The role of pneumolysin in pneumococcal pneumonia and meningitis

Diseases caused by Streptococcus pneumoniae include pneumonia, septicaemia and meningitis. All these are associated with high morbidity and mortality. The pneumococcus can colonize the nasopharynx, and this can be a prelude to bronchopneumonia and invasion of the vasculature space. Proliferation in the blood can result in a breach of the blood-brain barrier and entry into the cerebrospinal fluid (CSF) where the bacteria cause inflammation of the meningeal membranes resulting in meningitis. The infected host may develop septicaemia and/or meningitis secondary to bronchopneumonia. Also septicaemia is a common precursor of meningitis. The mechanisms surrounding the sequence of infection are unknown, but will be dependent on the properties of both the host and bacterium. Treatment of these diseases with antibiotics leads to clearance of the bacteria from the infected tissues, but the bacteriolytic nature of antibiotics leads to an acute release of bacterial toxins and thus after antibiotic therapy the patients can be left with organ-specific deficits. One of the main toxins released from pneumococci is the membrane pore forming toxin pneumolysin. Here we review the extensive studies on the role of pneumolysin in the pathogenesis of pneumococcal diseases.     

Activity of telithromycin and seven other agents against 1034 pediatric Streptococcus pneumoniae isolates from ten central and eastern European centers

Objective  To test the activity of telithromycin against 1034 Streptococcus pneumoniae isolates from pediatric patients in ten centers from ten central and eastern European countries during 2000–2001, and to compare it with the activities of erythromycin A, azithromycin, clarithromycin, clindamycin, and quinupristin–dalfopristin.
Methods  The minimum inhibitory concentrations (MICs) of telithromycin, erythromycin A, azithromycin, clarithromycin, clindamycin, levofloxacin, quinupristin–dalfopristin and penicillin G were tested by the agar dilution method with incubation in air, and mechanisms of resistance to macrolides and quinolones were investigated.
Results  Strains were isolated from sputum, tracheal aspirates, ear, eye, blood, and cerebrospinal fluid. Among S. pneumoniae strains tested, 36% had raised penicillin G MICs (≥ 0.12 mg/L). Susceptibilities were as follows: telithromycin, quinupristin–dalfopristin and levofloxacin, ≥ 99%; clindamycin, 83%; and erythromycin A, azithromycin and clarithromycin, 78%. Of 230 (22.3%) erythromycin A-resistant S. pneumoniae strains, 176 (79.6%) had erm(B) , 38 (16.1%) had mef(A) , and 10 (4.3%) had mutations in 23S ribosomal RNA or in ribosomal protein L4. The rates of drug-resistant S. pneumoniae are high in all centers except Kaunas, Riga, and Prague.
Conclusion  Telithromycin had low MICs against all strains, irrespective of macrolide, azalide or clindamycin resistance. Ribosomal methylation was the most prevalent resistance mechanism among all resistant strains, except in Sofia, where the prevalence of the efflux mechanism was higher.     

肺炎衣原体对血管内皮细胞的感染及其对ICAM-1表达的影响

目的:观察肺炎衣原体对人脐静脉内皮细胞(HUVECs)的感染及其对细胞分泌和表达细胞间粘附分子1(ICAM-1)的影响,探讨C.pneumoniae感染在动脉粥样硬化形成中的作用及其可能机制。方法:用人喉表皮癌(HEP-2)细胞培养C.pneumoniae,以C.pneumoniae感染HUVE细胞,经透射电镜及PCR检测有无感染。用流式细胞仪检测感染前后HUVE细胞表面ICAM-1蛋白的表达的变化,用荧光定量RT-PCR检测ICAM-1mRNA的变化。结果:C.pneumoniae能感染体外培养的HUVE细胞;感染后12h,细胞表面ICAM-1蛋白的表达即增加,其峰值约在感染后24h;荧光定量RT-PCR结果显示其增加在mRNA水平。结论:C.pneumoniae能感染体外培养的人脐静脉内皮细胞并增加ICAM-1的表达,提示C.pneumoniae感染可能是动脉粥样硬化的始动因子之一,其致动脉粥样硬化机制可能与感染后血管内皮细胞粘附分子表达的增加有关。     

Monocyte cytokine secretion induced by chemically-defined derivatives of Klebsiella pneumoniae.

The capacity of a K. pneumoniae membrane proteoglycan (Kp-MPG) and four of its chemically defined derivatives to activate human monocytes was studied by measuring immunoreactive IL-1 beta, IL-6 and tumour necrosis factor-alpha (TNF-alpha) in culture supernatants. Monocyte culture supernatants were also tested for their comitogenic activity on concanavalin A-stimulated thymocytes and for their cytotoxic activity on the mouse fibroblastic L929 cell line. The four Kp-MPG derivatives were: (i) an acylpoly(1-3)galactoside (APG); (ii) an APG preparation submitted to acid hydrolysis which removed all fatty acids but left intact the galactose chain of APG (GC-APG); (iii) a preparation obtained by mild alkaline hydrolysis, containing additional ester-linked C14 and C16 fatty acids bound to the APG molecule (EFA-APG); and (iv) a polymer of the latter compound (APG pol). Kp-MPG induced the synthesis of IL-1 beta, IL-6 and TNF-alpha with dose-responses and kinetics similar to those of Salmonella minnesota lipopolysaccharide (Sm-Re-LPS). APG pol and EFA-APG induced the secretion of the three cytokines with lower potency than Kp-MPG or Sm-Re-LPS. APG did not trigger any detectable cytokine production and GC-APG induced only borderline and inconsistent responses. Our data demonstrate the critical role of ester-linked C14 and C16 fatty acids in the triggering of monocyte response to Kp-MPG derivatives.     

Distribution of genotypes and antibiotic resistance genes among invasive Streptococcus agalactiae (group B streptococcus) isolates from Australasian patients belonging to different age groups.

Serotype distribution and antibiotic resistance (AR) among group B streptococci (GBS) affect GBS disease prevention strategies, but vary among patient groups. A multiplex PCR-based reverse line blot (mPCR/RLB) hybridisation assay was used to compare the distributions of GBS serotypes, serotype III subtypes and AR-associated genes among 666 invasive isolates from 663 patients, divided into five age groups: infants, early-onset (EO; 0-6 days) and late-onset (LO; 7-90 days); children (aged 3 months to 14 years); women of childbearing age (WCBA; aged 15-45 years); and other adults (males aged >15 years; females aged >45 years). Serotypes Ia and V and serosubtype III-1 accounted for 60% of infections. Serosubtype III-2, which corresponds to a virulent clone belonging to sequence type (ST)17, was relatively uncommon overall (7%), but was associated strongly with LO infant infections, in which it was significantly more common than in adult infections (25/104 (24%) vs. 9/392 (2%), p <0.0001) or in EO infections (25/104 (24%) vs. 14/155 (9%), p <0.005). Erythromycin resistance genes were found in 8% of all isolates (ermB 3%, ermA 2.5% and mefA/E 2%), in 11-15% of isolates of serotypes II and V and subtype III-1, but in none of the isolates of serosubtype III-2 (III-2, 0/49 vs. all others, 54/618 (9%), p <0.04). In summary, the virulent serosubtype III-2 was associated strongly with LO infant GBS infection, but was less likely than other serotypes or serosubtype III-1 to carry AR genes.     

运用变性高效液相色谱对肺炎克雷伯菌产ESBL进行基因分型

目的 通过运用变性高效液相色谱（DHPLC）技术对前期研究已确认产超广谱β-内酰胺酶（ESBL）的肺炎克雷伯菌临床分离株TEM型质粒进行基因分型,试图建立一种方便快捷的用于ESBL分子诊断及其流行病学监测的新方法.方法 利用PCR技术从肺炎克雷伯菌临床分离株中扩增出TEM型质粒的编码序列,扩增产物运用DHPLC技术进行分析,分析提示,异常的样本通过测序确定其基因突变的类型,最后通过比对确定其基因型.结果 共分析了101例肺炎克雷伯菌临床分离株,全部样本均扩增出TEM型质粒的编码序列,经过DHPLC分析,52例（51.4%）样本表现为单一的洗脱峰,其形态与TEM-1标准菌株的峰型相一致,测序确定它们的碱基序列亦相一致,不存在变异,为TEM-1型;49例（48.6%）样本表现为异常的洗脱峰,它们均为双峰,形态一致,但异源双链峰的高度有差异,测序结果表明它们均存在四种相同的基因突变,在NCBI网站比对后确定为TEM-116;测序结果还提示,部分样本中TEM-1和TEM-116混合存在,其比例的不同表现为DHPLC时异源双链峰高度的差异;文献检索表明,本次确定的TEM-116为一新的基因亚型,为国内首次报道.结论 DHPLC具有简便快捷、高通量和自动化的特点,重复性好,不仅可对已知突变作出即时诊断,还可发现新的基因亚型,不失为一种较好的ESBL分子诊断方法及其流行病学监测手段.     

Subtyping of Mycoplasma pneumoniae isolates based on extended genome sequencing and on expression profiles

Mycoplasma pneumoniae isolates from patients, collected over a period of 12 years in Germany, were characterized by various methods (parameters) including multilocus sequence typing, restriction fragment length polymorphisms, Western blotting with mono-specific antibodies directed against selected proteins or with polyspecific antibodies directed against the Triton X-114-soluble protein fraction, and two-dimensional gel electrophoresis. The results for 91 isolates from Germany, which were complemented with 14 isolates from the USA and 10 isolates from France, clearly showed that M. pneumoniae is a highly uniform species and that most of the isolates could be assigned to one of the two subtypes 1 and 2. The members of one subtype differ from the other with respect to the sequence of the P1 gene, the ORF6 gene, the P65 gene, and by a typical DNA restriction fragment pattern. We observed four isolates (variants), which seemed identical by the above mentioned criteria, but did not belong to either one of the two subtypes. They showed most of the subtype 2-specific features, but differed in the sequence of the P1 gene and showed a variation in the restriction fragment pattern. The appearance of subtype 1 or 2 over the last 12 years in Germany showed a dominance of subtype 1 between 1989 and 1996 and a dominance of subtype 2 between 1997 and 1998. The variant (neither subtype 1 nor subtype 2) was only detected in 1991 and 1995 but it had no epidemiological consequences.     

Evaluation of a commercial test for antibodies to the chlamydial lipopolysaccharide (Medac) for serodiagnosis of acute infections by Chlamydia pneumoniae (TWAR) and Chlamydia psittaci

A commercial test (rELISA) based on a recombinant chlamydial lipopolysaccharide (LPS) antigen has been evaluated for the diagnosis of acute infections caused by Chlamydia pneumoniae (TWAR) and Chlamydia psittaci. This test and a microimmunofluorescence test (MIF) were compared in 160 patients with community-acquired pneumonia. Seventeen of nineteen cases with significant titre changes detected by rELISA were confirmed by MIF. The two remaining cases not confirmed by MIF were considered false-positive reactions. One case positive by MIF only was judged not to be a true-positive reaction. All three cases occurred in patients with Mycoplasma pneumoniae infection and may be the result of a mitogenic effect. High antibody titres have been used to indicate acute C. pneumoniae infection. We found high MIF or rELISA titres to be equally common in patients and controls; no association between the two tests was detected. An unexpected cross-reactivity between the rELISA antigen and parvovirus was observed, which might have diagnostic implications. Both MIF and rELISA detected acute C. pneumoniae and C. psittaci infection, and there was good agreement between the tests. Single serum diagnosis was generally not feasible with either MIF or rELISA.