乳杆菌功能改变与外阴阴道假丝酵母菌病

目的研究VVC患者阴道乳杆菌菌种构成与功能特点。方法从VVC患者和健康女性阴道分泌物中分离乳杆菌并鉴定。测定乳杆菌抑制白假丝酵母菌的能力。结果对照组女性阴道分泌物分离出64株乳杆菌,81.3%能抑制白假丝酵母菌增殖,VVC组分离出51株乳杆菌,70.6%能抑制白假丝酵母菌增殖。对照组常见菌种为卷曲乳杆菌、加氏乳杆菌和詹氏乳杆菌,VVC组常见菌种为阴道乳杆菌、发酵乳杆菌和德氏乳杆菌。对照组所有患者均能分离到具有抑制白假丝酵母菌功能的乳杆菌,VVC组仅42%的患者能分离到具有抑制白假丝酵母菌作用的乳杆菌。结论 VVC患者阴道乳杆菌菌种构成与健康女性存在差别。VVC组患者阴道乳杆菌抑制白假丝酵母菌能力较对照组弱。     

阴道乳杆菌对常见阴道假丝酵母菌抑制作用的体外试验研究

目的:观察阴道乳杆菌菌株在体外实验中对假丝酵母菌生长的抑制作用。方法:分离健康妇女阴道中常见的乳杆菌和外阴、阴道假丝酵母菌病(vulvovaginal candidiasis,VVC)患者阴道中的常见假丝酵母菌,采用96孔板混合生长抑制法观察处于生长对数期的乳杆菌对假丝酵母菌生长的抑制作用。结果:23例健康妇女阴道分泌物中分离出3种乳杆菌:嗜酸乳杆菌(La)、干酪乳杆菌(Lca)和卷曲乳杆菌(Lcr),15例VVC初发患者阴道分泌物中分离出3种假丝酵母菌:白色假丝酵母菌(Ca)、热带假丝酵母菌(Ct)和克柔假丝酵母菌(Ck)。各种乳杆菌均表现出对假丝酵母菌生长的抑制能力,La对各种假丝酵母菌的抑制能力均优于Lcr;与Lca相比,La对Ct的抑制更为明显;产H2O2的Lcr与不产H2O2的Lca对假丝酵母菌的抑制能力无显著差异。结论:各种乳杆菌表现出对假丝酵母菌生长的不同抑制能力,La对研究的3种假丝酵母菌的抑制能力均优于Lcr;与Lca相比,La的唯一优势体现在对Ct的抑制上;乳杆菌对假丝酵母菌的抑制可能不仅仅与产生H2O2能力有关。     

健康妇女和外阴阴道假丝酵母菌病患者阴道乳杆菌对阴道假丝酵母菌抑制作用的实验研究

目的:探讨正常健康和外阴阴道假丝酵母菌病(VVC)妇女阴道分泌物中分离的乳杆菌对假丝酵母菌生长的抑制。方法:健康正常(正常组)和VVC初发(VVC组)妇女各30名,通过定量采集阴道分泌物培养乳杆菌,观察阴道乳杆菌数量及菌种分布情况,并将获得的乳杆菌及VVC患者阴道分泌物中假丝酵母菌分离纯化及鉴定后,采用牛津杯法观察乳杆菌对假丝酵母菌生长的抑制作用,同时比较VVC组和正常组乳杆菌产H2O2能力的差异。结果:①乳杆菌检出率及菌种分布组间无统计学差异(P>0.05);②VVC组阴道乳杆菌数量的平均对数值与正常组相比显著减少(P<0.01);③阴道内乳杆菌BV试剂盒检测为阴性者,正常组显著高于VVC组(P<0.01);④正常组阴道分离的乳杆菌与VVC组比较,对白色和热带假丝酵母菌的生长抑制有统计学差异(P<0.05),对克柔假丝酵母菌的抑制无统计学差异(P>0.05);正常组分离的嗜酸乳杆菌与VVC组的比较,对白色和热带假丝酵母菌的生长抑制均有统计学差异(P<0.01,P<0.05),对克柔假丝酵母菌的生长抑制无统计学差异(P>0.05);但是正常组和VVC组分离的詹氏乳杆菌对上述3种假丝酵母菌的生长抑制均无统计学差异(P>0.05)。结论:①正常妇女与VVC患者阴道内乳杆菌检出率、菌种分布相似,均以嗜酸、詹氏乳杆菌为优势菌种;②VVC患者阴道乳杆菌的数量显著减少;③VVC组产生H2O2的能力下降,可能与VVC发生相关;④正常状态下阴道乳杆菌对假丝酵母菌抑制作用优于VVC感染状态,其中以嗜酸乳杆菌的抑菌作用最为明显。     

阴道白假丝酵母菌酶活性影响因素的研究

目的:探讨外环境改变对阴道白假丝酵母菌酵母相和菌丝相的酶活性的影响。方法:收集标本50株菌株,用牛奶培养基和卵黄培养基法检测白假丝酵母菌酵母相与菌丝相分泌型酸性蛋白酶(SAP)和磷脂酶(PL)的酶活性;从中随机挑选20株菌株,分别用不同pH值的培养基检测其酵母相和菌丝相酶活性的改变;在不同乳杆菌浓度的环境下检测阴道白假丝酵母菌酵母相和菌丝相酶活性的变化。结果:酵母相和菌丝相磷脂酶的活性(PZ值)分别为0.546±0.132和0.471±0.119;分泌型酸性蛋白酶的活性(PA值)分别为0.605±0.125和0.568±0.086;菌丝相分泌型酸性蛋白酶和磷脂酶活性均显著高于酵母相(P<0.001)。pH=4时,白假丝酵母菌酶活性最低,随着pH值增加,酶活性升高,至pH=7时酶活性降低。未添加乳杆菌组白假丝酵母菌酶活性最高,逐渐增加乳杆菌浓度,酶活性随之降低。结论:菌丝相酶活性均高于酵母相;pH值可影响白假丝酵母菌酶活性的改变,过酸或过碱的环境均可抑制白假丝酵母菌酶活性;乳杆菌对白假丝酵母菌的酶活性有抑制作用,且对菌丝相的抑制作用较酵母相更强。     

苦参总碱对阴道常见乳杆菌增殖影响的体外研究

目的探讨苦参总碱对6种阴道常见乳杆菌体外增殖的影响。方法 2018年6月至2019年3月于清华大学附属北京清华长庚医院,将从中国育龄期健康女性阴道中分离、纯化的6种不同阴道乳杆菌进行体外培养,通过琼脂稀释法观察不同浓度的苦参凝胶主要有效成分苦参总碱对各种乳杆菌临床分离株体外增殖的影响。结果苦参总碱对鼠李糖乳杆菌的最低抑菌浓度(MIC)为40 mg/m L;对卷曲乳杆菌、詹氏乳杆菌、加氏乳杆菌、罗伊氏乳杆菌的MIC为20 mg/m L;对阴道乳杆菌的MIC则为10 mg/m L。当苦参总碱浓度低于MIC时,对加氏乳杆菌和罗伊氏乳杆菌具有一定促进增殖的作用,而对卷曲乳杆菌和阴道乳杆菌的增殖无影响。结论临床给药剂量下,苦参凝胶不会抑制阴道常见乳杆菌生长,有利于恢复患者阴道微生态平衡。     

223例健康育龄妇女阴道内乳酸杆菌菌群的鉴定

目的检测健康育龄妇女阴道内乳酸杆菌的主要菌群种类及其产生过氧化氢的能力。方法采集223例健康育龄妇女阴道侧壁的分泌物,经过乳酸杆菌选择性培养基（Rogosa）与肉汤培养基（MRS）分离培养后,提取细菌染色体DNA,PCR扩增细菌的16SrRNA基因,测定16SrRNA核苷酸序列,然后与基因库（GenBank）中的数据进行比对,确定细菌的种类。并对乳酸杆菌产生过氧化氢的情况进行分析。结果从223例健康育龄妇女阴道中分离到8种乳酸杆菌种属,较常见的种属为卷曲乳杆菌（53．7％）,詹氏乳杆菌（27．9％）与格氏乳杆菌（10．3％）。其他少见的细菌类型包括阴道乳杆菌（2．2％）,L．coleohominis（1．5％）,约氏乳杆菌（1．5％）,嗜酸乳杆菌（1．5％）及发酵乳杆菌（1．5％）;大多数乳酸杆菌均能产生过氧化氢,其中卷曲乳杆菌和詹氏乳杆菌产生过氧化氢的能力较强。结论卷曲乳杆菌、詹氏乳杆菌和格氏乳杆菌是健康育龄妇女阴道内乳酸杆菌的主要种属。卷曲乳杆菌和詹氏乳杆菌有较强的产生过氧化氢的能力。     

细胞溶解性阴道病的诊治进展

乳杆菌是女性阴道正常菌群中的重要成员,可通过产生乳酸、H2O2和其他抑制物质如嗜酸乳菌素,刺激免疫系统等方式来抑制大肠埃希菌、假丝酵母菌、阴道加德纳菌的过度生长,维持阴道微生态平衡。但乳杆菌的过量生长可造成阴道上皮鳞状细胞的破裂,导致一些类似外阴阴道假丝酵母菌病(VVC)的临床症状,即细胞溶解性阴道病(CV)。通过对CV的诊断、临床鉴别、治疗等方面进行文献回顾,提示本病发病率低于VVC、复发性VVC(RVVC)等,临床中易误诊误治。对于长期阴道分泌物异常、实验室真菌检查反复阴性、经验使用治疗真菌感染类药物无效的患者,应积极考虑本病的可能。     

细胞溶解性阴道病92例临床分析

乳酸杆菌是女性阴道正常菌群中的最重要成员,它通过产生乳酸、H2O2和其他抑制物质如嗜酸乳菌素,刺激免疫系统等来抑制大肠埃希菌、假丝酵母菌、阴道加德纳菌和动弯杆菌的过度生长[1],维持阴道微生态平衡.     

Farnesol对白假丝酵母菌细胞生长及活性的干扰作用

目的:评价Farnesol对白假丝酵母菌生长、细胞活性及形态的干扰作用;检测氟康唑、克霉唑、制霉菌素、特比萘酚、卡泊芬净在不同浓度Farnesol条件下对白假丝酵母菌的抗菌活性。方法:通过吸光浊度测定不同浓度组Farnesol对白假丝酵母菌生长作用的影响;采用MTT法检测Farnesol对白假丝酵母菌细胞活性的干扰作用;光镜及电镜扫描观察不同浓度组Farnesol作用下,白假丝酵母菌细胞形态改变;测定不同种类抗真菌药物在不同浓度组Farnesol条件下对白假丝酵母菌的抗菌活性。结果:Farnesol对白假丝酵母菌的生长及细胞活性具有抑制作用。低浓度Farnesol即可抑制假丝酵母菌菌丝的形成,随着Farnesol浓度的升高,菌株以孢子相存在且生长受到抑制;高浓度组Farnesol并不能完全抑制菌株生长。Farnesol对抗真菌药物的抗菌活性具有干扰作用,低浓度Farnesol环境可以改善氟康唑的"拖尾"现象。结论:Farnesol对白假丝酵母菌生长及细胞活性具有浓度依赖性的抑制作用。Farnesol抑制白假丝酵母菌菌丝形成,可干扰抗真菌药物的活性,低浓度改善氟康唑的"拖尾"现象。     

快速青春发育期女性外阴阴道感染特征分析

目的:探讨9~13岁快速青春发育期女性外阴阴道感染特征。方法:回顾性分析390例2015年7月至2018年6月在四川大学华西第二医院就诊的9~13岁快速青春发育期女性外阴阴道感染状况,并按年龄分为9岁、10岁、11岁、12岁、13岁5个年龄组。采用χ^2检验分析其假丝酵母菌外阴阴道病(VVC)总发生率、细菌性阴道病(BV)发生率、炎症反应率、优势菌异常率,及不同年龄组上述四者的差异。结果:研究对象中BV发生率、VVC总发生率、炎症反应率、优势菌异常率分别为:37.95%、19.24%、16.15%、8.21%,BV发生率显著高于VVC总发生率,差异有统计学意义(P<0.05)。VVC中白假丝酵母菌和非白假丝酵母菌阳性检出率分别为:15.39%、3.85%,两者比较差异有统计学意义(P<0.05)。5组间经两两比较,发现不同年龄组BV发生率和炎症反应率差异无统计学意义(P>0.05);9岁组VVC总发生率(7.56%)显著低于其他年龄段组(分别为20.75%、29.51%、25.42%、24.44%),差异有统计学意义(P<0.05);10岁组优势菌异常率(12.26%)显著高于13岁组(4.44%),差异有统计学意义(P<0.05)。乳杆菌数量1+/油镜随年龄增长呈下降趋势,而乳杆菌3+~4+/油镜随年龄增长呈上升趋势。结论:快速青春发育期女性外阴阴道感染好发BV和VVC,且BV比VVC更易感,而VVC中以白假丝酵母菌感染为主;9岁女性VVC总发生率最低;10岁女性优势菌异常率高于13岁,波动尚无规律。乳杆菌数量随年龄增长呈规律性增加。     

Antimicrobial components of vaginal fluid

OBJECTIVE: We examined the antimicrobial activity and composition of vaginal fluid. STUDY DESIGN: Vaginal fluid from preweighed tampons was assayed for pH, lactic acid, and antimicrobial polypeptides. The fluid was also fractionated by molecular filtration. Antimicrobial activity of whole fluid was determined against representative resident and exogenous microbes, and its fractions were tested against Escherichia coli. RESULTS: Vaginal fluids (5/5 donors) were permissive for Lactobacillus crispatus and vaginalis and Candida albicans, but not for Escherichia coli, Streptococcus group B, and Lactobacillus jensenii in three of five donors. The antimicrobial activity against E coli was predominantly in a <3-kd fraction and correlated with both low pH and high lactic acid content. Compared with a matched pH buffer, lactic acid markedly suppressed the growth of E coli. Concentrated 2- or 5-fold, the protein-rich fraction was active against E coli. CONCLUSION: Vaginal fluid exerts selective antimicrobial activity against nonresident bacterial species. The activity is mediated by lactic acid, low pH, and antimicrobial polypeptides.     

Bacterial flora of the female genital tract: function and immune regulation

The use of non-culture gene amplification techniques has improved our understanding of the composition of the vaginal bacterial ecosystem. In most healthy women in the reproductive period the predominant vaginal bacteria are one or more of the following species of Lactobacillus: L. crispatus, L. iners and L. gasseri. However, in other apparently healthy women lactobacilli may be deficient or absent, being replaced by other lactic-acid-producing bacteria: Atopobium, Megasphaera and/or Leptotrichia species. Infection and/or proliferation of pathogenic bacteria in the vagina is suppressed by lactic acid production, bacteria-generated antimicrobial products, and the local activities of the innate and cell-mediated immune systems. Vaginal epithelial cells produce a range of compounds with antimicrobial activities. These cells also possess membrane-bound Toll-like receptors that recognize pathogen-associated molecular patterns. Recognition leads to pro-inflammatory cytokine production and antigen-specific immunity. Local production of IgG and IgA antibodies can also be initiated in the endocervix and vagina in response to infection.     

Probiotic potential of 3 Lactobacilli strains isolated from breast milk.

Breast milk is an important factor in the initiation, development, and composition of the neonatal gut microbiota. In a previous study, the authors isolated lactic acid bacteria from milk of healthy mothers. Since some of the identified isolates belonged to the genus Lactobacillus, the objective of this work was to evaluate the probiotic potential of 2 Lactobacillus gasseri and 1 Lactobacillus fermentum strains. Different assays, including survival to conditions simulating those existing in the gastrointestinal tract, production of antimicrobial compounds, adherence to intestinal cells, production of biogenic amines, degradation of mucin, enzymatic profile, and pattern of antibiotic resistance, were performed. Globally, the results showed that the probiotic potential of lactobacilli isolated from milk of healthy mothers is, at least, similar to that of the strains commonly used in commercial probiotic products. This fact, together with the presence of prebiotic substances, indicates that breast milk is a natural synbiotic food.     

The in vitro activity of vaginal Lactobacillus with probiotic properties against Candida

Lactobacilli, the predominant vaginal microorganisms in healthy premenopausal women, control other members of the vaginal microflora and thus protect against bacterial vaginosis and urinary tract infections. It has been claimed that some lactobacilli are also protective against Candida vaginitis. Little is known, however, about the mechanisms by which these lactobacilli can control vaginal populations of Candida and prevent vaginitis. To address this question, vaginal Lactobacillus strains with known antagonistic properties against bacteria were tested for their cell surface properties, adhesion to vaginal cell lines in vitro and antagonistic activities against Candida. A small proportion of the lactobacilli tested adhered strongly to cultured vaginal epithelial cells and inhibited growth of Candida albicans but not of C. pseudotropicalis. This anticandidal activity was in some Lactobacillus strains related to hydrogen peroxide (H2O2) production, but catalase treatment did not suppress this activity in other Lactobacillus strains, suggesting alternative mechanism(s). Moreover, tested vaginal Candida strains were resistant to relatively high concentrations of H2O2 that markedly exceeded those produced by even the most active Lactobacillus strains.     

Defense factors of vaginal lactobacilli

OBJECTIVE: To determine the antagonistic relationship between vaginal lactobacilli and endogenous vaginal microflora. STUDY DESIGN: Twenty-two Lactobacillus strains were studied for the production of lactic acid, hydrogen peroxide, and bacteriocin. RESULTS: Under standardized growth conditions, most strains increased their biomass by more than 4 times. Lactobacillus species grew best at a pH > or = 4.5, and growth was retarded at a pH < 4.5. Lactic acid levels were 0.68 to 2.518 mg/mL and were not related to the number of cells or the pH of media. The pH of the media was caused by the secretion of lactic and other organic acids. Approximately 80% of the strains produced H(2)O(2) and were graded as 2+ in one third of the strains and 1+ in others. No statistical correlation was found between H(2)O(2) lactic acid and bacteriocin production. Bacteriocin activity was tested on 4 strains of Gardnerella vaginalis. Approximately 80% of the lactobacilli tested produced bacteriocin that inhibited growth of G vaginalis. Six of the strains did not produce bacteriocin. Thirteen strains produced all 3 defense factors, whereas the others lacked 1 or 2 properties. CONCLUSIONS: Lactobacillus species grow best at a pH > 4.5. The pH of the media is dependent on the cell mass and on all organic acids produced by Lactobacillus species. Although all species produce organic acids, not all produce H(2)O(2) and bacteriocin. Not all strains of G vaginalis can be inhibited by lactobacilli-producing bacteriocin.     

Could bacterial vaginosis be due to the competitive suppression of lactobacilli by aerobic microorganisms?

Bacterial vaginosis (BV) is characterized by lack of lactobacilli and predominance of anaerobic species. Ciprofloxacin is active against a great number of aerobic bacteria. The effect of ciprofloxacin on the vaginal microbial flora was studied both in healthy subjects and in patients with BV in order to test if BV may be due to competitive suppression of lactobacilli by aerobic species. About half of the patients were relieved of their symptoms. No adverse effects on the microbial profile or colonization by lactobacilli were produced by the drug. Lactobacilli known to be able to produce hydrogen peroxide were found to be significantly more common in healthy women than in women with BV. The species most frequently related to vaginal health were Lactobacillus jensenii and Lactobacillus rogosae.     

Novel insight into the vaginal microflora in postmenopausal women under hormone replacement therapy as analyzed by PCR-denaturing gradient gel electrophoresis

OBJECTIVES: A study was conducted to use new molecular technologies to identify the vaginal bacterial species of postmenopausal women under oral estrogen therapy (Premarin-conjugated equine estrogen, CEE). STUDY DESIGN: Nineteen postmenopausal women under CEE treatment were recruited and their vaginal flora were analyzed during 3 months, using polymerase chain reaction in combination with denaturing gradient gel electrophoresis (DGGE) and sequencing of DNA fragments. RESULTS: Sixty-eight percent of the women presented with a 'Normal' Nugent score at the first sampling time (d0). All the subjects had bacterial species detected in their vaginal flora. Lactobacillus species were detected in all samples. Moreover, 53% of the women had lactobacilli exclusively at d0. Two Lactobacillus species were dominant, and were recovered from the majority of samples (L. iners and L. crispatus). Forty-four percent of the samples also contained other bacterial species, which were potential urogenital pathogens. Candida albicans was detected in 26% of the subjects. The vaginal flora of the women under CEE treatment were different from that of postmenopausal women not receiving hormone therapy, but very similar to premenopausal women. CONCLUSIONS: The present study indicates that one benefit from hormone replacement therapy (HRT) is the restoration of a lactobacilli vaginal flora associated with a protective effect against urogenital infections.     

In vitro adherence of Lactobacillus species to vaginal epithelial cells

Because of the reported use of yogurt douches for the treatment of vaginitis, the adherence of Lactobacillus species to normal human vaginal epithelial cells was tested by in vitro methods. L. acidophilus and L. bulgaricus isolated from three commercial yogurts were compared to L. acidophilus, L. fermentum, L. cellobiosus, and L. casei isolated from clinical specimens and from American Type Culture Collection cultures. L. acidophilus isolated from Lactinex tablets was also tested. No significant difference was found among the 19 clinical isolates, the American Type Culture Collection cultures, and the Lactinex isolate, but all showed significantly higher (p less than 0.01) adherence than any Lactobacillus isolated from yogurt. The presence of Gardnerella vaginalis, which adheres strongly to vaginal epithelial cells, was found to interfere with the adherence of lactobacilli from all sources studied.