An Integrated Metagenomics/Metaproteomics Investigation of the Microbial Communities and Enzymes in Solid-state Fermentation of Pu-erh tea.

Microbial enzymes during solid-state fermentation (SSF), which play important roles in the food, chemical, pharmaceutical and environmental fields, remain relatively unknown. In this work, the microbial communities and enzymes in SSF of Pu-erh tea, a well-known traditional Chinese tea, were investigated by integrated metagenomics/metaproteomics approach. The dominant bacteria and fungi were identified as Proteobacteria (48.42%) and Aspergillus (94.98%), through pyrosequencing-based analyses of the bacterial 16S and fungal 18S rRNA genes, respectively. In total, 335 proteins with at least two unique peptides were identified and classified into 28 Biological Processes and 35 Molecular Function categories using a metaproteomics analysis. The integration of metagenomics and metaproteomics data demonstrated that Aspergillus was dominant fungus and major host of identified proteins (50.45%). Enzymes involved in the degradation of the plant cell wall were identified and associated with the soft-rotting of tea leaves. Peroxiredoxins, catalase and peroxidases were associated with the oxidation of catechins. In conclusion, this work greatly advances our understanding of the SSF of Pu-erh tea and provides a powerful tool for studying SSF mechanisms, especially in relation to the microbial communities present.

Novel mutations in DAX1 of X-linked adrenal hypoplasia congenita over several generations in one family.

OBJECTIVE: X-linked adrenal hypoplasia congenital (AHC) is a rare disorder caused by mutations in DAX1 gene. We report a case of X-linked AHC in a large family to analyze the pathogenesis of this rare disease and to add to our clinical knowledge of it. METHODS: We describe 3-year-old boy's clinical features and laboratory test results, as well as the patient's nuclear family members' clinical symptoms, especially those with features of adrenal insufficiency. Genomic deoxyribonucleic acid (DNA) was extracted from the patient's and the family members' peripheral blood leukocytes, and the coding region and promoter region of DAX1 were directly sequenced. RESULTS: A 3-year-old boy who was diagnosed with X-linked AHC presented with atypical symptoms, and his laboratory test results revealed elevated serum adrenocorticotropic hormone levels (ACTH) and decreased serum cortisol levels. Three novel mutations were detected in the DAX1 coding sequence in this family: a missense mutation (c.376G>A, p.Val126Met), a synonymous mutation (c.498G>A, p.Arg166Arg), and a nonsense mutation (c.1225C>T, p. Gln409X). CONCLUSIONS: This report describes the familial transmission of AHC over several generations and further expands the number of DAX1 mutations reported in the literature. Early diagnosis and prompt treatment of X-linked AHC are important and may provide a good prognosis.

A serum metabonomic study on the difference between alcohol- and HBV-induced liver cirrhosis by ultraperformance liquid chromatography coupled to mass spectrometry plus quadrupole time-of-flight mass spectrometry.

BACKGROUND: Liver cirrhosis is the fatal consequence of chronic hepatitis, making early diagnosis of liver cirrhosis critical. Liver biopsy is still the standard diagnostic method for liver cirrhosis, although its use in a broad population with alcoholism or hepatitis B virus (HBV) infection remains difficult. In this study, we used a metabonomic approach to detect potential biomarkers for early diagnosis of liver cirrhosis. METHODS: Serum specimens were collected prospectively from normal control subjects (n = 22) and patients with alcoholic cirrhosis (n = 18) or HBV-induced cirrhosis (n = 19). The serum metabonome was analyzed using ultraperformance liquid chromatography (LC)/time-of-flight mass spectrometry (MS) integrated with chemometrics. The acquired LC-MS data were normalized and processed using principal component analysis (PCA) and partial least squares discrimination analysis (PLS-DA). RESULTS: Significant differences in the metabonomics among the three groups were observed. Lysophosphatidyl cholines (LPCs) (LPC C16:0, LPC C18:0, LPC C18:2, LPC C18:3, LPC C20:3, LPC C20:5) were decreased in the serum of patients with hepatic cirrhosis, whereas bile acids (glycocholic acid, glycochenodeoxycholic acid), hypoxanthine, and stearamide were increased in the serum of patients with hepatic cirrhosis. These metabolites are considered "common" biomarkers for hepatic cirrhosis. Oleamide and myristamide were increased in the serum of patients with alcoholic cirrhosis but decreased in those with HBV-induced cirrhosis. These could be specific biomarkers for differential diagnosis between alcohol- and HBV-induced hepatic cirrhosis. CONCLUSIONS: There are significant metabonomic differences between alcohol- and HBV-induced liver cirrhosis. Metabonomics is a top-down systems biology tool for conducting research on clinical problems.

Determination and comparison of γ-aminobutyric acid (GABA) content in pu-erh and other types of Chinese tea.

Two previous studies have reported that pu-erh tea contains a high level of γ-aminobutyric acid (GABA), which is the major inhibitory neurotransmitter in the central nervous system and has several physiological functions. However, two other researchers have demonstrated that the GABA content of several pu-erh teas was low. Due to the high value and health benefits of GABA, analysis of mass-produced pu-erh tea is necessary to determine whether it is actually enriched with GABA. A high-performance liquid chromatography (HPLC) method was developed for the determination of GABA in tea, the results of which were verified by amino acid analysis using an Amino Acid Analyzer (AAA). A total of 114 samples of various types of Chinese tea, including 62 pu-erh teas, 13 green teas, 8 oolong teas, 8 black teas, 3 white teas, 4 GABA teas, and 16 process samples from two industrial fermentations of pu-erh tea (including the raw material and the first to seventh turnings), were analyzed using HPLC. Statistical analysis demonstrated that the GABA content in pu-erh tea was significantly lower than that in other types of tea (p < 0.05) and that the GABA content decreased during industrial fermentation of pu-erh tea (p < 0.05). This mass analysis and comparison suggested GABA was not a major bioactive constituent and resolved the disagreement GABA content in pu-erh tea. In addition, the GABA content in white tea was found to be significantly higher than that in the other types of tea (p < 0.05), leading to the possibility of producing GABA-enriched white tea.

A hevein-like protein and a class I chitinase with antifungal activity from leaves of the paper mulberry.

Paper mulberry (Broussonetia papyrifera, syn. Morus papyrifera L.) is a Chinese traditional medicine and its low-molecular-weight extracts are reported to have antifungal activity. In this study, two proteins (PMAPI and PMAPII) with activity against Trichoderma viride were obtained from paper mulberry leaves with a fast protein liquid chromatography (FPLC) unit. The purification protocol employed (NH(4))(2)SO(4) precipitation, ion-exchange chromatography and hydrophobic-interaction chromatography on FPLC. Molecular masses were 18,798 Da for PMAPI, and 31,178 Da for PMAPII determined by Matrix-assisted laser desorption ionization time-of-flight mass spectrometry. Peptide mapping fingerprint analysis showed that PMAPI has no peptides similar to PMAPII. N-terminal amino acid sequencing revealed that PMAPI is a hevein-like protein, and PMAPII is a class I chitinase. They both had a half-maximal inhibitory concentration (IC50) of 0.1 µg/µL against T. viride. This is the first report of high-molecular-weight extracts with antifungal activity from paper mulberry.

Clinical significance of C-reactive protein values in antibiotic treatment for pyogenic liver abscess.

AIM: To investigate the clinical significance of C-reactive protein (CRP) values in determining the endpoint of antibiotic treatment for liver abscess after drainage. METHODS: The endpoints of antibiotic treatment in 46 patients with pyogenic liver abscess after complete percutaneous drainage were assessed by performing a retrospective study. After complete percutaneous drainage, normal CRP values were considered as the endpoint in 18 patients (experimental group), and normal body temperature for at least 2 wk were considered as the endpoints in the other 28 patients (control group). RESULTS: The duration of antibiotic treatment after complete percutaneous drainage was 15.83 ± 6.45 d and 24.25 ± 8.18 d for the experimental and the control groups, respectively (P = 0.001), being significantly shorter in the experimental group than in the control group. The recurrence rate was 0% for both groups. However, we could not obtain the follow-up data about 3 patients in the control group. CONCLUSION: CRP values can be considered as an independent factor to determine the duration of the antibiotic treatment for pyogenic liver abscess after complete percutaneous drainage.