Shengjiang Xiexin Decoction ameliorates antibiotic-associated diarrhea by altering the gut microbiota and intestinal metabolic homeostasis.

BACKGROUND: Antibiotic-associated diarrhea (AAD) has had a significant increase in the last years, with limited available effective therapies. Shengjiang Xiexin Decoction (SXD), a classic traditional Chinese medicine formula for treating diarrhea, is a promising alternative for reducing the incidence of AAD. PURPOSE: This study aimed to explore the therapeutic effect of SXD on AAD and to investigate its potential therapeutic mechanism by integrated analysis of the gut microbiome and intestinal metabolic profile. METHODS: 16S rRNA sequencing analysis of the gut microbiota and untargeted-metabolomics analysis of feces were performed. The mechanism was further explored by fecal microbiota transplantation (FMT). RESULTS: SXD could effectively ameliorate AAD symptoms and restore intestinal barrier function. In addition, SXD could significantly improve the diversity of the gut microbiota and accelerate the recovery of the gut microbiota. At the genus level, SXD significantly increased the relative abundance of Bacteroides spp (p < 0.01) and decreased the relative abundance of Escherichia_Shigela spp (p < 0.001). Untargeted metabolomics showed that SXD significantly improved gut microbiota and host metabolic function, particularly bile acid metabolism and amino acid metabolism. CONCLUSION: This study demonstrated that SXD could extensively modulate the gut microbiota and intestinal metabolic homeostasis to treat AAD.

Quinolone Resistance Genes and Their Contribution to Resistance in Vibrio cholerae Serogroup O139.

BACKGROUND: Quinolones are commonly used for reducing the duration of diarrhea, infection severity, and limiting further transmission of disease related to Vibrio cholerae, but V. cholerae susceptibility to quinolone decreases over time. In addition to mutations in the quinolone-resistance determining regions (QRDRs), the presence of qnr and other acquired genes also contributes to quinolone resistance. RESULTS: We determined the prevalence of quinolone resistance related genes among V. cholerae O139 strains isolated in China. We determined that eight strains carried qnrVC, which encodes a pentapeptide repeat protein of the Qnr subfamily, the members of which protect topoisomerases from quinolone action. Four qnrVC alleles were detected: qnrVC1, qnrVC5, qnrVC12, and qnrVC9. However, the strains carrying qnrVC1, qnrVC5, and qnrVC12 were ciprofloxacin (CIP)-sensitive. Contrastingly, the strain carrying qnrVC9 demonstrated high CIP resistance. qnrVC9 was carried by a small plasmid, which was conjugative and contributed to the high CIP resistance to the receptor V. cholerae strain. The same plasmid was also detected in V. vulnificus. The qnrVC1, qnrVC5, and qnrVC12 were cloned into expression plasmids and conferred CIP resistance on the host V. cholerae O139 strain. CONCLUSIONS: Our results revealed the contribution of quinolone resistance mediated by the qnrVC9 carried on the small plasmid and its active horizontal transfer among Vibrio species. The results also suggested the different effects of qnrVC alleles in different V. cholerae strains, which is possibly due to differences in sequences of qnrVC alleles and even the genetic characteristics of the host strains.

Characterization of the Clinical Features in HBV-Related Acute-on-Chronic Liver Failure.

OBJECTIVE: Acute-on-chronic liver failure (ACLF) is a type of liver failure commonly found in China, and currently the mechanism of the disease remains unknown. This study aimed to investigate the epidemiology, clinical features and prognostic factors in ACLF. METHODS: This study retrospectively included 170 patients with ACLF admitted to Beijing Friendship Hospital in Beijing, China from November 2017 to May 2019. Patients were divided into 2 groups: the improved group and the deteriorated group, according to the severity of their disease. Patients' demographic data; clinical manifestations; complications; laboratory indicators including platelets (PLT), alanine aminotransferase (ALT), aspartate amino transferase (AST), total bilirubin (TBIL), prothrombin time (PT), activated partial thromboplastin time (APTT), prothrombin activity (PTA), international normalized ratio (INR), and alkaline phosphatase (ALP) were collected. The relationship between these factors and the patients' prognosis were analyzed by logistic multivariate regression analysis. RESULTS: The highest morbidity rate was in the age group 40 to 49 years (29.41%). The age group with the second highest morbidity was between 50 and 59 years (25.29%), followed by >60 (21.18%), 30 to 39 (20.59%), 20 to 29 (2.94%) and <20 years (0.59%). A total of 53 patients (31.18%) had a family history of hepatitis B virus infection. The patients' main clinical manifestations were ascites (77.65%) and weakness (68.23%). The most common complications were hypoalbuminemia (80%), infection (67.65%) and electrolyte imbalance (44.12%). In addition, the PTA (P = .009), hepatorenal syndrome (P = .005) and hepatic encephalopathy (level IV) (P = .005) were independently related to the prognosis of ACLF. There is a significant relationship between complications and prognosis (χ2 = 8.502; P = .004). CONCLUSION: This study showed that prothrombin activity, hepatorenal syndrome and hepatic encephalopathy were independently related to the prognosis of ACLF. This outcome provided more options for reducing patient mortality in clinic.

Changes of soil bacterial community structure at the secondary successional stages in the Pinus yunnanensis forest.

Soil bacterial diversity is a key factor for the maintanence of forest ecosystem function. Soil bacterial community would change along forest succession. We analyzed the variations of soil bacterial diversity and community composition at different successional stages in the Pinus yunnanensis forest, which would help understand the mechanism underlying forest restoration. We investigated soil bacterial diversity, community composition, and effect factors at different successional stages (including coniferous forest, mixed coniferous and broadleaf forest, and evergreen broadleaf forest) using Illumina Hiseq platform. The results showed that OTUs, Chao1 index, Ace index, and Shannon index of soil bacterial community decreased with the process of secondary succession. The highest soil bacterial total OTUs, richness, and complexity appeared at early successional stage. Soil bacterial community composition varied across different stages, with the mixed coniferous and broadleaf forest showing largest variation. Proteobacteria and Acidobacteria were common dominant phyla at secondary successional stages. Actinobacteria, Chloroflexi, and Patescibacteria were dominant phyla at the early successional stage, the abundance of which decreased with successional process in the P. yunnanensis forest. Proteobacteria and WPS-2 increased with the succession. Soil pH and tree species richness were key factors in driving soil bacterial community structure. The soil bacterial diversity decreased with forest succession, while the variations of soil bacterial community composition became larger.

Modulation of gut microbiota and intestinal barrier function during alleviation of antibiotic-associated diarrhea with Rhizoma Zingiber officinale (Ginger) extract.

Antibiotic-associated diarrhea (AAD) is typically mediated by antibiotic therapy, which has increased in prevalence in recent years. Previous studies have suggested that ginger, a common spice and herbal medicine, can modulate the composition of gut microbiota and is beneficial against gastrointestinal disease. This study investigates the therapeutic effects of fresh ginger extract on AAD in a rat model. Gut microbiota and intestinal barrier function were also studied. Ginger was administered to rats with AAD. Diarrhea symptoms were assessed, and 16s rRNA sequencing analysis of gut microbiota was performed. An AAD model was successfully established, and ginger was found to effectively ameliorate AAD-related diarrhea symptoms. After the intervention of ginger decoction, the diversity (rather than richness) of gut microbiota was significantly improved, and the gut microbiota recovery was accelerated. At the genus level, Escherichia_Shigella and Bacteroides levels decreased and increased the most, respectively. Additionally, these changes were demonstrated to be coincidental with the moderate restoration of intestinal barrier function, especially the restoration of tight junction protein ZO-1. Our data indicate that ginger could restore gut microbiota and intestinal barrier function during alleviation of AAD.

The tcdA-negative and tcdB-positive Clostridium difficile ST81 clone exhibits a high level of resistance to fluoroquinolones: a multi-centre study in Beijing, China.

Clostridium difficile infection (CDI) is the leading cause of antibiotic-associated diarrhoea worldwide. In order to gain a better understanding about the molecular epidemiology of C. difficile in Beijing, China, molecular typing, antimicrobial susceptibility testing and drug resistance gene sequencing were performed on 174 strains of C. difficile collected from four large tertiary hospitals in Beijing. In total, 31 sequence types (STs) were identified among the 174 strains. ST81 was found to be the most prevalent (26.4%, 46/174), followed by ST2 (16.7%, 29/174) and ST54 (9.8%, 17/174). All isolates were susceptible to metronidazole and vancomycin. The test strains displayed resistance rates of 97.1%, 44.3% and 44.3% for ciprofloxacin, levofloxacin and moxifloxacin, respectively. ST81 isolates displayed a drug resistance rate of 97.8% for levofloxacin and moxifloxacin, which was significantly higher than ST2 (0%), ST54 (17.6%) and ST42 (0%) isolates (P<0.05). An amino acid mutation (T82I) was identified in GyrA, and the total mutation rate of the C. difficile strains was 40.8% (71/174). The mutation rate of ST81 isolates was 95.7% (44/46). Three amino acid mutations (D426N, S366A and D426V) were identified in GyrB, and the total mutation rate of GyrB was 39.1%. A double-site mutation in GyrB (S366A+D426V) was identified in all ST81 (n=46) isolates. In conclusion, the C. difficile ST81 clone showed a high level of resistance to fluoroquinolones in Beijing, highlighting the need for nationwide surveillance of CDI.

In vitro Activity of a New Fourth-Generation Cephalosporin, Cefoselis, Against Clinically Important Bacterial Pathogens in China.

The objective of this study was to systematically evaluate the in vitro activity of cefoselis and other comparators against common bacterial pathogens collected from 18 hospitals across China. Minimum inhibitory concentrations (MICs) were determined by the broth microdilution method following Clinical and Laboratory Standards Institute (CLSI) guidelines. Cefoselis showed poor activity against extended-spectrum ß-lactamase (ESBL)-producing Escherichia coli, Klebsiella pneumoniae, and Proteus mirabilis, with susceptibility rates of < 10% each, while the susceptibility rates of this antibiotic against non-ESBL-producing strains of these organisms were 100%, 94.3%, and 97.0%, respectively. Cefoselis exhibited susceptibility rates of 56.7-83.3% against other tested Enterobacteriaceae isolates. For Acinetobacter baumannii and Pseudomonas aeruginosa isolates, the susceptibility rates to cefoselis were 18.7% and 73.3%, respectively. All methicillin-resistant Staphylococcus aureus (MRSA) strains were resistant to cefoselis, while all methicillin-sensitive S. aureus (MSSA) strains were susceptible to this antibiotic. In conclusion, cefoselis showed good activity against non-ESBL-producing E. coli, K. pneumoniae, and P. mirabilis, MSSA, and was also potent against Enterobacteriaceae, P. aeruginosa, and Streptococcus.

[Non-structural carbohydrates characteristics of different forest layers in monsoon broad-leaved evergreen forest in Pu'er, Yunnan Province, China]. / 云南普洱季风常绿阔叶林不同林层非结构性碳水化合物特征.

Based on cluster analysis of species vertical stratification, the variations in concentration, allocation and seasonal dynamics of non-structural carbohydrates were investigated in a monsoon broad-leaved evergreen forest in Pu'er, Yunnan Province. The results showed that the concentrations of soluble sugar (3.9%) and nonstructural carbohydrates (NSC, 13.3%) were the highest in the sub-canopy. In contrast, the sugar/starch (0.76) of understory was the lowest. There was no significant difference in starch content among three forest layers. For all forest stories, the soluble sugar was mainly allocated to leaves and the roots had more starch and NSC. The soluble sugar contents in leaves and trunks of sub-canopy species were higher than canopy and understory species. There were no significant differences in soluble sugar content of twigs and roots among three different forest layers. The starch content of leaves increased with decreasing height of forest layers. In contrast, the lowest starch content (10.7%) of roots was in understory. There was no significant difference in starch content of twigs and trunks among three different forest layers. The NSC content of leaves was lower in canopy (10.7%) than that in sub-canopy (12.3%) and understory (12.0%). The lowest NSC content of roots was in understory (14.2%). The lowest sugar/starch of leaves, twigs, and trunks presented in understory, while the lowest sugar/starch of roots presented in canopy (0.79). There were dramatic seasonal variations in concentrations and components of NSC. The concentration of soluble sugar and the sugar/starch were higher in the rainy season than that in the dry season. However, the concentration of starch and NSC were higher in the dry season than in the rainy season. The variations in concentration of NSC and its components among forest layers indicated that species with different heights varied in their utilization strategies of carbon, which partly explained species coexistence.

Inhibitory Effect of Vaginal Lactobacillus Supernatants on Cervical Cancer Cells.

Lactobacilli have been shown to inhibit the proliferation of several types of cancer cells, but the effects of vaginal Lactobacilli on cervical cancer cells have seldom been reported. We incubated Caski cells with supernatants of predominant strains in the vagina and investigated their effects on cell growth and the possible mechanisms. Cell-free supernatants of Lactobacillus crispatus, L. jensenii, and L. gasseri were prepared and purified. Caski cells were treated with various concentrations of Lactobacillus supernatants (LS). The effect of LS on cell growth was investigated using MTT assays. The influence of LS on the cell cycle and expression of human papillomavirus (HPV) E6 and E7 oncogenes was determined by flow cytometry and RT-PCR, respectively. LS-inhibited Caski cell proliferation caused morphological changes in a pH-independent manner. Flow cytometric analysis revealed that cells exposed to LS exhibited a significant increase of cell number in S phase and a strong decrease of cell number in G2/M phase. Expression of HPV E6 and E7 oncogenes, as well as CDK2 and cyclin A was decreased after treatment with LS, while expression of p21 was increased. Supernatants of L. crispatus, L. jensenii, and L. gasseri have inhibitory effects on the viability of cervical cancer cells via regulation of HPV oncogenes and cell cycle-related genes. Lactobacillus, as a promising treatment for cancer, is being assessed for its effect, and these results provide further evidence in this respect.

Epidemiology and antifungal susceptibilities of yeast isolates causing invasive infections across urban Beijing, China.

AIM: To investigate the species distribution and antifungal susceptibility profiles of yeast isolates causing invasive infections across Beijing. MATERIALS & METHODS: A total of 1201 yeast isolates recovered from blood and other sterile body fluids were correctly identified by matrix-assisted laser desorption/ionization TOF MS supplemented by DNA sequencing. Antifungal susceptibility testing was performed according to the Clinical and Laboratory Standards Institute broth microdilution method. RESULTS: Candida (95.5%) remained the most common yeast species isolated; Candida albicans (38.8%) and Candida parapsilosis (22.6%) were the leading species of candidemia. Azole resistances were mainly observed in Candida glabrata and Candida tropicalis isolates. CONCLUSION: This study outlined the epidemiologic data of invasive yeast infections and highlighted the need for continuous monitoring of azole resistances among C. glabrata and C. tropicalis isolates in Beijing.

Crystal structure of µ-4-oxidobenzoato-κ2O1:O4-bis-[bis-(1,10-phenanthroline-κ2N,N')copper(II)] bis-(4-hy-droxy-benzoate) 7.5-hydrate.

The title hydrated complex, [Cu2(C7H4O3)(C12H8N2)4](C7H5O3)2·7.5H2O, is composed of dinuclear CuII complex cations, noncoordinating 4-hy-droxy-benzoate anions and water mol-ecules of crystallization. In the dinuclear complex cation, the CuII ions are bridged by a 4-oxidobenzoate ligand and thus each metal ion is five-coordinated by two chelating 1,10-phenanthroline (phen) mol-ecules and one anionic O atom in a distorted trigonal-bipyramid geometry. In the crystal, aromatic π-π stacking occurs between phen rings of neighbouring dinuclear CuII complex cations, forming two-dimensional supra-molecular systems parallel to (100).

Preferable procedure for the screening of syphilis in clinical laboratories in China.

BACKGROUND: With the rapid expansion of infectious syphilis all over the world, optimal procedures for screening syphilis are urgently required. Conventional methods for the diagnosis of syphilis are time- and labor-consuming. We compared automated chemiluminescent micro-particle immunoassay (CLIA) with conventional methods to verify whether CLIA is feasible for syphilis screening. METHODS: A cross-sectional assay was conducted on 3962 serum samples tested by CLIA, rapid plasma reagin test (RPR), and Treponema pallidum particle agglutination (TPPA). Meanwhile, another 36 000 sera were screened for syphilis using CLIA and the positive samples were confirmed using TPPA, RPR or Western blotting. RESULTS: The sensitivity and specificity were 100% and 99.8% for CLIA, and 65% and 99.6% for RPR. With the elevation of the optical density value of samples to cut-off ratio (S/CO) value, the true-positive rate of CLIA increased significantly, and when the S/CO value exceeded 10, the true-positive rate of CLIA reached 100%. The false-positive rate of CLIA was 0.22%; pregnant women had the most false-positive results, followed by elderly people and cancer patients. CONCLUSION: CLIA is suggested as a screening test for the diagnosis of syphilis, while TPPA and RPR are required for confirming the positive samples and monitoring their activity.

Quantification of Atopobium vaginae loads may be a new method for the diagnosis of bacterial vaginosis.

BACKGROUND: Traditional methods for the diagnosis of BV is either with poor sensitivity or poor specificity. Thus, establishing a new method based on the vaginal flora is vital for the diagnosis of BV. METHODS: This article is a retrospective research. Based on the Amsel criteria and Nugent score, 230 BV-positive patients and 360 healthy women were enrolled, specific PCR and quantitative PCR were applied to quantify 5 BV-associated bacteria, including Gardnerella vaginalis, Atopobium vaginae, Leptotrichia/Sneathia species, Megasphaera species and Mobiluncus mulieris. ROC curve was facilitated to screen a bacterial panel with optimal sensitivity and specificity. RESULTS: Specific PCR showed that the area under ROC curve of A.vag, G.vag + A.vag, G.vag + A.vag + Lepto, G.vag + A.vag + Mega and G.vag + A.vag + M.mul were 0.845, 0.862, 0.865, 0.869 and 0.867, the sensitivity and specificity were higher than 80%, which were practicable methods for the diagnosis of BV. Quantitative PCR showed the area under ROC curve of Gardnerella vaginalis, Atopobium vaginae, Leptotrichia/Sneathia species, Megasphaera species and Mobiluncus mulieris were 0.959, 0.996, 0.933, 0.748 and 0.639, when the cutoff value of Atopobium vaginae loads was 247,800 copies/mL, the optimal sensitivity and specificity were 0.979 and 0.952, which was distinctly better than specific PCR. CONCLUSIONS: Quantification ofAtopobium vaginae loads may be a new method of excellent sensitivity and specificity for the diagnosis of BV.

Antimicrobial activity of linezolid combined with minocycline against vancomycin-resistant Enterococci.

BACKGROUND: Vancomycin-resistant Enterococci (VRE) cause serious infections that are difficult to treat. We carried out this study to determine the mutant prevention concentration (MPC) of linezolid when combined with minocycline against VRE strains, to determine the mechanism of drug resistance in vitro, and to provide a theoretical basis for the rational use of drugs against VRE. METHODS: The minimum inhibitory concentrations (MICs) of linezolid and minocycline against 30 Enterococci (E.) isolates (including 20 VRE strains) were determined by the broth microdilution method. Drug interactions were assessed by the checkerboard microdilution tests and confirmed by time-kill studies. Two vancomycin-susceptible strains N27 and N40 (linezolid MIC, 2 g/ml; minocycline MIC, 4 µg/ml) and control strains E. faecalis ATCC 29212 and ATCC 51299 were also tested. The MPCs of linezolid and minocycline (alone and combined) were determined using the agar dilution method. Strains showing stable resistance were analyzed by polymerase chain reaction (PCR) amplification of domain V of the 23S rRNA gene. RESULTS: Checkerboard titration studies revealed synergistic effects of combination therapy in 26.7% of 30 E. isolates. Antagonism was not observed. The G2576U mutation was detected in stable linezolid-resistant strains of ATCC 29212, N40, and N27 before and after resistance screening, and MIC values increased with the number of G2576U mutations. The MPC of linezolid against E. decreased dramatically when combined with minocycline, and vice versa. CONCLUSION: Linezolid or minocycline alone produce resistant strains; however, their joint use may reduce the MPC of each agent against VRE, thereby decreasing resistant mutants and bacterial infections.

[Species-area relationship at different succession stages of monsoon evergreen broad-leaved forest in south subtropical area of Yunnan Province].

Based on the investigation data of monsoon evergreen broad-leaved forest at its different succession stages (primary, CP; 15 years of succession, CF; and 30 years of succession, CT) in Pu' er of Yunnan Province, this paper studied the species-area relationship of this forest at each succession stage. It was found that in the communities at each succession stage, the number of total species, trees, shrubs, and lianas had a significant correlation with sampling area, with the area explained over 94% of the total variation. The Z value of the total species (0.334) and trees (0.394) was the lowest at CT, whereas that of shrubs (0.437) and lianas (0.326) was the lowest at CF. No significant differences were observed in the intercepts of the species-area curve of total species, trees, shrubs, and lianas among different succession stages, but the coefficient of determination (R2) of the species-area curve of total species and lianas was the highest at CP. The richness of trees and shrubs at CF explained 99.9% of the variance of Z value, but the richness of total species, trees, shrubs, and lianas at CP and CT had no significant correlations with the Z value.

Chemokine CXC Ligand 16 serum concentration but not A181V genotype is associated with atherosclerotic stroke.

BACKGROUND: Serum chemokine CXC Ligand 16 (CXCL16) concentration is associated with atherosclerosis and CXCL16 expression may be influenced by the polymorphism, A181V. We established whether serum CXCL16 concentration or the A181V genotype is more strongly associated with atherosclerotic stroke and its associated risk factor, carotid atherosclerosis. METHODS: PCR-RFLP was used to genotype 244 atherosclerotic stroke patients (AS group), 153 stroke-free controls (patient controls) and 167 healthy controls. Serum CXCL16 concentration was determined for a subset of patients (n=135) and all controls. The same subset of patients was then examined using ultrasound to evaluate their carotid atherosclerotic lesions, including intima-media thickness (IMT), plaque stability and carotid plaque area (CPA). RESULTS: Compared with the patient controls and healthy controls, serum CXCL16 concentration was significantly increased in the AS group (P<0.05, and 0.01). It was also strongly associated with increased IMT, vulnerable plaque and increased CPA (P<0.05, <0.001, and <0.01). However, the CXCL16 A181V genotype distribution and allele frequencies showed no differences between AS and control groups, nor did it influence serum CXCL16 concentration. CONCLUSION: Serum CXCL16 concentration is significantly associated with atherosclerotic stroke and carotid atherosclerosis, suggesting that this biochemical test may be useful to identify patients at increased risk of atherosclerosis.

Monomer-dimer transition of the conserved N-terminal domain of the mammalian peroxisomal matrix protein import receptor, Pex14p.

Pex14p is a central component of the peroxisomal matrix protein import machinery. In the recently determined crystal structure, a characteristic face consisting of conserved residues was found on a side of the conserved N-terminal domain of the protein. The face is highly hydrophobic, and is also the binding site for the WXXXF/Y motif of Pex5p. We report herein the dimerization of the domain in the isolated state. The homo-dimers are in equilibrium with the monomers. The homo-dimers are completely dissociated into monomers by complex formation with the WXXXF/Y motif peptide of Pex5p. A putative dimer model shows the interaction between the conserved face and the PXXP motif of another protomer. The model allows us to discuss the mechanism of the oligomeric transition of the full-length Pex14p modulated by the binding of other peroxins.

Bis(µ-2,4-dihy-droxy-benzoato-κO:O')bis-[aqua-(2,4-dihy-droxy-benzoato-κO)(1,10-phenanthroline-κN,N')cadmium(II)].

In the title centrosymmetric dimeric Cd(II) complex, [Cd(2)(C(7)H(5)O(4))(4)(C(12)H(8)N(2))(2)(H(2)O)(2)], the Cd(II) cation is coord-inated by a bidentate phenanthroline (phen) ligand, three dihy-droxy-benzoate (dhba) anions and one water mol-ecule in a distorted CdN(2)O(4) octa-hedral geometry. Among the dhba anions, two anions bridge two Cd(II) cations to form the dimeric complex with significant different Cd-O bond distances of 2.2215 (19) and 2.406 (2) Å. The centroid-centroid distance of 3.4615 (19) Šbetween two nearly parallel benzene rings of the dhba and phen ligands coordinating to the same Cd(II) cation indicates the existence of intra-molecular π-π stacking in the complex. Extensive O-H⋯O hydrogen bonding and inter-molecular weak C-H⋯O hydrogen bonding help to stabilize the crystal structure. One hy-droxy group of the monodentate dhba ligand is disordered over two sites with a site-occupancy ratio of 0.9:0.1.

Comparison of main lactobacillus species between healthy women and women with bacterial vaginosis.

BACKGROUND: The normal microbial flora of the vagina plays an important role in preventing genital and urinary tract infections in women. Thus an accurate understanding of the composition and ecology of the ecosystem is important to understanding the etiology of these diseases. This study aimed to compare the characteristics of main lactobacillus species between healthy women and women with bacterial vaginosis (BV) by quantitative culture and PCR methods. Main lactobacillus species include L. crispatus, L. gasseri, L. jensenii and L. iners. METHODS: A total of 150 women attending Gynecology Outpatient Clinic of Beijing Friendship Hospital, were diagnosed as having BV because three or more of the following criteria were met (standard of Amsel's composite criteria): homogeneous discharge, elevated vaginal pH (pH > 4.5), production of amines, and presence of "clue" cells. Those with less than three of the criteria were considered as healthy. Simultaneously, smears were made of vaginal fluid and Gram stained, then were assessed qualitatively as normal (grade I), intermediate (grade II), or consistent with BV (grade III). Gardnerella vaginalis were identified by using Vitek 2 Compact and PCR methods. Lactobacillus species were identified by PCR methods. Gardnerella vaginalis and lactobacilli colony counts were determined by calculating the most number of colonies of each species in the appropriate plates (colonies between 10 and 300), corrected by the dilution of the sample in the plates, and multiplied by 10 (to account for plating 100 microl), in order to get colony forming units per milliliter of vaginal secretion. RESULTS: BV was diagnosed in 31% (46/150) patients using the composite criteria, the remainder being regarded as healthy. The majority of patients with BV had a smear assessed as grade III (91%, 42/46) and minority of them had a smear assessed as grade II (9%, 4/46). The majority of healthy women had a smear assessed as grade I (64%, 67/104). Smears assessed as grade II were found (36%, 37/104) among patients diagnosed as healthy, with less than three of the composite criteria. L. crispatus was cultured from 94% of healthy women and 83% of women with BV, with the former colonies count average value of 10(6) and the latter of 10(3). L. gasseri, L. iners, and L. jensenii were cultured from 85%, 68% and 43% of healthy women; and 28%, 89% and 44% of BV women, respectively. CONCLUSIONS: The quantities of four lactobacillus species except L. jensenii had a significant difference between healthy women and women with BV. Our results provide support for the negative association between L. iners and L. gasseri. Although L. crispatus were existent both in healthy and BV positive women's vagina, the numbers of L. crispatus were significantly different for the dominant number in healthy women. Smears of vaginal fluid and Gram stain play an important guiding role in bacteria culture.

Crystal structure of the conserved N-terminal domain of the peroxisomal matrix protein import receptor, Pex14p.

Pex14p is a central component of the peroxisomal protein import machinery, in which the conserved N-terminal domain mediates dynamic interactions with other peroxins including Pex5p, Pex13p, and Pex19p. Here, we report the crystal structure of the conserved N-terminal domain of Pex14p with a three-helix bundle. A hydrophobic surface is composed of the conserved residues, of which two phenylalanine residues (Phe-35 and Phe-52) protrude to the solvent. Consequently, two putative binding pockets suitable for recognizing the helical WXXXF/Y motif of Pex5p are formed on the surface by the two phenylalanine residues accompanying with positively charged residues. The structural feature agrees well with our earlier findings where F35A/L36A and F52A/L53A mutants were impaired in the interactions with other peroxins such as Pex5p and Pex13p. Pex14p variants each with Phe-to-Ala mutation at positions 35, 52, and 35/52, respectively, were defective in restoring the impaired peroxisomal protein import in pex14 Chinese hamster ovary mutant ZP161 cells. Moreover, in GST pull-down assays His(6)-Pex5pL bound only to GST-Pex14p(25-70), not to any of GST-Pex14p(25-70)F35A, GST-Pex14p(25-70)F52A, and GST-Pex14p(25-70)F35A/F52A. Endogenous Pex5p was recruited to FLAG-Pex14p on peroxisomes in vivo but barely to FLAG-Pex14pF35A, FLAG-Pex14pF52A, and FLAG-Pex14pF35A/F52A. Collectively, Phe-35 and Phe-52 are essential for the Pex14p functions, including the interaction between Pex14p and Pex5p.