东莞地区痤疮患者痤疮丙酸杆菌对大环内酯-林可酰胺类耐药基因分析

目的 了解东莞地区痤疮患者痤疮丙酸杆菌对大环内酯-林可酰胺类耐药的基因分布情况,探讨其耐药机制。方法 分离、培养、鉴定痤疮丙酸杆菌后经药敏实验选取大环内酯类-林可酰胺类抗生素耐药菌株53株(单一耐药13株,多重耐药40株)和敏感菌株4株,进行全基因组鸟枪法重测序,与痤疮丙酸杆菌标准株ATCC 6919进行对照,分析本地区痤疮丙酸杆菌中耐药基因情况。结果 敏感菌株均无23S rRNA基因点突变或携带ermX耐药基因;单一耐药(克拉霉素或阿奇霉素)均无携带ermX耐药基因,但3株对克拉霉素低、中度耐药的菌株在23S rRNA基因287、555、287\514位点存在突变;40株多重耐药菌株中,12株23S rRNA基因发生2058点突变(10株由A→G突变,2株由A→T突变),26株携带ermX耐药基因,且携带ermX耐药基因与发生2058点突变的菌株不存在交叉重叠,剩余2株未发现点突变或携带ermX耐药基因。结论 东莞地区痤疮患者痤疮丙酸杆菌对大环内酯-林可酰胺类多重耐药的主要基因型为携带ermX耐药基因(65%)和23S rRNA基因2058点突变(30%)。此外,观察到23S rRN...     

淋球菌对大观霉素耐药基因的初步检测

【摘要】 目的 检测导致大观霉素耐药的淋球菌16S rRNA基因中的突变位点。 方法 对6株大观霉素耐药淋球菌［最小抑菌浓度（MIC） ≥ 128 mg/L］、20株大观霉素敏感菌株（MIC 32 mg/L和16 mg/L各10株）的16S rRNA基因进行DNA扩增和序列测定,分析16S rRNA基因突变情况。 结果 6株大观霉素耐药淋球菌的16S rRNA基因均发生了突变,其中2株（MIC > 256 mg/L）为C1192T突变,1株（MIC 256 mg/L）为C1344T和T1345A突变,1株（MIC 256 mg/L）为T990G和T991C突变,1株（MIC 128 mg/L）为T990G、G1343C和C1344T突变,1株（MIC 128 mg/L）为T991C突变。20株大观霉素敏感菌株均未发生突变。 结论 淋球菌16S rRNA基因不同位点突变可能与大观霉素不同程度的耐药相关,C1192T突变可能导致高度耐药,其他单一位点或多位点突变与不同程度的耐药相关。 【关键词】 奈瑟球菌,淋病; 壮观霉素; 点突变; RNA,核糖体,16S; 抗药性,细菌     

沙眼衣原体对大环内酯类药物敏感性试验及23S rRNA基因突变研究

目的 检测天津地区近年来大环内酯类抗生素对泌尿生殖道沙眼衣原体临床分离株的抗菌活性,筛查耐药株,从基因分子水平上探讨耐药机制。方法 将经McCoy细胞培养法检测出的42株沙眼衣原体临床株,传代培养至感染率达90%以上,收集标本进行3种常用大环内酯类抗生素的药敏试验,用RT-PCR、PCR方法分别扩增与大环内酯类耐药相关的23S核蛋白体RNA基因、核糖体蛋白L4基因,产物直接测序检测基因突变。结果 最小抑菌浓度（MIC）结果为红霉素0.5 ～ 2 mg/L,克拉霉素0.008 ～ 0.032 mg/L,阿奇霉素0.125 ～ 0.5 mg/L。发现2株红霉素耐药株,MIC值为2 mg/L,2株耐药株的23S rRNA基因有C2452A、T2611C的突变（大肠杆菌序列编号）,红霉素耐药株及敏感株L4基因均发现脯氨酸113→亮氨酸、脯氨酸156→丙氨酸两个位点的突变。结论 沙眼衣原体对红霉素已产生一定的耐药性。C2452A、T2611C的突变导致红霉素低水平耐药,L4基因的点突变可能与红霉素耐药无关。     

12种抗菌药物对痤疮丙酸杆菌的体外抗菌活性研究

目的 评价12种抗菌药物对临床分离的痤疮丙酸杆菌（P.acnes）的体外抗菌活性。方法 本院2014年8月至2016年4月收集的100株P.acnes,采用肉汤稀释法测定其对四环素、多西环素、米诺环素、红霉素、罗红霉素、克拉霉素、阿奇霉素、甲氧苄啶、左氧氟沙星、氯霉素、克林霉素和夫西地酸的敏感率和最低抑菌浓度（MIC）。结果 米诺环素对P.acnes的MIC90与敏感率分别为8.0 mg/L、66%,多西环素为32 mg/L、36%,左氧氟沙星为16 mg/L、34%,氯霉素为128 mg/L、17%,克林霉素为 > 128 mg/L、7%;夫西地酸为16 mg/L、6%;四环素为 > 128 mg/L、4%,甲氧苄啶和阿奇霉素为 > 128 mg/L、3%;红霉素、罗红霉素和克拉霉素为> 128 mg/L、0。结论 临床分离的P.acnes对12种抗菌药物的敏感性由高到低为米诺环素、多西环素、左氧氟沙星、氯霉素、克林霉素、夫西地酸、四环素、甲氧苄啶、阿奇霉素,对红霉素、罗红霉素和克拉霉素耐药。     

淋球菌rpsE基因突变与大观霉素耐药性的研究

目的 探讨淋球菌rpsE基因突变与淋球菌大观霉素耐药的相关性。 方法 对临床分离的4株大观霉素耐药的淋球菌株（MIC128 μg/ml、256 μg/ml）的rpsE基因进行PCR扩增测序分析,寻找可能的突变位点,通过DNA转化技术将含有突变基因的细菌基因组DNA转化入敏感的淋球菌株,检测转化成功的淋球菌的MIC并进行PCR扩增测序,分析发现的突变位点与淋球菌大观霉素耐药的相关性。 结果 4株大观霉素耐药的菌株均发现rpsE基因A70C（Thr24Pro）突变,而16S rRNA大观霉素耐药决定区（SRDR）未发现任何突变;大观霉素敏感菌株的16S rRNA和rpsE基因均未发现突变。转化后获得的大观霉素耐药淋球菌中亦发现了同样的rpsE基因突变。 结论 rpsE基因单点突变与淋球菌对大观霉素耐药相关。     

夫西地酸对痤疮丙酸杆菌的体外抗菌活性研究

目的 评价夫西地酸对痤疮丙酸杆菌的体外抗菌活性。 方法 上海华山医院2013年3 - 9月收集的50株痤疮丙酸杆菌,按美国临床实验室标准化研究所推荐的琼脂稀释法测定夫西地酸及其他抗菌药的最低抑菌浓度（MIC）。数据分析采用WHONET 5.4软件。 结果 夫西地酸对痤疮丙酸杆菌的MIC50和MIC90分别为0.5和1.0 mg/L,敏感率为90%;莫西沙星与夫西地酸大致相仿,敏感率为90%;克林霉素和红霉素的敏感率分别为54%和46%,MIC90值均 > 128 mg/L;对甲硝唑耐药。在药物质量浓度为128 mg/L时,克林霉素的累积抑菌率达到70%,红霉素只有48%,而夫西地酸在2 mg/L时的累积抑菌率已达到100%。 结论 夫西地酸对临床分离的痤疮丙酸杆菌具有良好的体外抗菌活性。     

我国痤疮患者皮损中痤疮丙酸杆菌对抗生素耐药性的Meta分析

目的：系统评价我国痤疮患者的皮损分离的痤疮丙酸杆菌(P.acnes)对抗生素的耐药情况，为痤疮抗生素合理治疗提供临床依据。方法：计算机网络检索中国知网(CNKI)、中文科技期刊文数据库(VIP)、万方数据库和PubMed中的相关研究，收集痤疮皮损分离痤疮丙酸杆菌对四环素类、大环内酯类等抗菌药物耐药率的相关研究，对符合纳入标准研究资料进行提取，采用R语言3.3.2 Meta程序包进行Meta分析。结果：共纳入文章13篇，合计1770例痤疮患者，共分离痤疮丙酸杆菌合计1259株。对我国P.acnes的抗生素耐药率进行单组率Meta分析，结果显示P.acnes对红霉素、克拉霉素、阿奇霉素耐药率分别为：45%（95% CI 0.35～0.58)，74%（95% CI 0.51～0.89)，59%（95% CI 0.33～0.84)；对米诺环素、多西环素、四环素耐药率分别为：2%（95% CI 0.00～0.07)，6%（95% CI 0.00～0.26)，8%（95% CI 0.00～0.24)；对甲硝唑耐药率为96%（95% CI 0.80～1.00）。不同抗生素之间存在交叉耐药。结论：我国痤疮患者皮损分离的P.acnes对大环内酯类及甲硝唑耐药率高，对四环素类、喹诺酮类抗生素较为敏感，其中以米诺环素敏感性最高。     

外排泵抑制剂对耐药痤疮丙酸杆菌的体外作用

目的 通过外排泵抑制剂羰基氰氯苯腙(CCCP)和苯丙氨酸-精氨酸-β-萘酰胺(PAβN)联合大环内酯-林可酰胺类抗菌药物进行药敏试验，了解外排泵抑制剂对痤疮丙酸杆菌耐药作用的影响。方法 对前期收集的39株痤疮丙酸杆菌，采用琼脂稀释法进行大环内酯-林可酰胺类抗菌药物体外药敏试验，根据药敏试验结果分组进行外排泵抑制剂联合抗菌药物的外排泵抑制试验，以MIC值降低≥4倍为外排泵表型阳性。结果 39株痤疮丙酸杆菌：多重耐药菌株27株、单一耐药菌株2株、敏感菌株10株。多重耐药组：CCCP联合阿奇霉素筛选出5株外排泵表型阳性菌株；CCCP联合克拉霉素筛选出2株外排泵阳性菌株；CCCP联合红霉素、克林霉素、林可霉素时，菌株外排泵表型均为阴性。PAβN联合克拉霉素筛选出2株外排泵表型阳性菌株；PAβN联合红霉素、阿奇霉素、克林霉素、林可霉素时，菌株外排泵表型均为阴性。单一耐药组：菌株外排泵表型均为阴性。结论 外排泵抑制剂CCCP和PAβN能一定程度上降低多重耐药痤疮丙酸杆对大环内酯-林可酰胺类抗生素的耐药性，痤疮丙酸杆菌耐药与外排泵相关。     

痤疮患者痤疮丙酸杆菌对大环内酯类抗生素的敏感性分析

目的:了解广东省东莞地区痤疮患者痤疮丙酸杆菌对三种大环内酯类抗生素的敏感性情况。方法:采用质谱检测系统鉴定痤疮丙酸杆菌,琼脂稀释法测定痤疮丙酸杆菌对红霉素、阿奇霉素和克拉霉素的最小抑菌浓度(MIC)。结果:104例痤疮患者中分离出70株痤疮丙酸杆菌,对大环内酯类抗生素的耐药率从高到低依次为:阿奇霉素71.43%(50/70)、克拉霉素65.71%(46/70)、红霉素32.86%(23/70)。三种抗生素间存在交叉耐药。结论:东莞地区痤疮患者痤疮丙酸杆菌对大环内酯类抗生素有较高的耐药率并且存在交叉耐药,为该地区痤疮治疗的临床用药提供了借鉴。     

大连地区沙眼衣原体泌尿生殖道感染及耐药性检测分析

目的对大连地区沙眼衣原体泌尿生殖道感染患者进行流行病学调查;比较观察不同药物不同疗程治疗沙眼衣原体泌尿生殖道感染的疗效;通过检测基因23S rRNA的突变情况探讨沙眼衣原体对大环内酯类药物的分子耐药机制。方法对2013年1月-2013年12月本院皮肤性病科门诊诊治的126例沙眼衣原体感染的尿道炎患者进行流行病学调查;统计分析不同方法的治疗结果;同时筛选出6例沙眼衣原体对大环内酯类药物耐药株,进行耐药基因23S rRNA突变的检测。结果男女患者沙眼衣原体阳性率差异无统计学意义(P0.05);阿奇霉素长疗程治疗方案与其他三组治疗方案对沙眼衣原体感染患者的治愈率差异有统计学意义(P0.05);其他三组治疗方案之间的治愈率差异无统计学意义(P0.05);扩增23S rRNA的基因,PCR产物中耐药株测序结果中有1株A2059G突变,有3株A2057G突变,1株A2093T突变,5株有T2611C突变。结论男女沙眼衣原体感染的患病率无差别;阿奇霉素长疗程治疗方案对沙眼衣原体感染的患者治愈率较高;大连地区沙眼衣原体对大环内酯类药物的耐药分子基础是23S rRNA基因的点突变。     

Erythromycin resistant propionibacteria in antibiotic treated acne patients: association with therapeutic failure

Erythromycin resistant (EmR) propionibacteria were isolated from the skin surface of 51% of patients treated with oral erythromycin and 42% of patients treated with topical clindamycin compared with 3% of untreated control subjects (P less than 0.001). Amongst the topical clindamycin-treated patients, there was a higher incidence of EmR propionibacterial carriage in those patients who had previously been treated with oral erythromycin (64%) than in patients with no known previous exposure to erythromycin (20%; 0.01 greater than P greater than 0.001). Patients responding to oral erythromycin treatment carried EmR propionibacteria less frequently (24%) than patients who were not responding or who had relapsed (70%; P less than 0.001). These observations suggest that the use of oral erythromycin and/or topical clindamycin encourages the development of resistant propionibacteria and that the emergence of resistant strains is associated with therapeutic failure in erythromycin-treated patients. In total 63 resistant isolates were obtained from 52 subjects. There were 42 strains of Propionibacterium acnes, 16 strains of Propionibacterium granulosum and five strains of Propionibacterium avidum. The majority of isolates were inducibly or constitutively resistant to macrolide (e.g. erythromycin), lincosamide (e.g. clindamycin) and streptogramin B type antibiotics. Therefore, the isolates are phenotypically indistinguishable from the majority of EmR bacteria in which resistance is due to methylation of 23S ribosomal RNA.     

The bacteriology of acne vulgaris and antimicrobial susceptibility of Propionibacterium acnes and Staphylococcus epidermidis isolated from acne lesions

We examined the species of bacteria aerobically and anaerobically isolated from 30 acne lesions and determined antimicrobial susceptibilities of Propionibacterium acnes (P. acnes) and Staphylococcus epidermidis (S. epidermidis) using nine antimicrobial agents. Among the bacteria isolated, S. epidermidis was most dominant. Both P. acnes and S. epidermidis were isolated from half of the acne lesions. The MIC of seven antimicrobials (ampicillin, erythromycin, roxithromycin, clindamycin, tetracycline, minocycline, nadifloxacin) against P. acnes was under 3.13 micrograms/ml. There were very few resistant strains of P. acnes, but many of S. epidermidis. More than 30% of the S. epidermidis isolates were resistant to erythromycin, roxithromycin, and clindamycin. After long-term systemic antibiotic therapy, the resistant strains of S. epidermidis increased, but P. acnes resistance was still limited. When we use antimicrobial agents for the treatment of acne, it should be noticed that not only P. acnes but also S. epidermidis in the acne lesions may acquire resistance to antimicrobials.     

Phenotypic and genotypic characterization of antibiotic-resistant Propionibacterium acnes isolated from acne patients attending dermatology clinics in Europe, the U.S.A., Japan and Australia

BACKGROUND: Propionibacterium acnes is the target of antimicrobial treatments for acne vulgaris. Acquired resistance to erythromycin, clindamycin and tetracyclines has been reported in strains from diverse geographical loci, but the molecular basis of resistance, via mutations in genes encoding 23S and 16S rRNA, respectively, has so far only been elucidated for isolates from the U.K. OBJECTIVES: To determine whether similar or different resistance mechanisms occur in resistant P. acnes isolates from outside the U.K. METHODS: The phenotypes and genotypes of 73 antibiotic-resistant strains of P. acnes obtained from the skin of acne patients in the U.K., U.S.A., France, Germany, Australia and Japan were compared. Antibiotic susceptibilities were determined by minimum inhibitory concentration (MIC) measurements, and polymerase chain reaction and DNA sequencing were used to identify mutations in genes encoding rRNA. RESULTS: Most erythromycin-resistant isolates (MIC(90) > or = 512 microg mL(-1)) were cross-resistant to clindamycin but at a much lower level (MIC(90) > or = 64 microg mL(-1)). As in the U.K., resistance to erythromycin was associated with point mutations in 23S rRNA in 49 of 58 strains. An A-->G transition at Escherichia coli equivalent base 2058 was present in 24 strains. This gave a unique cross-resistance phenotype against a panel of macrolide, lincosamide and type B streptogramin antibiotics. Two further point mutations (at E. coli equivalent bases 2057 and 2059) were identified (in three and 22 isolates, respectively) and these were also associated with specific cross-resistance patterns originally identified in isolates from the U.K. However, nine of 10 erythromycin resistant-strains from Germany did not exhibit any of the three base mutations identified and, in six cases, cross-resistance patterns were atypical. Consistent with previous U.K. data, 34 of 38 tetracycline-resistant strains carried a base mutation at E. coli 16S rRNA equivalent base 1058. Tetracycline-resistant isolates displayed varying degrees of cross-resistance to doxycycline and minocycline, but isolates from the U.S.A. had higher MICs for minocycline (4--16 microg mL(-1)) than isolates from other countries and, in particular, Australia. All the P. acnes isolates resistant to one or more of the commonly used antiacne antibiotics were sensitive to penicillin, fusidic acid, chloramphenicol and the fluoroquinolone, nadifloxacin. All but one isolate (from the U.K.) were sensitive to trimethoprim. CONCLUSIONS: This study shows that 23S and 16S mutations identified in the U.K. conferring antibiotic resistance in P. acnes are distributed widely. However, resistant strains were isolated in which mutations could not be identified, suggesting that as yet uncharacterized resistance mechanisms have evolved. This is the first report of high-level resistance to minocycline and is of concern as these strains are predicted to be clinically resistant and are unlikely to remain confined to the U.S.A. Epidemiological studies are urgently required to monitor how resistant strains are selected, how they spread and to ascertain whether the prevalence of resistance correlates with antibiotic usage patterns in the different countries.     

Propionibacterium acnes resistance: a worldwide problem

Antibiotic therapy directed against Propionibacterium acnes has been a mainstay of treatment for more than 40 years. Despite years of widespread use of systemic tetracyclines and erythromycin, change in P. acnes sensitivity to antibiotics was not seen until the early 1980s. The first clinically relevant changes in P. acnes antibiotic sensitivity were found in the USA shortly after the introduction of topical formulations of erythromycin and clindamycin. By the late 1980s, P. acnes strains with very high MIC levels for erythromycin and elevated MICs for tetracycline were increasingly found in the UK and the USA. Mutations in the genes encoding the 23S and 16S subunits of ribosomal RNA were first identified in the UK and also seen in a recent survey from clinics in Europe, Japan, Australia and the USA. In addition, strains were found in which these known mutations could not be identified, indicating that as yet unidentified resistance mechanisms have evolved. These findings indicate the need to develop strategies to minimize the use of antibiotics in acne therapy.     

Inhibition of lipase activity in antibiotic-resistant propionibacterium acnes strains

BACKGROUND AND OBJECTIVE: Erythromycin-sensitive and/or clindamycin-sensitive strains of Propionibacterium acnes show a reduced lipase production at levels below the minimal growth-inhibitory concentration (MIC). The objective of this study was to determine whether erythromycin and clindamycin concentrations far below the MIC inhibit lipase production in P. acnes strains resistant to these antibiotics. METHODS: Of 42 P. acnes strains, 10 showed an MIC >256 micro g/ml for erythromycin. Two strains showed MICs of 0.19 and 0.25 micro g/ml, while the MIC of the remaining strains was 256 micro g/ml were also tested for lipase inhibition by clindamycin. While this method fails to differentiate between inhibition of lipase production and inhibition of lipase activity, the absence of inhibition of lipase activity rules out inhibition of lipase production. RESULTS: Inhibition of lipolysis by sub-MIC concentrations was demonstrated only for clindamycin in 3 P. acnes strains. However, lipase inhibition was seen only at the dilution level immediately below the MIC. CONCLUSIONS: Resistant P. acnes strains with high erythromycin and/or clindamycin MICs can be ruled out to show in vitro inhibition of lipase production at antibiotic concentrations far below the MIC.     

Antibiotic susceptibility of Propionibacterium acnes isolated from acne vulgaris in Korea

Propionibacterium acnes plays an important role in the development of acne, and inflammatory lesions are improved by antibiotics. Long-term use of antibiotics may result in development of resistant strains and treatment failure. The aim of the present study was to investigate the isolation rate of P. acnes and to evaluate its antibiotic susceptibility to widely used antibiotics in acne in Korea. Among 46 patients, 31 P. acnes strains were cultured. Isolated P. acnes was measured for minimum inhibitory concentration (MIC) of tetracycline, doxycycline, minocycline, erythromycin and clindamycin using an Epsilometer test. Age, disease duration and previous history of antibiotic therapy for acne were compared in relation to the MIC. The mean MIC of tetracycline, minocyclines, doxycycline, clindamycin and erythromycin were all below the breakpoint of antibiotic resistance. The patients with acne vulgaris with disease duration of more than 2 years documented higher MIC values in doxycycline, erythromycin, and clindamycin than those of less than 2 years. The patients who were previously treated with topical or systemic antibiotics showed higher MIC in doxycycline. Antibiotic resistance of P. acnes is still low in Korea, but at this point, there is an increasing trend of MIC. Caution and acknowledgement of increased risk of antibiotic resistant P. acnes should be advised in acne antibiotic treatment to minimize and avoid the emergence of the resistant strain.     

Effect of zinc gluconate on propionibacterium acnes resistance to erythromycin in patients with inflammatory acne: in vitro and in vivo study

Tetracyclines and macrolide antibiotics have been in use for acne treatment for more than 20 years. Since 1992 increasing resistance to these antibiotics, and especially to erythromycin, is reported with Propionibacterium acnes. Zinc salts have demonstrated their efficacy in inflammatory acne treatment as well as their bacteriostatic activity against Propionibacterium acnes. The objective of our work was firstly to determine whether the clinical anti-inflammatory efficacy of zinc salts was altered in the presence of erythromycin resistant strains in vivo, and secondly to study the in vitro and in vivo effect of zinc on the sensitivity of Propionibacterium acnes strains to erythromycin. Thirty patients with inflammatory acne were treated by zinc gluconate with a daily dose of 30 mg for two months and bacteriologic samples were taken at D0, D30 and D60. In vivo, this study displayed a reduction in the number of inflammatory lesions after a 2-month treatment whether or not Propionibacterium acnes carriage was present. Concurrently, in vitro addition of zinc salts in the culture media of Propionibacterium acnes reduced resistance of Propionibacterium acnes strains to erythromycin. Thus, association of zinc salts via a systemic route and topical erythromycin treatment seems an interesting option in the light of an increasing number of patients carrying erythromycin resistant Propionibacterium acnes strains.     

Effects of subminimal inhibitory concentrations of erythromycin, tetracycline, clindamycin, and minocycline on the neutrophil chemotactic factor production in Propionibacterium acnes biotypes 1-5.

Biotypes 1-5 Propionibacterium acnes (P. acnes) strains were grown in the presence of 1/10 minimal inhibitory concentrations (sub-MIC) of erythromycin (EM), tetracycline (TC), clindamycin (CLDM), or minocycline (MINO) and their culture filtrates were assayed for human neutrophil chemotactic activity using Boyden chamber methods. The addition of sub-MIC of MINO to the medium strongly suppressed the neutrophil chemotactic activity of the culture filtrates of P. acnes strains of all biotypes. In contrast, with sub-MIC of EM, TC, or CLDM, the activity of the culture filtrates of P. acnes strains of biotypes 2 and 3 were suppressed but those of biotypes 1, 2, and 5 were not. These results indicate that sub-MIC of MINO is capable of decreasing the inflammatory capacity of P. acnes strains of all biotypes.