葡萄球菌属的分离及耐药性研究

目的　对 1 974株临床分离的葡萄球菌的耐药性进行研究 ,以观察葡萄球菌属的耐药现状。方法　药敏试验采用纸片扩散法 (K -B法 )。结果　临床分离到的葡萄球菌中 ,MRSA的分离率为 2 0 .8% (66/ 31 7) ,MRCNS的分离率为 66 .9% (1 1 0 9/ 1 657) ,MRSA主要来源于生殖道分泌物、痰、伤口分泌物 ,分别占 2 7.3 % ,2 2 .7% ,1 8.2 % ;血液标本中MRSA和MRCNS分别占总分离数的 0 .2 5 %和 4 .9% ,而MRSA只占 0 .2 5 %。MRSA和MRCNS对红霉素、庆大霉素、氯霉素、克林霉素、环丙沙星、复方磺胺甲恶唑、四环素的耐药率分别为 86 .2 %、47.6 %、50 .8%、61 .3 %、54 .5 %、83 .3 %、66 .1 %和 84.9%、37.5 %、45 .9%、53 .7%、58.2 %、84.9%、67.6 % ;MSCNS中环丙沙星、复方磺胺甲恶唑、四环素、氯霉素的耐药率分别为 33 .0 %、64 .1 %、58.5 %、2 7.8%均高于MSSA的 1 4 .6 %、42 .8%、41 .5 %、1 7.7% ,但对于MSS环丙沙星、庆大霉素、氯霉素、克林霉素有较高的敏感性。结论　重视对葡萄球菌的耐药性监测 ,合理使用抗生素、特别是合理使用糖肽类抗生素是非常必要的。     

Isolation and characterization of a monoclonal anti-P30 antibody resistant mutant of Toxoplasma gondii

Of the possible iodine-labelled Toxoplasma gondii surface proteins, P30 (apparent Mr 30,000) is the principal one recognized by acute and convalescent anti-toxoplasma sera. This protein which comprises from 3 to 5% of the total parasite protein was used to raise a panel of parasiticidal monoclonal anti-P30 antibodies. One of these monoclonal antibodies was able to select a resistant mutant from a large population of chemically mutagenized wild-type P strain parasites. This mutant retained the wild type sensitivity to other non-P30 parasiticidal monoclonal antibodies as well as polyclonal anti-P30 rabbit sera. Analysis of surface radioiodinated wild type and mutant parasites showed that the mutant had a quantitative reduction in the amount of P30. A comparison of surface biotin labelled wild type and resistant parasites by two dimensional electrophoresis showed that the mutant lacked one and possibly two of several proteins that make up wild type P30. Western blot analysis indicated that the mutant was devoid of antigenically reactive P30. These findings further support the hypothesis that antigenic variants of T. gondii can be induced and may involve the major surface membrane antigens of the parasite.     

三种实验性IgA肾病模型的比较

目的探讨建立一种理想的IgA肾病（IgAN）动物模型方法。方法分别采用葡聚糖G200、大肠杆菌外膜蛋白和金葡菌的细胞膜20肽抗原决定簇诱导小鼠IgA肾病模型。用分子生物学和病理学方法对3组IgAN模型小鼠进行鉴定和比较。结果（1）葡聚糖组尿蛋白增高，伴有血尿；免疫荧光显示部分肾小球大量IgA沉积；光镜下肾小球系膜细胞增多，肝和脾可见弥漫性的粉染物质沉积；电镜下肾小球系膜区少量低电子密度的致密沉积物，肝和脾可见淀粉丝样物质沉积。（2）大肠杆菌外膜蛋白组尿蛋白增高，伴有血尿；免疫荧光显示肾小球有少量IgA沉积；光镜下肾小球系膜细胞轻度增多，间质炎细胞浸润明显；电镜下肾小球系膜区无电子致密沉积物。（3）金葡菌细胞膜20肽抗原决定簇组尿蛋白增高，伴有血尿；免疫荧光显示多数肾小球均可见大量IgA沉积；光镜下肾小球系膜细胞增多，伴系膜基质轻度增生；电镜下肾小球系膜区和基底膜的内皮细胞下可见高电子密度的致密沉积物。结论金葡菌细胞膜20肽抗原决定簇组诱导的IgAN模型从临床表现和病理学变化与人IgAN极其相似，是3种IgAN模型中最理想的IgAN模型。     

Distribution of the exfoliative toxin D gene in clinical Staphylococcus aureus isolates in France

Exfoliative toxin D (ETD) was identified recently as a new exfoliative toxin serotype. Like other exfoliative toxins, ETD induces intra-epidermal cleavage through the granular layer of the epidermis of neonatal mice. The distribution of ETD production was investigated in Staphylococcus aureus isolates from infected and colonised patients in France. The etd gene was found in 55 (10.5%) of 522 isolates tested. Isolates responsible for bullous impetigo and generalised staphylococcal scalded skin syndrome did not harbour etd, but etd was significantly more frequent in isolates causing cutaneous abscesses and furuncles. Most etd- and Panton-Valentine leukocidin-positive strains belonged to the clone of community-acquired methicillin-resistant S. aureus spreading currently throughout France.     

Possible role of IL-2 deficiency for hypogammaglobulinaemia in patients with common variable immunodeficiency.

Common variable immunodeficiency (CVID) patients are unable to produce specific immunoglobulins after antigen contact in vivo. The aim of this study was to investigate whether in some cases of CVID a decreased de novo synthesis of IL-2 might be the cause of immunodeficiency and whether this deficiency can be corrected by IL-2 supplementation in vitro. Mononuclear cells from 17 CVID patients and from 10 healthy controls were cultured with monoclonal anti-CD3 antibody OKT3, pokeweed mitogen (PWM) or tetanus toxoid (TT) to stimulate IL-2 synthesis. In parallel, in vitro IgG and IgM synthesis was stimulated with Staphylococcus aureus Cowan I (SAC), PWM or TT in the presence or absence of IL-2. While lymphocytes of 11 out of 17 patients produced low to normal amounts of IL-2 upon stimulation with anti-CD3, only three patients showed low IL-2 production in response to PWM and five in response to TT. Regarding immunoglobulin synthesis in vitro, five patients completely failed to produce IgM or IgG upon stimulation with PWM, SAC or TT irrespective of the addition of IL-2. By contrast, four patients did not show any defect in vitro and synthesized normal amounts of IgM and IgG with any of the three stimuli. Finally, eight patients could be reconstituted for PWM-, SAC- and TT-induced IgM and/or IgG synthesis in vitro, by adding IL-2 to the culture system. This enhancing effect of IL-2 could be blocked by adding anti-IL-2 receptor antibodies to the cultures. Our findings indicate that a defective IL-2 synthesis after antigen stimulation may be one reason for the impaired immunoglobulin production in some cases of CVID.     

Adherence of Staphylococcus aureus to cultures of human peritoneal mesothelial cells

Strains of Staphylococcus aureus, isolated from the effluentof patients with peritonitis on CAPD (continuous ambulatoryperitoneal dialysis), adhered well to both cultured human mesothelialcells and to fibronectin, but not to laminin or gelatin. Mesothelialcells grown in medium M199 exhibited more surface fibronectincompared to cells grown in MEM-Dval and demonstrated higherlevels of S. aureus adherence. Soluble fibronectin concentrations up to lOµg/ml increasedthe adherence of S. aureusto cultured mesothelial cells. Thedose-response curve was consistent with the binding of fibronectinto a saturable receptor of apparent dissociation constant (K_D)= 1.7xlO^–10 M. This corresponds closely to the K_D (2xlO^–10M) of the staphylococcal fibronectin-binding protein. S. aureus adherence was increased following the preincubationof mesothelial cell monolayers with interleukin-1 and was maximalafter 6 h preincubation. Treating mesothelial cells with interferon-gammafor 48–72 h reduced the adherence of S. aureus.     

长、短链探针以链延伸反应提高基因传感器检测MRSA灵敏度的实验研究

目的：利用链延伸反应的原理延长引物链以提高石英谐振式基因传感器表面的质量负载，从而提高传感器检测MRSA的灵敏度。方法：以不对称PCR的反应产物做为长链探针，人工合成的寡核苷酸片段为短链探针，在基因传感器阵列表面分别固定上金黄色葡萄球菌mecA和femA的长链及短链探针共4种探针片段，与相应的靶序列杂交后，加入Klenow酶，37℃进行链延伸反应1h。结果：长、短链探针均有效地固定了传感器表面。mecA,femA短链探针在链延伸反应前的检测灵敏度为0.5nmol/L，但经链延伸反应后检测灵敏提高到了0.05nmol/L；而长链探针却无法与相应的基因组靶序列发生杂交。结论：短链探针链延伸反应有效地提高了石英谐振式基因传感器的灵敏度，但该法不适用于链探针。     

The effect of 2,4-diamino-6-hydroxy-pyrimidine on postburn Staphylococcus aureus sepsis in rats

GTP-cyclohydrolase I (GTP-CHI) is the first and rate-limiting enzyme for the de novo biosynthesis of biopterin. The present study was to observe the effect of 2,4-diamino-6-hydroxy-pyrimidine (DAHP),an inhibtor of GTP-CHI, on the development of postburn Staphylococcus aureus sepsis. Methods: 56 male Wistar rats were randomly divided into four groups as follows: normal control group (n= 10), scald control group(n= 10),pos tburn sepsis group (n= 20) and DA HP treatment group (n= 16). In the scald control group, rats were subjected to a 20% total body surface area (TBSA) Ⅲ° scald injury, then sacrificed at 24 hrs. In the postburn sepsis group (n=20), rats were inflicted with 20% TBSA Ⅲ° scald followed by Staphylococcus aureus challenge, and they were further divided into 2 and 6 hrs groups. In the DAHP treatment group (n= 16), animals were intraperitoneally injected with a dose of 1g/kg DAHP prior to Staphylococcus aureus challenge, and then further divided into 2, 6 hrs groups. Tissue samples from liver, kidneys, lungs and heart were collected to determine GTP-CHI, inducible nitric oxide synthase (iNOS) and tumor necrosis factor-α (TNF-α) mRNA expression. Meanwhile, biopterin and nitric oxide (NO) levels in these tissues were also measured. Results: After the scald injury followed by Staphylococcus aureus challenge, GTP-CHI mRNA expression and biopterin levels significantly elevated in various tissues such as liver, heart, kidneys and lungs, so did the values of iNOS mRNA expression and NO formation (P＜0.01). Pretreatment with DAHP could significantly reduce GTP-CHI/biopterin induction (P＜0. 05～0. 01), and the up-regulation of iNOS/NO was also suppressed. Furthermore, DAHP administration could also inhibit the gene expression of TNF-α. 2 hrs after septic challenge, TNF-α mRNA expression in liver, kidneys and lungs in DAHP-treated group were 35.7%, 37.3% and 33.0% of those in postburn septic group, respectively. Additionally, in animals without DAHP treatment, the 6-hour mortality was 55.6% (20/36), while it was only 25.0% in DAHP-treated animals (4/16, P=0. 08). Conclusions: Early treatment with DAHP might be a potential strategy to prevent the development of postburn Staphylococcal sepsis, which appears to be associated with down-regulation of biopterin and NO formation by DAHP.     

凝固酶阴性葡萄球菌检测在慢性前列腺炎中的意义

目的　研究慢性前列腺炎与凝固酶阴性葡萄球菌 (CNS)的关系 ,探讨前列腺液中CNS检测的临床意义。　方法　对 4 2 8例慢性前列腺炎患者采用Meares Stamey四段取样法作前列腺液细菌培养和药敏试验。患者年龄 18～ 4 6岁 ,平均 31岁。病程 3～ 32个月 ,平均 6个月。慢性前列腺炎症状指数 (NIH CPSI)平均 2 3.2分。　结果　 4 2 8例均行细菌培养 ,其中 2 4 8例 (5 7.94 % )分离出细菌。革兰阳性菌 195例 (78.6 3% ) ,其中葡萄球菌 16 0例 (6 4 .5 2 % ,16 0 / 2 4 8) ,CNS 89例 (35 .89% ,89/ 2 4 8) ;CNS中以表皮葡萄球菌为主者 81例 (32 .6 6 % ) ,其次为腐生葡萄球菌 3例和溶血性葡萄球菌 2例 ;NIH CPSI积分与细菌培养结果无明显相关。CNS对常用抗菌药物 (β 内酰胺类、喹诺酮类、氨基糖苷类 )耐药率较高 (5 1.9%～ 10 0 % )。　结论　CNS为慢性前列腺炎的主要致病菌 ,应引起高度重视 ;适时监测前列腺液病原菌及药敏试验对临床诊断和治疗慢性前列腺炎具有重要意义。     

Mecillinam susceptibility as an indicator of β-lactamase production in Staphylococcus aureus

The use of direct susceptibility testing from specimens has led to the fortuitous observation that penicillin-susceptible strains have larger inhibition zones for mecillinam than do β -lactamase producers. The current study was, therefore, undertaken to test 179 Staphylococcus aureus isolates for mecillinam susceptibility by Rosco Neo-Sensitabs and to compare the results with commonly used tests for β -lactamase production, i.e. size and character of penicillin inhibition zones, chromogenic cephalosporin (nitrocefin) tests and clover leaf assays. Agreement between methods was reached for 175 of 179 strains when disregarding the results of the nitrocefin tests, 88 isolates being found susceptible and 87 being found to be β -lactamase producers. All 88 susceptible isolates had mecillinam zones of >22 mm, with the great majority being >25 mm; double zones did, however, occur. The 87 β -lactamase producers had zones <14 mm or no zones. Four isolates presenting problems in had mecillinam zones of ≤20 mm and were without tapering growth at the penicillin inhibition zone edge. In conclusion, the size of the mecillinam inhibition zone is found to be a useful supplementary test in the clinically important distinction between β -lactamase-producing and non-producing isolates of S. aureus .