-MS metabolomics combined with 16S rRNA gene sequencing was employed to explore the effects of BR on rumen microbiota and metabolites in Shanbei Fine-Wool Sheep (SFWS), and Spearman correlation analyses of rumen microbiota, metabolites, and OS were performed to investigate the mechanism of antioxidant function of BR. Our results indicated that as SFWS grows, levels of OS and antioxidant capacity increase dramatically, but providing BR to SFWS enhances antioxidant capacity while decreasing OS. Rumen microbiota and OS are strongly correlated, with total antioxidant capacity (T-AOC) showing a significant negative correlation with Succiniclasticum and a positive correlation with Ruminococcus. Importantly, the Chao1 index was significantly negatively correlated with malondialdehyde (MDA) and positively correlated with superoxide dismutase (SOD) and T-AOC. Two biomarkers connected to the antioxidant effects of BR, 5,6-DHET and LPA (a-25:0/0:0), were screened according to the results of metabolomics and Spearman analysis of rumen contents, and a significant relationship between the concentration of rumen metabolites and OS was found. Five metabolic pathways, including glycerolipid, glutathione, nucleotide, D-amino acid, and inositol phosphate metabolism, may have a role in OS. The integrated results indicate that rumen microbiota and metabolites are strongly related to OS and that BR is responsible for reducing OS and improving antioxidant capacity in post-weaned SFWS. These findings provide new strategies to reduce OS occurring during SFWS growth.

Complete genome sequence of a recombinant nephropathogenic infectious bronchitis virus strain in China.

Recently, nephropathogenic infectious bronchitis virus (IBV) outbreaks have occurred in commercial broiler flocks and have been associated with a high incidence and morbidity in China. The CK/CH/Zhejiang/06/10 strain (IBV-YX10) was isolated from a 12-day-old broiler chicken in a flock of chickens with swollen speckled kidneys and distended ureters filled with uric acid in China in 2010. Here we reported the complete genomic sequence of the IBV-YX10 which was a natural recombinant nephropathogenic infectious bronchitis virus strain. These findings will contribute additional insights into the molecular characteristics of evolving IBV genomes and the need for effective control of IBV in China.

Complete genome sequence analysis of a natural reassortant infectious bursal disease virus in China.

A novel isolate of infectious bursal disease virus (IBDV) was designated GX-NN-L. The GX-NN-L IBDV was a very virulent infectious bursal disease virus (vvIBDV) isolated from broiler flocks in Guangxi province, China, in 2011. The GX-NN-L IBDV caused high mortality, immunosuppression, low weight gain, and bursal atrophy in commercial broilers. Here, we report the complete genome sequence of the GX-NN-L IBDV, a reassortment strain with segments A and B derived from very virulent strains and attenuated IBDV, respectively. These findings from this study provide additional insights into the genetic exchange between attenuated and very virulent strains of IBDV and continuous monitoring of the spread of the virus in chicken.

Complete genome sequence of a Newcastle disease virus strain isolated from broiler breeder flocks in China.

In 2010 and 2011, several devastating Newcastle disease (ND) outbreaks occurred in China, affecting broilers, layers, and breeders. The CK-JSX1-201005 virus was isolated from broiler breeder flocks vaccinated with the classical ND virus (NDV) vaccine program, but laying rate decreased from 80% to 30 to 40% in the clinic. Here, we report the complete genome sequence and molecular characteristic of the CK-JSX1-201005 NDV. These findings provide additional insights into the genetic variation of NDV circulating in China and are useful for vaccine development for NDV.