Protective Effects of 28-O-Caffeoyl Betulin (B-CA) on the Cerebral Cortex of Ischemic Rats Revealed by a NMR-Based Metabolomics Analysis.

28-O-caffeoyl betulin (B-CA) has been demonstrated to reduce the cerebral infarct volume caused by transient middle cerebral artery occlusion (MCAO) injury. B-CA is a novel derivative of naturally occurring caffeoyl triterpene with little information associated with its pharmacological target(s). To date no data is available regarding the effect of B-CA on brain metabolism. In the present study, a 1H-NMR-based metabolomics approach was applied to investigate the therapeutic effects of B-CA on brain metabolism following MCAO in rats. Global metabolic profiles of the cortex in acute period (9 h after focal ischemia onset) after MCAO were compared between the groups (sham; MCAO + vehicle; MCAO + B-CA). MCAO induced several changes in the ipsilateral cortex of ischemic rats, which consequently led to the neuronal damage featured with the downregulation of NAA, including energy metabolism dysfunctions, oxidative stress, and neurotransmitter metabolism. Treatment with B-CA showed statistically significant rescue effects on the ischemic cortex of MCAO rats. Specifically, treatment with B-CA ameliorated the energy metabolism dysfunctions (back-regulating the levels of succinate, lactate, BCAAs, and carnitine), oxidative stress (upregulating the level of glutathione), and neurotransmitter metabolism disturbances (back-regulating the levels of γ-aminobutyric acid and acetylcholine) associated with the progression of ischemic stroke. With the administration of B-CA, the levels of three phospholipid related metabolites (O-phosphocholine, O-phosphoethanolamine, sn-glycero-3-phosphocholine) and NAA improved significantly. Overall, our findings suggest that treatment with B-CA may provide neuroprotection by augmenting the metabolic changes observed in the cortex following MCAO in rats.

Insufficient CD100 shedding contributes to suppression of CD8+ T-cell activity in non-small cell lung cancer.

CD100 is an immune semaphorin constitutively expressed on T-cells. Matrix metalloproteinase (MMP) is an important mediator of membrane-bound CD100 (mCD100) cleavage to generate soluble CD100 (sCD100), which has immunoregulatory activity in immune cell responses. The aim of the study was to investigate the level and role of sCD100 and mCD100 in modulating CD8+ T-cell function in non-small cell lung cancer (NSCLC). sCD100 and MMP-14 levels in the serum and bronchoalveolar lavage fluid (BALF), and mCD100 expression on peripheral and lung-resident CD8+ T-cells were analysed in NSCLC patients. The ability to induce sCD100 and the effect of MMP-14 on mCD100 shedding for the regulation of non-cytolytic and cytolytic functions of CD8+ T-cells were also analysed in direct and indirect contact co-culture systems. NSCLC patients had lower serum sCD100 and higher mCD100 levels on CD8+ T-cells compared with healthy controls. BALF from the tumour site also had decreased sCD100 and increased mCD100 on CD8+ T-cells compared with the non-tumour site. Recombinant CD100 stimulation enhanced non-cytolytic and cytolytic functions of CD8+ T-cells from NSCLC patients, whereas blockade of CD100 receptor CD72 attenuated CD8+ T-cell activity. NSCLC patients had lower MMP-14 in the serum and in BALF from the tumour site. Recombinant MMP-14 mediated mCD100 shedding from CD8+ T-cell membrane, and led to promotion of CD8+ T-cell response in NSCLC patients. Overall, decreased MMP-14 resulted in insufficient CD100 shedding, leading to suppression of peripheral and lung-resident CD8+ T-cell activity in NSCLC.

Infection deteriorating hepatitis B virus related acute-on-chronic liver failure: a retrospective cohort study.

BACKGROUND: Infection is common in acute-on-chronic liver failure (ACLF), which may worsen the clinical condition and prognosis. However, the characteristics of infection and its influence on prognosis in hepatitis B virus related ACLF (HBV-ACLF) as defined by the European Association for the Study of the Liver (EASL) have not been clarified. We aimed to investigate the characteristics of infection and its influence on mortality in patients with HBV-ACLF defined by EASL in China. METHODS: We performed a retrospective cohort study in patients with HBV-ACLF defined by EASL in a single center from January 2015 to December 2017. These patients were divided into two groups with and without infection. The incidence, sites of infection, isolated strains, and risk factors associated with mortality were evaluated. RESULTS: A total of 289 patients were included, among them 185 (64.0%) were diagnosed with an infection. The most common type of infection was pneumonia (55.7%), followed by spontaneous bacterial peritonitis (47.6%) and others. The gram-negative bacteria were the most frequent (58.3%). Patients with one, two, and three or more infection sites had a gradually increasing incidence of sepsis (P < 0.01), septic shock (P < 0.001), and ACLF-3 (P < 0.05). Also, patients with infection isolated one, two, and three or more strains showed a growing incidence of sepsis (P < 0.01) and septic shock (P < 0.001). Patients with infection showed a significantly higher 28-day mortality than those without (P < 0.01), especially in patients with ACLF-3. Infection was identified as an independent risk factor for 28-day mortality in all HBV-ACLF patients. Pneumonia and sepsis were identified as independent predictors of 28-day mortality for patients with infection. CONCLUSIONS: Infection is associated with severe clinical course and high mortality in HBV-ACLF defined by EASL. The increased number of infection sites or isolated strains was associated with the occurrence of sepsis and septic shock. Pneumonia and sepsis were independent predictors for mortality in HBV-ACLF patients with infection.

M860, a Monoclonal Antibody against Human Lactoferrin, Enhances Tumoricidal Activity of Low Dosage Lactoferrin via Granzyme B Induction.

Lactoferrin (LF) is a soluble glycoprotein of the transferring family found in most biological fluids, functioning as a major first line defense molecule against infection in mammals. It also shows certain anti-tumor activity, but its clinical application in tumor therapy is limited because high dosage is required. In this study, we demonstrate that M860, a monoclonal antibody against human LF (hLF), could significantly increase the anti-tumor potential of low dosage hLF by forming LF-containing immune complex (IC). Human monocytes primed with LF-IC, but not hLF or M860 alone, or control ICs, showed strong tumoricidal activity on leukemia cell lines Jurkat and Raji through induction of secreted Granzyme B (GzB). LF-IC is able to colligate membrane-bound CD14 (a TLR4 co-receptor) and FcγRIIa (a low affinity activating Fcγ receptor) on the surface of human monocytes, thereby triggering the Syk-PI3K-AKT-mTOR pathway leading to GzB production. Our work identifies a novel pathway for LF-mediated tumoricidal activity and may extend the clinical application of LF in tumor therapy.

Interleukin-35 Suppresses the Antitumor Activity of T Cells in Patients with Non-Small Cell Lung Cancer.

BACKGROUND/AIMS: Interleukin (IL)-35 has immunosuppressive functions in autoimmune diseases, infectious diseases, and certain cancers. However, few studies have focused on its immunoregulatory activity in non-small cell lung cancer (NSCLC). Thus, we investigated the role of IL-35 in the pathogenesis of this disease. METHODS: A total of 66 NSCLC patients and 21 healthy individuals were enrolled. IL-35 expression in peripheral blood and bronchoalveolar lavage fluid (BALF) was measured. The modulatory functions of IL-35 on purified CD4+ and CD8+ T cells from NSCLC patients were investigated in direct and indirect coculture systems with NSCLC cell lines. RESULTS: IL-35 expression was significantly increased in BALF from the tumor site, but not in the peripheral blood of NSCLC patients. IL-35 did not affect the bioactivity including proliferation, cytokine production, cell cycle, and cellular invasion of NSCLC cells. It suppressed responses from type 1 T helper (Th1) and Th17 cells but elevated the regulatory T cell response in cultured CD4+ T cells from NSCLC patients, and reduced cytokine-mediated CD4+ T cells cytotoxicity to NSCLC cells. Moreover, IL-35 also inhibited cytotoxic gene expression in CD8+ T cells from NSCLC, reducing their cytolytic and noncytolytic functions. CONCLUSION: The results of this study suggest that IL-35 contributes to the dysfunction/exhaustion of T cells and limited antitumor immune responses in NSCLC.

Dual anti-ischemic effects of rosmarinic acid n-butyl ester via alleviation of DAPK-p53-mediated neuronal damage and microglial inflammation.

The discovery of efficacious anti-ischemic drugs remains a challenge. Recently we have found that rosmarinic acid n-butyl ester (RABE), a derivative of rosmarinic acid, significantly protects SH-SY5Y cells against oxygen glucose deprivation (OGD)-induced cell death. In the present study we simultaneously investigated the effects of RABE on the two key players in the pathophysiology of cerebral ischemia, ischemic neuronal damage and microglial inflammation. Pretreatment with RABE (1, 10 µmol/L) dose-dependently attenuated OGD- or H2O2-induced reduction of the viability of SH-SY5Y neuroblastoma cells. RABE pretreatment concurrently reduced the apoptotic cell rate, down-regulated the expression of the pro-apoptotic proteins Bax and p53, and up-regulated the expression of the anti-apoptotic protein phosphorylated death-associated protein kinase (DAPK). Furthermore, pretreatment with RABE (3 µmol/L) markedly inhibited lipopolysaccharide (LPS)-induced increases in the release of TNF-α, IL-1ß, NO and PGE2, and the expression levels of iNOS, and COX-2 in cultured rat microglial cells. In conclusion, these results reveal for the first time the potential anti-ischemic effects of RABE on neuronal and glial cells and elucidate the molecular mechanisms involved in its dual beneficial profiles in vitro. RABE may be a promising drug lead/candidate for the treatment of ischemic stroke.

Caffeoyl Triterpenoid Esters as Potential Anti-ischemic Stroke Agents from Celastrus orbiculatus.

Three new triterpenoids, celastrusins A-C (1-3), together with 3-O-caffeoyl-α-amyrin (4) were isolated from the root bark of Celastrus orbiculatus. Their structures were identified by spectroscopic analysis, X-ray crystallography using Cu Kα radiation, and the comparison of both observed and reported spectroscopic data. An in vitro bioassay revealed that the caffeoyl triterpenoid esters 1, 3, and 4 possess neuroprotective effects against oxygen-glucose deprivation (OGD) induced SH-SY5Y cell damage. Further animal studies indicated that compound 1 significantly reduced brain infarction after transient middle cerebral artery occlusion (MCAO) in rats using a 10 mg/kg (i.v.) dose.

A novel caffeoyl triterpene attenuates cerebral ischemic injury with potent anti-inflammatory and hypothermic effects.

Despite the intense efforts in searching for stroke therapies, an urgent need still exists to explore novel neuroprotective agents for ischemic stroke that have high efficacy and wide therapeutic time-window. Here, we provide the first demonstration that 28-O-caffeoyl betulin (B-CA), a novel derivative of naturally occurring caffeoyl triterpene, could significantly alleviate brain infarction and neurological deficit when given as late as 6 h after transient middle cerebral artery occlusion in the rat. Moreover, post-ischemia B-CA administration exhibited long-term (14 days post stroke) protective effects on both brain infarction and functional (i.e., motor and sensory) deficits. Protective B-CA effects correlated with decreased inflammatory responses as indicated by inhibition of microglia and astrocyte activation [stained with ionized calcium-binding adapter molecule 1 (Iba-1) and glial fibrillary acidic protein (GFAP) antibody, respectively], as well as suppression of tumor necrosis factor-α, interleukin-1ß, and cyclooxygenase-2 overproduction in the ipsilateral cortex of ischemic rat. B-CA administration caused significant hypothermia in the focal cerebral ischemic rat, which may contribute to its ameliorative effects on brain damage and inflammation. In view of its potency in wide therapeutic time-window, robust anti-inflammatory and hypothermic effects, this novel caffeoyl triterpene derivative may lead toward the development of effective therapeutic strategies for the treatment of ischemic stroke.

Lithocarpic acids O-S, five homo-cycloartane derivatives from the cupules of Lithocarpus polystachyus.

Chemical investigation of the cupules of Lithocarpus polystachyus resulted in the identification of four 3,4-seco-homo-cycloartane and one homo-cycloartane derivatives named lithocarpic acids O-S. Their structures were determined based on extensive 1D/2D NMR, IR, MS spectroscopic analyses and chemical methods. Lithocarpic acid O (1) exhibited inhibitory activities on mouse and human isozymes of 11ß-hydroxysteroid dehydrogenase type 1 (11ß-HSD1) with IC50 values of 0.49 and 1.1 µM, respectively.

Four new triterpenoids isolated from the resin of Garcinia hanburyi.

Four new triterpenoids, 2-O-acetyl-3-O-(4'-O-acetyl)-α-l-arabinopyranosylmaslinic acid (1), 2-O-acetyl-3-O-(3'-O-acetyl)-α-l-arabinopyranosylmaslinic acid (2), 2-O-acetyl-3-O-(3',4'-O-diacetyl)-α-l-arabinopyranosylmaslinic acid (3), and 3-O-(3'-O-acetyl)-α-l-arabinopyranosyloleanolic acid (4), together with six known triterpenoids, 3-O-(4'-O-acetyl)-α-l-arabinopyranosyloleanolic acid (5), maslinic acid (6), 2-O-acetylmaslinic acid (7), 3-O-acetylmaslinic acid (8), betulinic acid (9), and 2α-hydroxy-3ß-O-acetylbetulinic acid (10), were isolated from the EtOAc extract of Garcinia hanburyi resin. Their structures were elucidated by analysis of the spectroscopic data and chemical methods.

(M)- and (P)-bicelaphanol A, dimeric trinorditerpenes with promising neuroprotective activity from Celastrus orbiculatus.

(M)-Bicelaphanol A (1) and (P)-bicelaphanol A (2), two unprecedented dimeric trinorditerpenes existing as atropisomers, together with their monomer celaphanol A (3), were isolated from the root bark of Celastrus orbiculatus. The structures and absolute configurations of 1 and 2 were determined by spectroscopic and single-crystal X-ray diffraction analyses. Compound 1 exhibited a significant in vitro neuroprotective effect against a hydrogen peroxide-induced cell viability decrease in PC12 cells at 1 µM, while compounds 2 and 3 showed such effects at 10 µM.

Membrane Bound CRT Fragment Accelerates Tumor Growth of Melanoma B16 Cell In Vivo through Promoting M2 Polarization via TLR4.

Calreticulin (CRT) is a major calcium-binding luminal resident protein on the endoplasmic reticulum that can also be released extracellular as well as anchored on surface of cells. Previously, we demonstrated that soluble recombinant CRT fragment 39-272 (CRT/39-272) exhibited potent immunostimulatory effects as well as immunoregulation effects on immune cells. Here, we constructed stable B16 melanoma cell lines expressing recombinant CRT/39-272 on the membrane (B16-tmCRT/39-272) to investigate the roles of cell surface CRT on tumor progression. We found that B16-tmCRT/39-272 cells subcutaneously inoculated into C57BL/6 mice exhibited stronger tumorigenicity than the B16-EGFP control cells. The tumor associated macrophages infiltrated in tumors were mainly M2 phenotype. Regulatory T cells (Tregs) were also expanded more in bearing mice. Consistent with the in vivo results, B16-tmCRT/39-272 promoted macrophage polarization toward F4/80+CD206+ M2 macrophages and promoted transforming growth factor beta (TGF-ß) secretion in vitro, which could promote naïve CD4+ T cell differentiation into Tregs. These results imply that the tmCRT/39-272 could accelerate tumor development by enhancing M2 macrophage polarization to induce TGF-ß secretion, and then promoted Treg differentiation in the tumor microenvironment. Our data may provide useful clues for better understanding of the potentiating roles of CRT in tumorigenesis.

Altered Phenotype and Enhanced Antibody-Producing Ability of Peripheral B Cells in Mice with Cd19-Driven Cre Expression.

Given the importance of B lymphocytes in inflammation and immune defense against pathogens, mice transgenic for Cre under the control of Cd19 promoter (Cd19Cre/+ mice) have been widely used to specifically investigate the role of loxP-flanked genes in B cell development/function. However, impacts of expression/insertion of the Cre transgene on the phenotype and function of B cells have not been carefully studied. Here, we show that the number of marginal zone B and B1a cells was selectively reduced in Cd19Cre/+ mice, while B cell development in the bone marrow and total numbers of peripheral B cells were comparable between Cd19Cre/+ and wild type C57BL/6 mice. Notably, humoral responses to both T cell-dependent and independent antigens were significantly increased in Cd19Cre/+ mice. We speculate that these differences are mainly attributable to reduced surface CD19 levels caused by integration of the Cre-expressing cassette that inactivates one Cd19 allele. Moreover, our literature survey showed that expression of Cd19Cre/+ alone may affect the development/progression of inflammatory and anti-infectious responses. Thus, our results have important implications for the design and interpretation of results on gene functions specifically targeted in B cells in the Cd19Cre/+ mouse strain, for instance, in the context of (auto) inflammatory/infectious diseases.

[The effect on anti oxidative damage with aspirin pretreatment in primary cultured rat type II alveolar epithelial cell].

OBJECTIVE: To investigate the protective effect of aspirin on primary cultured type II alveolar epithelial cell (AEC II), and the mechanism of its effect on anti oxidation damage. METHODS: The original generation of adult rat AEC II were cultured and purified. They were divided into normal saline (NS) group, hydrogen peroxide injury group (H(2)O(2) group), and 1, 2, 3 aspirin pretreatment groups (A1-3 groups).In H(2)O(2) group, 0.5 mmol/L H(2)O(2) was added to AEC II after 40 hours of culture to reproduce a cell oxidative injury model. In NS group, only NS was added to AEC II culture. To the A1-3 groups aspirin 50, 100 and 200 µmol/L were added respectively. Cell form, cell count and cell survival rate were observed at 3 hours after H(2)O(2) was given . Immunohistochemical and polymerase chain reaction (PCR) methods were used for the determination of heme oxygenase-1 (HO-1) protein and HO-1 mRNA (20, 40, 60 hours of culture). RESULTS: With trypsin digestion and immune adherence method AEC II could be harvested (2.0-2.5)× , and the purity and activity were both over 90%. Compared with NS group, gaps between cells were widened in H(2)O(2) group, cell account was reduced, and the survival rate (A value) was reduced significantly (0.054 6±0.004 0 vs. 0.103 8±0.009 9, P<0.01). Compared with H(2)O(2) group, in A1-3 groups the number of adherent cells was increased, cell morphology was intact, and no obvious cell shrinkage was found. Higher survival rate (A value) was found in A1-3 groups than that of H(2)O(2) group (0.066 9±0.003 9, 0.071 0±0.006 5, 0.078 7±0.009 2 vs. 0.054 6±0.004 0, all P<0.01). Compared with NS group, HO-1 protein and HO-1 mRNA expression in AEC II after 20, 40 and 60 hours of culture reached peak level at 60 hours, and they were increased significantly in A1-3 groups [protein (A value): 1.59±0.12, 1.60±0.09, 1.61±0.08 vs. 1.25±0.11; mRNA (the ratio of Ct value: 24.31±1.74, 30.45± 2.53, 32.63±3.74 vs. 22.99±1.95, all P<0.05]. There was no significant difference in HO-1 protein expression among A1-3 groups. CONCLUSION: There are significant protective effects of aspirin against anti oxidative damage in cultured AEC II cell. As expression of HO-1 is increased in aspirin groups, it may be considered as a protective factor against anti oxidative damage in AEC II cell culture.

[The role of heme oxygenase 1 in hydrogen peroxide induced apoptosis and mitochondrial trans membrane potential change in rat primary type II alveolar epithelium cells].

OBJECTIVE: To understand the role of heme oxygenase-1 (HO-1) in hydrogen peroxide [H(2)O(2)] induced apoptosis and mitochondrial trans-membrane potential (MTMP) change in primary alveolar epithelial cell type II(AEC II). METHODS: Primary AEC II collected from healthy Sprague Dawley (SD) rats were cultured for 24 hours, then divided into four groups to be treated with: (1) saline; (2) H(2)O(2) (0.5 mmol/L); (3) H(2)O(2) +HO-1 (0.2 mmol/L); (4) H(2)O(2) +zinc original porphyrin IX (HO-1 inhibitor, 20 µmol/L). The morphology of cells in the cultures was examined by fluorescent microscopy 2.5 hours later, and the number of apoptotic cells / the MTMP determined by flow-cytometry 0.5, 1.0, 1.5, 2.0 and 2.5 hours later. RESULTS: Large number of cells in with green (early apoptotic) or red (later apoptotic) fluorescence were observed by microscope in cultures treated with H(2)O(2) , and H(2)O(2) + HO-1 inhibitor, but such cells were obviously fewer in HO-1 treated cultures. Compared with saline treated cells, H(2)O(2) treated cells had significantly higher apoptosis rate, that increased with time, reaching peak value 2.5 hours into the treatment [0.5 hour: (30.27 ± 0.74)% vs. (3.76 ± 0.81)%, 2.5 hours: (40.46 ± 0.91)% vs. (22.74 ± 0.60)%, both P < 0.05], while the rate of MTMP depolarization was significantly lower (0.99 ± 0.21 vs. 1.91 ± 0.16, P < 0.05) in these cells. Compared with H(2)O(2) treated cells, the apoptosis rate in HO-1 treated cells was significantly lower [0.5 hour: (5.99 ± 0.60)% vs. (30.27 ± 0.74)%, 2.5 hours: (22.69 ± 1.69)% vs. (40.46 ± 0.91)%, both P < 0.05], and their rate of MTMP depolarization higher (2.02 ± 0.12 vs. 0.99 ± 0.21, P < 0.05). Compared with HO-1 treated cells, HO-1 inhibitor treated cells had significantly higher apoptosis rate which reached peak value 2.5 hours into the treatment [0.5 hour: (30.73 ± 1.08)% vs. (5.99 ± 0.60)%, 2.5 hours: (41.38 ± 0.57)% vs. (22.69 ± 1.69)%, both P < 0.05], while rate of MTMP depolarization in these cells was significantly lower (0.98 ± 0.09 vs. 2.02 ± 0.12, P < 0.05). CONCLUSION: HO-1 could maintain the integrity of AEC II and stabilize their mitochondria membrane potential, protecting the cells from H(2)O(2) induced damage.

Botanical Volatiles Selection in Mediating Electrophysiological Responses and Reproductive Behaviors for the Fall Webworm Moth Hyphantria cunea.

Host-plant volatiles play vital roles for insects to locate foraging, mating, and oviposition sites in the environment. As one of the devastating invasive forestry pests, Hyphantria cunea causes a great annual loss in China, and understanding its chemical ecology is an important task. The current research was done in terms of chemical analysis, electrophysiology, and behavioral assays on H. cunea to assess its olfactory reception toward host-plant volatiles. A screen of possible common host volatiles was done, targeting on five favored hosts of H. cunea, harvesting six potential bioactive compounds from a total of 78 odorant components. Six types of antennal sensilla were investigated on their distributions on the antennae, and sexual dimorphism was described. H. cunea showed responses to all selected host-related volatiles in electroantennogram tests, and linalyl butyrate elicited the strongest responses. Furthermore, mating rates in adult pairs that are exposed to dibutyl phthalate and phytol have been significantly increased, while oviposition rates and female fecundity were not influenced. The results of the current study provide initial evidence showing that universal host-derived volatile cues are essential for H. cunea moth in terms of mating, which can also provide insights into the development of botanical attractants.

[Effects of protease inhibitors on the activities of midgut proteases in Callosobruchus chinensis (Linnaeus) larvae.] / è›‹ç™½é ¶æŠ‘åˆ¶å‰‚å¯¹ç»¿è±†è±¡å¹¼è™«ä¸­è‚ è›‹ç™½é ¶æ´»æ€§çš„å½±å“.

To clarify the effects of protease inhibitors on the activities of midgut proteases in Callosobruchus chinensis (L.) larvae, the inhibitory effects of four kinds of protease inhibitors on the activities of midgut proteases of C. chinensis larvae were examined in vitro and in feeding conditions with indoor artificial insect inoculation. The activities of total protease, trypsin-like enzyme and chymotrypsin-like enzyme in the midgut of C. chinensis larvae fed on artificial mung bean that contained different contents of mung bean types in inhibitor (MBTI) were examined. The results showed that those four protease inhibitors had significant inhibitory effects on the activities of total protease, trypsin-like enzyme and chymotrypsin-like enzyme in C. chinensis larvae. The inhibitory effect was more significant when the concentration was higher. Among those inhibitors, the inhibitory effect of 20 µg·mL-1 MBTI on three kinds of enzyme activities was the strongest, with reduction of 62.5%, 41.2% and 38.7%, respectively. Ovomucoid inhibitor (OI) had the lowest inhibitory effect. The activities of three enzymes in the midgut were also inhibited by C. chinensis larvae fed with artificial mung bean containing different inhibitors. The activities of three enzymes increased with the prolongation of the instar after feeding, but they were significantly lower than that in control. The inhibitory effect of MBTI was the strongest. The inhibitory effect on the activities of total protease and trypsin-like enzyme was gradually enhanced with the increases of MBTI contents when C. chinensis larvae were fed with artificial mung bean with different contents MBTI. The inhibitory effect on chymotrypsin-like enzyme activity was not significant. When the content of MBTI was up to 20%, the activity of chymotrypsin-like enzyme was obviously inhibited.

[Identifying group G beta-hemolytic Streptococci strains by 16S rDNA sequencing].

OBJECTIVE: To identify the DNA sequences of 26 beta-hemolytic group G streptococci strains isolated from the swabs of patients in a glomerulonephritis outbreak event. METHODS: API20 Strep system and 16S rDNA sequencing were performed on all of the 26 isolated strains respectively. The dendrogram based on the analysis of the 16S rDNA sequences of four groups of G streptococci strains was constructed by the neighbor-joining method and boot-strap test. RESULTS: The API20 Strep system failed to identify the strains with less than 62% of success rates. All of the 26 group G beta-hemolytic streptococci isolates were identified as Streptococcus anginosus by the 16S rDNA sequencing, with more than 97% of success rates. The dendrogram illustrated that the four groups of G streptococci strains were clustered at the same branch with Streptococcus anginosus. CONCLUSION: The 16S rDNA sequencing has unique advantages over the phenotypic methods. It can provide detailed information of nucleotides and identify species of beta-hemolytic G group streptococci.