Role of the fsr Quorum-Sensing System in Enterococcus faecalis Bloodstream Infection.

Enterococcus faecalis is an important intestinal colonizing bacteria and can cause various tissue infections, including invasive blood infection (BI). The annual incidence of E. faecalis BI has been estimated to be ~4.5 per 100,000, with a fatality rate that can reach 20%. However, whether bacterial colonization or invasive infections are tissue based has not been thoroughly studied. In this study, we analyzed 537 clinical isolates from 7 different tissues to identify the key genomic elements that facilitate the colonization and invasive infection of E. faecalis. Comparative genomic analysis revealed that the BI E. faecalis isolates had the largest genome size but the lowest GC content, fsr quorum-sensing system genes were enriched in the BI E. faecalis, and the fsr gene cluster could enhance biofilm formation and serum resistance ability. Our findings also provide deep insight into the genomic differences between different tissue isolates, and the fsr quorum-sensing systems could be a key factor promoting E. faecalis invasion into the blood. IMPORTANCE First, we conducted an advanced study on the genomic differences between colonizing and infecting E. faecalis, which provides support and evidence for early and accurate diagnoses. Second, we discovered that fsr was significantly associated with blood infections, which also provides additional information for studies exploring the invasiveness of E. faecalis. Most importantly, we found that fsr played an important role in both biofilm formation and serum resistance ability in E. faecalis.

Molecular characterization of Cryptococcus neoformans isolated from the environment in Beijing, China.

The molecular type of environmental Cryptococcus neoformans in Beijing was not clear. Our study aims to reveal the molecular characterization of C. neoformans complex from environment in Beijing, China. A total of 435 samples of pigeon droppings from 11 different homes in Beijing were collected from August to November in 2015. Pigeon droppings were inoculated onto caffeic acid cornmeal agar (CACA) to screen C. neoformans complex. Bruker Biotyper matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) was performed for species identification. Serotype and mating type was determined by specific primers. Restriction fragment length polymorphisms of URA5 (URA5-RFLP) were applied to genotype. Multi-locus sequence typing (MLST) was done for further identification and sequence type (ST) determination. Altogether, 81 isolates of C. neoformans AFLP1/VNI were recognized from 435 pigeon droppings in this study. The positive rate for C. neoformans AFLP1/VNI from pigeon droppings in different homes varied from 5.0% to 52.6%, the average was 20.2%. All of these cryptococcal strains were serotype A, MATα. They were genotyped as VNI by URA5-RFLP and were confirmed by MLST. No other molecular types of C. neoformans and Cryptococcus gattii isolates were isolated. Their STs were identified as ST 31 (n = 54, 66.7%), followed by ST 53 (n = 10), ST 191 (n = 8), ST 5 (n = 5), ST 57 (n = 3), and ST 38 (n = 1). We concluded that not only clinical but also environmental isolates of C. neoformans need to be investigated more deeply and more extensively. The virulence difference between ST 5 and ST 31 need to be explored in the future.

[Study on inflammatory effect of toxic raphides from Pinellia ternate and its correlation with macrophages].

OBJECTIVE: To study the toxic mechanism of toxic raphides from Pinellia ternata. METHOD: Mouse peritoneal macrophage in vitro culture model was adopted to study dose-dependent and time-dependent curves of toxic raphides, with TNF-alpha, IL-1beta and IL-6 in supernatant as indexes. Scanning electron microscopy was used to observe the changes in surface morphology of raphides-treated macrophages. Macrophages-neutrophils co-cultured the transport model to study the effect of toxic raphides' stimulation of macrophages on neutrophils migration. RESULT: Toxic raphides' stimulation of macrophages could cause the increase in the levels of TNF-alpha, IL-1beta and IL-6 released, and showed dose dependence and time dependence. Scanning electron microscopy showed that toxic raphides were swallowed by macrophages, with notable cell membrane creases, increase in the number of pseudopods and decrease in integrity of cell membranes, and could significantly induce migration of neutrophils. CONCLUSION: The inflammatory process induced by toxic raphides is mainly mediated by macrophages. The toxic mechanism of toxic raphides from P. ternata is that toxic raphides penetrate into tissues to activate resident macrophages, release phagocytic and inflammatory cytokines, and cause migration of neutrophils, which finally results in acute inflammatory response.

[Effect of Yupingfeng San against OVA-induced allergic asthma in mice].

OBJECTIVE: To observe the effect of Yupingfeng San (YPFS) against OVA-induced allergic asthma in mice. METHOD: Mice were injected with OVA to establish the allergic asthma model. They were abdominally injected with 20 microg OVA on day 0 and 14, and inhaled aerosol 0.5% OVA solution for 20 min for seven days. The blank control group was administrated with equal volume of saline. YPFS groups with different doses were administrated intragastrically with YPFS every day, with the crude drug dosage of 3.25, 6.5, 13 g x kg(-1), respectively. The model group and control group were administrated with equal volume of saline. The positive control group was given intraperitoneally injected with 1 mg x kg(-1) DEX since aerosol inhalation. Blood was drawn after the last OVA aerosol inhalation to count the number of Eosnophils (Eos) in blood and detect IgE in serum; BALF was collected to count the number of cells and classify; right lung tissues were evenly grinded to detect cytokines IL-4 and IFN-gamma, and left upper lung lobes were collected for pathologic histology. RESULT: The level of Eos and IgE in serum increased significantly in the model group, and a large number of Eos were detected in BALF. Histopathological changes in lung showed bronchial serous exudation, tubular epithelial cells exfoliation, tube narrowing, widened alveolar septum, and bronchial periarterial lymphocytes infiltration. Homogenate of lung tissues showed increase of IL-4, and decrease in IFN-gamma/IL-4 ratio. YPFS groups with different doses displayed decrease of Eos in blood and BALF and IgE content in serum, and relief of pathologic changes in above models. Meanwhile, IL-4 content in homogenate of lung tissues decreased, with the increase in IFN-gamma/IL-4 ratio. CONCLUSION: YPFS shows the inhibitory effects on OVA-induced allergic asthma, involving down regulation of Eos and IgE levels in blood of asthma mice, and infiltration of inflammatory cells in lung tissues. Meanwhile, it can reduce IL-4 in lung homogenates, increase IFN-gamma/IL-4, and inhibits Th2 polarization.

The Influence of Government Regulation on Farmers' Green Production Behavior-From the Perspective of the Market Structure.

To clarify the factors influencing the green production behavior of peach farmers, this paper uses the survey data of 741 peach farmers in 19 provinces and cities in China, it uses a multiple ordered probit model to empirically analyze the impact of the government regulations on the green production behavior of peach farmers, from the perspective of the market structure. This paper also analyzes its intermediary role in this process, and it analyzes the possible heterogeneity at both the planting scale and the regional level. The results show the following: (1) Government regulation has a positive and significant impact on the green production behavior of peach farmers. Specifically, government supervision and inspection, alongside green subsidies, can positively promote the implementation of green production behavior by peach farmers, but government publicity and training have not played a good role. (2) The market structure plays a partial intermediary role, rather than a complete intermediary role, in the government regulation affecting the green production behavior of peach farmers. (3) The impact of the government regulation on the green production behavior of peach farmers is heterogeneous. Specifically, compared with small farmers, the impact on large-scale farmers is higher; however, the influence of the three methods of government regulation on the green production behavior of peach farmers varies from region to region. Therefore, in order to promote the implementation of green production, the government should introduce appropriate local policies, strongly support new agricultural business entities, draw clear guidelines for the market, and play the role of "night watchman".

Evaluation of the Performance of a Multiplex Real-Time PCR Assay for the Identification of Aspergillus, Cryptococcus neoformans, and Pneumocystis jirovecii Simultaneously from Sputum in Multicenter.

Purpose: To evaluate the performance of a multiplex real-time polymerase chain reaction (PCR) assay for the simultaneous identification of Aspergillus, Cryptococcus neoformans, and Pneumocystis jirovecii from sputum. Patients and Methods: Sputum samples (n=537) from patients with suspected invasive fungal infection (IFI) were collected from four centers; they were tested by both multiplex real-time PCR assay and DNA sequencing. DNA sequencing was considered as the reference method, and the performance of the multiplex real-time assay was evaluated by determining the sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV). The interference experiment, repeatability, reproducibility, and stability of the multiplex real-time PCR assay were also evaluated. Results: The detection performance of the multiplex real-time assay, compared with that of DNA sequencing, for the three pathogens was as follows: sensitivity, specificity, PPV, and NPV for Aspergillus, Cryptococcus neoformans, and Pneumocystis jirovecii were 99.40%, 98.64%, 97.09%, and 99.73%; 100%, 99.59%, 96.36%, and 100.00%; and 99.28%, 98.50%, 95.80%, and 99.75%, respectively. The consistency of the two methods was almost perfect: the kappa value was between 0.97 and 0.98. The minimum detection limit of the multiplex real-time assay for each of the three pathogens was 1250 copies/mL. Interference experiment showed that blood and normally used antifungal drugs had no effect on the results. No cross-reactivity was detected for any bacteria or fungi. In 40 patients, mixed infections by Aspergillus and/or Cryptococcus neoformans and/or Pneumocystis jirovecii were detected by the multiplex real-time assay. Among these patients, those with acquired immune deficiency syndrome (AIDS) ranked first, with Aspergillus and Pneumocystis mixed infection accounting for 75%. Conclusion: The multiplex real-time PCR assay is fast, sensitive, and specific and has good clinical application prospects.

Super Dominant Pathobiontic Bacteria in the Nasopharyngeal Microbiota Cause Secondary Bacterial Infection in COVID-19 Patients.

Coronavirus disease 2019 (COVID-19) is a respiratory infectious disease responsible for many infections worldwide. Differences in respiratory microbiota may correlate with disease severity. Samples were collected from 20 severe and 51 mild COVID-19 patients. High-throughput sequencing of the 16S rRNA gene was used to analyze the bacterial community composition of the upper and lower respiratory tracts. The indices of diversity were analyzed. When one genus accounted for >50% of reads from a sample, it was defined as a super dominant pathobiontic bacterial genus (SDPG). In the upper respiratory tract, uniformity indices were significantly higher in the mild group than in the severe group (P < 0.001). In the lower respiratory tract, uniformity indices, richness indices, and the abundance-based coverage estimator were significantly higher in the mild group than in the severe group (P < 0.001). In patients with severe COVID-19, SDPGs were detected in 40.7% of upper and 63.2% of lower respiratory tract samples. In patients with mild COVID-19, only 10.8% of upper and 8.5% of lower respiratory tract samples yielded SDPGs. SDPGs were present in both upper and lower tracts in seven patients (35.0%), among which six (30.0%) patients possessed the same SDPG in the upper and lower tracts. However, no patients with mild infections had an SDPG in both tracts. Staphylococcus, Corynebacterium, and Acinetobacter were the main SDPGs. The number of SDPGs identified differed significantly between patients with mild and severe COVID-19 (P < 0.001). SDPGs in nasopharyngeal microbiota cause secondary bacterial infection in COVID-19 patients and aggravate pneumonia. IMPORTANCE The nasopharyngeal microbiota is composed of a variety of not only the true commensal bacterial species but also the two-face pathobionts, which are one a harmless commensal bacterial species and the other a highly invasive and deadly pathogen. In a previous study, we found that the diversity of nasopharyngeal microbiota was lost in severe influenza patients. We named the genus that accounted for over 50% of microbiota abundance as super dominant pathobiontic genus, which could invade to cause severe pneumonia, leading to high fatality. Similar phenomena were found here for SARS-CoV-2 infection. The diversity of nasopharyngeal microbiota was lost in severe COVID-19 infection patients. SDPGs in nasopharyngeal microbiota were frequently detected in severe COVID-19 patients. Therefore, the SDPGs in nasopharynx microbiota might invade into low respiratory and be responsible for secondary bacterial pneumonia in patients with SARS-CoV-2 infection.

[Clinical feature of cryptosporidium infection in HIV/AIDS patients with chronic diarrhea].

OBJECTIVE: To investigate the clinical feature of cryptosporidium infection in HIV/AIDS patients with chronic diarrhea. METHODS: 253 Stool samples were collected from HIV/AIDS patients with chronic diarrhea during Nov.2009 to Dec.2010. The samples were concentrated by Formalin-Ethyl Acetate Sedimentation technique and stained by Modified acid-fast stain (AFS) for the identification of oocysts by microscopy. Divided the cases into three groups according to their CD4 cell counts (< 200, ≥ 200 and < 500, ≥ 500/µl); meanwhile the cases were divided into three groups by their age (20 - 35, 36 - 55, 56 - 69). Analyzed the groups by comparison. RESULTS: The total infection rate of Cryptosporidium in AIDS patients was 12.6% in 253 cases. CD4(+) T-lymphocyte counts was related to the infection rates of cryptosporidium, the difference was statistically significant (χ(2) = 10.33, P < 0.01): the infection rate were 20.4% (20/98), 9.23% (12/130), 0 (0/25) separately. In the 32 Cryptosporidium positive cases, 22 (68.8%) cases stood with diarrhea above 5 times per day, the three kind of CD4 counts level were separately 15 (15/20), 7 (7/12), 0. The difference was no statistically significant (P > 0.05). HIV/AIDS patients with chronic diarrhea who progressed during asymptomatic period, pre-AIDS period, AIDS period, had the infection rate of 0(0/7), 21.3% (19/89), 8.3% (13/157) respectively, the difference was statistically significant (χ(2) = 9.822, P < 0.01); 22 out of 32 Cryptosporidium positive cases in HIV patients were diagnosed with enteritis, the infection rate in urban area and rural area was 6.5% (7/104) and 16.8% (25/149) separately, the difference was statistically significant (χ(2) = 5.596, P < 0.05). Comparing different age groups, Cryptosporidium infection status were separately 7.3% (4/55), 13.4% (22/164), 17.6% (6/34). Each group's comparative difference was no statistically significant (χ(2) = 2.29, P > 0.05). CONCLUSION: The infection rate of cryptosporidium and clinical severity of cryptosporidium infection are statistically correlated with CD4(+) T-lymphocyte counts, with AIDS stage, with HIV associated proctitis.

Clinical characteristics, Cryptococcus neoformans genotypes, antifungal susceptibility, and outcomes in human immunodeficiency virus-positive patients in Beijing, China.

Cryptococcus neoformans is an environmental fungal pathogen that causes opportunistic infections and severe disseminated meningoencephalitis, mainly in immunocompromised patients such as those with acquired immunodeficiency syndrome (AIDS). In this study, the clinical characteristics, treatment protocols, and outcomes of 70 patients with AIDS and Cryptococcus neoformans infection at Beijing Ditan Hospital were retrospectively analyzed. We performed antimicrobial sensitivity tests and multilocus sequence typing (MLST) on C. neoformans isolates from these patients. The most common symptoms were headache (58.6%), fever (54.3%), and high cerebrospinal fluid pressure (≥200 mm H2O) (71.4%). All patients were positive for C. neoformans antigen in blood or cerebrospinal fluid. The CD4 cell counts of 92.8% (65/70) of patients were <100 cells/µL. In total, 74 C. neoformans isolates were obtained from the 70 patients. The 65 isolates that could be typed fell into 12 sequence types (STs) by MLST: ST5, ST31, ST63, ST202, ST237, ST289, ST295, ST296, ST298, ST324, ST337, and ST359. ST5 was the major type, accounting for 78.5% of isolates (51/65). This study comprehensively assessed the clinical and molecular epidemiology of C. neoformans in patients with AIDS and may inform the development of targeted prevention and treatment strategies for immunocompromised patients with C. neoformans infection.

Urinary 8-oxo-7,8-dihydroguanosine levels are elevated in HCV-infected patients.

HCV patients are usually under substantial oxidative stress because of viral infection. A total of 177 patients with HCV infection and 198 age- and sex-matched healthy controls were enrolled in this study. We evaluated the urinary levels of 8-oxo-7, 8-dihydro-2'deoxyguanosine (8-oxodGuo) and 8-oxo-7, 8-dihydroguanosine (8-oxoGuo) in patients with HCV infection and explored the factors affecting the urinary 8-oxodGuo or 8-oxoGuo levels. Biomarkers of liver function, cancer, and inflammation were determined. Nonparametric correlations were used to evaluate the correlation between 8-oxoGuo or 8-oxodGuo and various laboratory biochemical indicators. Results showed that the levels of urinary 8-oxoGuo both in male and female patients with HCV infection were significantly higher than those in healthy controls (both p < 0.0001), while the urinary 8-oxodGuo levels only in male patients with HCV infection were significantly higher than those in healthy controls (p < 0.01). Urinary 8-oxoGuo was significantly associated with the white blood cell count, C-reactive protein level, and 8-oxodGuo level (p = 0.016, p = 0.003, and p = 0.000, respectively). Urinary 8-oxodGuo was significantly associated with the white blood cell count and 8-oxoGuo level (p = 0.018 and p = 0.000, respectively). A regression equation of urinary 8-oxoGuo or 8-oxodGuo was also established using the biomarkers in plasma. The results suggested that patients with a high C-reactive protein level are likely to have high urinary 8-oxoGuo levels as well, which may be useful for assessing the level of inflammation and oxidative stress in HCV patients.Supplemental data for this article is available online at https://doi.org/10.1080/15257770.2021.1961272 .

The novel outer membrane protein from OprD/Occ family is associated with hypervirulence of carbapenem resistant Acinetobacter baumannii ST2/KL22.

Acinetobacter baumannii has become a major healthcare threat that causes nosocomial infections, especially in critically ill patients. The spread of carbapenem-resistant A. baumannii (CRAB) strains has long been a clinical concern. It is important to study the epidemiology and virulence characteristics of different CRAB isolates in order to tailor infection prevention and antibiotic prescribing. In this study, a total of 71 CRAB isolates were collected in the hospital, and clinical characteristics of infections were analyzed. The genomic characteristics and phylogenetic relationships were elucidated based on genome sequencing and analysis. The isolates were assigned to three sequence types (STs, Pasteur) and nine capsular polysaccharide (KL) types, among which ST2/KL22 was the most prevalent CRAB in the hospital. Even though all the ST2/KL22 isolates contained the same reported virulence genes, one specific clade of ST2/KL22 showed more pathogenic in mouse infection model. Complete genomic analysis revealed differences at the oprD locus between the low- and high-virulent isolates. More specifically, a premature stop codon in the low-virulence strains resulted in truncated OprD expression. By evaluating pathogenicity in C57BL/6 J mice, knock-out of oprD in high-virulent isolate resulted in virulence attenuation, and complementing the avirulent strain with full-length oprD from high-virulent isolate enhanced virulence of the former. The oprD gene may be associated with the enhanced virulence of the specific ST2/KL22 clone, which provides a potential molecular marker for screening the hypervirulent A. baumannii strains.

Case Report: Identification of SARS-CoV-2 in Cerebrospinal Fluid by Ultrahigh-Depth Sequencing in a Patient With Coronavirus Disease 2019 and Neurological Dysfunction.

We reported that the complete genome sequence of SARS-Coronavirus-2 (SARS-CoV-2) was obtained from a cerebrospinal fluid (CSF) sample by ultrahigh-depth sequencing. Fourteen days after onset, seizures, maxillofacial convulsions, intractable hiccups and a significant increase in intracranial pressure developed in an adult coronavirus disease 2019 patient. The complete genome sequence of SARS-CoV-2 obtained from the cerebrospinal fluid indicates that SARS-CoV-2 can invade the central nervous system. In future, along with nervous system assessment, the pathogen genome detection and other indicators are needed for studying possible nervous system infection of SARS-CoV-2.

The Diagnostic Value of 3D Power Doppler Ultrasound Combined With VOCAL in the Vascular Distribution of Breast Masses.

RATIONALE AND OBJECTIVES: This study uses a three-dimensional energy Doppler technique combined with the Virtual Organ Computer-aided Analysis (VOCAL) method in order to determine the diagnostic threshold of blood flow index in breast tumors to provide a reference for evaluation and treatment options. MATERIALS AND METHODS: We collected 322 solid lesions which had been operated. Each lesion met the definite pathological diagnosis; collected lesions included 262 cases of benign lesions and 60 cases of malignant lesions. All examinations were performed by using GE LOGIQ E9 with VOCAL software. Volume and four distinct vascular indices of gray mean (MG), power mean, ratio (R), and vascular flow index (VFI) were calculated by using the VOCAL software. Sampling and calculation were repeated three times and the mean value was calculated. RESULTS: The average age and power of the malignant group were greater than those of the benign group, ie p < .01 which had significant differences. The gray mean of the malignant group was lower than that of the benign group, ie p > .05 which had no significant differences between benign and malignant groups. The ratio, vascular flow index and volume had significant differences, i.e. p < .01. The area under the receiver operating characteristic curve (AUC) were 0.864, 0.830, 0.800, 0.758, and 0.764 for age, power, ratio, vascular flow index, and volume, respectively. The research indicators were higher than 50% of the curve showing their diagnostic value. The cut-off points of age, power, ratio, vascular flow index, and volume were 37.5, 26.56, 0.031, 0.846, and 1.75, respectively. Their corresponding sensitivity were 93.3%, 75%, 81.7%, 68.3%, 63.3%, and the specificity were 68.7%, 81%, 70.2%, 75.6%, and 81.7%, respectively. Comparison of vascular indices combined with the Breast imaging reporting and data System (BI-RADS) score and simple BI-RADS method, the AUC of power + BI-RADS, ratio + BI-RADS, VFI + BI-RADS, and BI-RADS alone are 0.928, 0.903, 0.895, and 0.796, respectively, which were higher than 50% of the curve. Sensitivity was 81.7%, 80%, 88.3%, 86.7%, and specificity was 88.5%, 85.5%, 77.1%, 69.5%, respectively. The power + BI-RADS method has the highest AUC among these three methods. CONCLUSIONS: Quantitative measurement of blood flow and blood vessel distribution in breast tumors by three-dimensional power Doppler ultrasound combined with the VOCAL method is more accurate and sensitive than the traditional two-dimensional ultrasound. And this method has potential promising applications in many current active research areas, such as the studies of random distribution of intratumoral blood vessels or the normalization of tumor blood vessels. Three-dimensional power Doppler ultrasound combined with the VOCAL method provides a new approach to achieving accurate judgments and the method evaluates the curative effect in breast cancer patients.

Emerging Gram-positive bacteria and drug resistance in cirrhosis patients with spontaneous bacterial peritonitis: A retrospective study.

Spontaneous bacterial peritonitis (SBP) is one of the most severe complications in liver cirrhosis (LC) patients with ascites. The aim of the present study was to retrospectively analyze the bacterial spectrum and drug resistance in ascites culture of patients with SBP. A total of 3, 189 patients with ascites were enrolled in the present study, including 912 LC patients, of which 247 had SBP. It was revealed that in the 3, 189 patients, the ratio of SBP exhibited annual increases, especially in 2015, and this trend remained when cases were divided into two groups: Group A (admission, 2011-2013) and Group B (admission, 2014-2016). The 247 SBP patients were then stratified into two groups: Group 1 (admission, 2011-2013) and Group 2 (admission, 2014-2016). The rate of infection with gram-positive bacteria (GPB) was markedly higher in Group 2 compared with Group 1. Over time, GPB and gram-negative bacteria (GNB) were increased, while the increase of GPB was greater than that of GNB. Direct bilirubin and C-reactive protein levels, and the positive rate of ascites culture in Group 2 were greater than in Group 1. Furthermore, marked differences in serological and ascitic indexes or pathogeny, as well as complications between the patients with GPB and GNB infection were observed. The results regarding drug sensitivity revealed that the resistance rate of GPB and GNB to penicillin (ampicillin) was 100%, while the resistance rate to amikacin, imipenem, meropenem and piperacillin/tazobactam was 0% for GNB, and similarly, the resistance rate to vancomycin, teicoplanin, amikacin and linezolid was 0% for GPB. The results suggested that combined use of ampicillin/sulbactam or piperacillin/tazobactam should be selected forempirical therapy. In cases of nosocomial infection, these drugs should be combined with vancomycin, linezolid or teicoplanin when required.

Jinlong capsule inhibits migration and invasion in human glioblastoma cells via the modulation of mTOR/S6 signaling pathway.

Aim: To investigate the anticancer effects of Jinlong capsule (JLC) against human glioblastoma cells and the possible underlying mechanism. Methods: Cell Counting Kit-8 and colony formation assay were adopted for the analysis of cell viability. Cell invasion and migration were evaluated by transwell and wound healing assays. Then, the expression level of mammalian target of rapamycin (mTOR), phosphorylated mTOR (p-mTOR), S6 and phosphorylated S6 (p-S6) were determined by western blotting. Results: The results showed that JLC significantly inhibited human glioblastoma cell proliferation, invasion and migration in a dose-dependent manner. The expressions of p-mTOR and p-S6 were dramatically suppressed by JLC. Furtherly, inhibition of mTOR reduced the cell migration and invasion, while the mTOR agonist (MHY1485) could partially reverse the anti-migration and anti-invasion activity of JLC. Conclusion: The above results suggested that JLC would be a potential candidate for the treatment of glioblastoma.

Diagnostic determination of Norovirus infection as one of the major causes of infectious diarrhea in HIV patients using a multiplex polymerase chain reaction assay.

Although infectious diarrhea is one of the most common complications in human immunodeficiency virus (HIV)-infected patients, robust diagnostic methods for determining potential pathogens are still limited in the clinic. Norovirus, a type of calicivirus, has been shown to be the most common cause of gastroenteritis. Here, we used multiplex polymerase chain reaction as a diagnostic tool to verify Norovirus as the diarrhea-related pathogen in HIV-infected patients with unknown etiological information. Stool specimens obtained from 81 HIV-infected patients with diarrhea were analyzed by BioFire FilmArray Gastrointestinal (GI) panel. Among 26 HIV-infected patients with unknown etiological information, we detected Norovirus in 14 stool specimens of these patients with 100% sensitivity and specificity as confirmed by reverse transcription polymerase chain reaction (RT-PCR), and one specimen showed both Norovirus and enterotoxigenic Escherichia coli infection. Among the remaining 55 patients with verified Clostridium difficile infection, nine patients also detected positive for Norovirus infection. In conclusion, using FilmArray GI panel and RT-PCR, we determined that Norovirus infection as one of the main pathogens responsible for diarrhea in HIV patients.

Optimization of the microwave-assisted enzymatic extraction of Rosa roxburghii Tratt. polysaccharides using response surface methodology and its antioxidant and α-d-glucosidase inhibitory activity.

An extraction assay applying microwave-assisted enzymatic treatment for polysaccharides in Rosa roxburghii was developed using response surface methodology. The process parameters were optimized using Plackett-Burman (PB) design and central composite design to enhance the Rosa roxburghii polysaccharide extraction yield. Specific conditions (microwave power, 575W; microwave time, 18min; liquid-to-material ratio, 13.5:1mL/g; and enzyme dose, 6.5g/mL) generated an experimental yield of 36.21±0.62%, which closely agreed with the predicted value of 35.75%. Purification with a DEAE-52 cellulose column generated two fractions, PR-1 (from 6.2×103 to 7.4KDa) and PR-2 (from 559.8 to 106.6KDa). Subsequently, the antioxidant activity and α-d-glucosidase inhibitory activity of the two polysaccharide fractions were assessed; PR-1 exhibited stronger antioxidant activity and α-d-glucosidase inhibitory activity than PR-2. Finally, the monosaccharide composition of PR-1 was determined by HPLC using a 1-phenyl-3-methyl-5-pyrazolone precolumn derivatization method. The result showed that PR-1 contained mannose, ribose, rhamnose, glucosamine hydrochloride, glucuronic acid, galacturonic acid, glucose, galactose, arabinose and fucose with molar percentages of 2.1%, 0.54%, 2.1%, 0.26%, 1.5%, 22.7%, 24.0%, 26.4%, 19.6% and 0.89%, respectively.

The emergence of multi-resistant Enterococcus faecalis clonal complex, CC4, causing nosocomial infections.

PURPOSE: Enterococcus faecalis is commonly found as a commensal gut bacteria, but some linages have caused increasing extra-gastrointestinal infections. In particular, strains with high-level virulence or antimicrobial resistance are prevalent in healthcare settings as nosocomial pathogens. This study was performed to elucidate the epidemiological characteristics and antimicrobial susceptibility profiles of E. faecalis causing nosocomial infections in a Chinese general hospital over a 4-year period. METHODOLOGY: We collected 77 isolates causing extra-gastrointestinal infections from patients at 14 different wards in a tertiary hospital from 2011 to 2014. The population relationship was assessed by multilocus sequence typing and multilocus variable-number tandem repeat analysis. The Kirby-Bauer disk diffusion method was used to evaluate susceptibility against 11 antimicrobial agents. RESULTS: The isolates showed high-level resistance to tetracycline (86.5 %), erythromycin (78.4 %), rifampin (62.2 %), etc. The major clonal complexes (CCs) included CC4, CC16 and CC21. As the most dominant subtype, CC16 was identified in almost all of the wards and all types of samples, but the isolation rate decreased continually. In contrast, the isolation rates of CC4 and CC21 increased and the proportion of these two CCs in 2014 was more than three times that in 2011. In addition, CC4 showed higher resistance than CC16. CONCLUSIONS: This study demonstrated the prevalent subtypes and resistance profiles of E. faecalis causing nosocomial infection, and indicated that CC4 may be a newly emerging high-risk, multi-resistant cluster. More surveillance is urgently needed, which will increase our understanding of the prevention and treatment of such infections.

Increased oxidative damage of RNA in liver injury caused by hepatitis B virus (HBV) infection.

To evaluate the urinary levels of 8-oxo-7,8-dihydro-2'deoxyguanosine (8-oxo-dGsn) and 8-oxo-7,8-dihydroguanosine (8-oxo-Gsn) in liver injury patients with hepatitis B virus (HBV) infection and to explore the relationship between urinary 8-oxo-dGsn or 8-oxo-Gsn and degree of liver damage. We enrolled 138 liver injury patients with HBV infection and 169 age- and sex-matched healthy controls in this study. A sensitive and accurate isotope-diluted liquid chromatograph mass spectrometer/mass spectrometer (LC-MS/MS) method was used to measure the urinary levels of 8-oxo-Gsn and 8-oxo-dGsn. Simultaneously, pathological analysis of liver biopsy tissues was carried out, and immunohistochemistry was carried out for 8-oxo-Guo, 8-oxo-dGuo and MTH1 protein in some liver injury tissues. We analysed the correlation between the degrees of inflammation and fibrosis and levels of 8-oxo-Gsn and 8-oxo-dGsn. We also analysed the levels of urinary 8-oxo-Gsn and 8-oxo-dGsn with clinical data of HBeAg, HBsAg, and HBV genotype and detected the levels of plasma aspartate aminotransferase, alanine aminotransferase (AST), platelet, alkaline phosphatase, prothrombin time (PT) and HBV DNA, and calculated the aspartate amino transferase-to-platelet ratio index (APRI) score. Nonparametric correlations were used to evaluate the correlation between 8-oxo-Gsn, 8-oxo-dGsn or APRI and various laboratory biochemical indicators. Results showed that the levels of urinary 8-oxo-Gsn and 8-oxo-dGsn in patients with liver injury were significantly higher than those of healthy controls (both p < .001). Urinary 8-oxo-Gsn was significantly associated with AST, APRI and PT (p = .013, p = .026 and p = .049). The receiver operating characteristic curves of 8-oxo-Gsn were 0.696 (0.632-0.759) and 0.731 (0.672-0.790) for inflammatory activity and fibrosis, respectively. Patients with higher levels of urinary 8-oxo-Gsn are more likely to have a high degree of fibrosis and urinary 8-oxo-Gsn may have a great potential in assessing liver fibrosis.

MicroRNA signatures from multidrug­resistant Mycobacterium tuberculosis.

Tuberculosis (TB) infections, caused by multidrug­resistant Mycobacterium tuberculosis (MDR MTB), remain a significant public health concern worldwide. The regulatory mechanisms underlying the emergence of MDR MTB strains remain to be fully elucidated, and further investigation is required in order to develop better strategies for TB control. The present study investigated the expression profile of microRNA (miRNA) in MTB strains, and examined the differences between sensitive MTB and MDR MTB using next generation sequencing (NGS) with Illumina Deep Sequencing technology to better understand the mechanisms of resistance in MDR MTB, A total of 5, 785 and 195, and 6, 290 and 595 qualified Illumina reads were obtained from two MDR MTB strains, and 6, 673 and 665, and 7, 210 and 217 qualified Illumina reads were obtained from two sensitive MTB strains. The overall de novo assembly of miRNA sequence data generated 62 and 62, and 95 and 112 miRNAs between the 18 and 30 bp long from sensitive MTB strains and MDR MTB strains, respectively. Comparative miRNA analysis revealed that 142 miRNAs were differentially expressed in the MDR MTB strain, compared with the sensitive MTB strain, of which 48 were upregulated and 94 were downregulated. There were six similarly expressed miRNAs between the MDR and sensitive MTB strains, and 108 miRNAs were expressed only in the MDR MTB strain. The present study acquired miRNA data from sensitive MTB and MDR MTB strains using NGS techniques, and this identification miRNAs may serve as an invaluable resource for revealing the molecular basis of the regulation of expression associated with the mechanism of drug­resistance in MTB.