儿童李斯特菌脑炎两例

本研究通过回顾性分析2例诊断为LM脑膜炎患儿的临床资料,总结两例儿童单核细胞增多性李斯特菌(LM)脑膜炎的临床特点及治疗转归。此两例患儿以发热、头痛、呕吐起病,脑脊液培养均为LM生长,早期经头孢类药物抗感染治疗无效,换用氨苄青霉素联合效果显著。LM脑膜炎在免疫功能正常儿童发生率低,但病死率及后遗症发生率高。发病多以发热、呕吐、头痛为主要临床表现,使用氨苄青霉素效果较好,脑脊液培养有助于诊断。     

3例婴儿脑膜炎败血伊丽莎白菌脑膜炎临床分析

总结脑膜炎败血伊丽莎白菌感染患儿的临床特点,以期提高临床诊治水平。收集中山市博爱医院2010年1月至2020年12月收治的3例脑膜炎败血伊丽莎白菌引起的脑膜炎患儿临床资料,结合相关文献进行总结分析。结果显示,3例患儿脑脊液中培养出脑膜炎败血伊丽莎白菌,其中2例血培养为同种细菌。3例患儿均为足月顺产,2例为新生儿期发病,1例为出生后35天发病,起病前均未发现致病高危因素。患儿以发热为主要起病表现,无抽搐及脑膜刺激征表现。3例患儿外周血常规白细胞总数、C反应蛋白、降钙素原均升高;脑脊液潘氏试验均为阳性,伴脑脊液白细胞数增高,脑脊液蛋白明显增高,脑脊液葡萄糖降低。头颅磁共振成像(magnetic resonance imaging,MRI)或电子计算机断层扫描术(computer tomography,CT)可见脑膜强化、软脑膜增厚、脑外间隙增宽,均无脑实质受累。3例检出菌株的药敏结果表现出高度一致性,均提示对环丙沙星、哌拉西林/他唑巴坦、左氧氟沙星、复方新诺明敏感。2例患儿以环丙沙星治疗,1例患儿以美罗培南联合万古霉素治疗。其中1例治愈,2例临床症状好转后出院。经电话随访,3例患儿一般情况尚可,无生长发育异常。本病例报道提示,脑膜炎败血伊丽莎白菌脑膜炎主要见于新生儿期,以发热起病为主要表现,该菌对儿科常用抗菌药物多显示体外耐药,对喹诺酮类药物敏感。     

血清降钙素原在小儿早期中枢神经系统感染鉴别诊断中的价值

目的研究细菌性脑膜炎和病毒性脑炎患儿血清降钙素原（PCT）水平的变化,探讨其在小儿早期中枢神经系统感染鉴别诊断中的价值。方法采用双抗夹心免疫发光测量法,检测78例早期中枢神经系统感染患儿的血清及脑脊液降钙素原水平,分析它们之间的相关性;并和脑脊液白细胞数、血白细胞数、血清C反应蛋白相对比;PCT阳性细菌性脑膜炎患儿使用抗生素治疗后再测定血清PCT。结果33例急性细菌性脑膜炎患儿的血清降钙素原浓度为（18．46±9．18）ng／mL,45例急性病毒性脑炎的血清降钙素原水平轻度升高（0．58±0．31）ng／mL,P〈0．01,两者相比较差异有统计学意义。细菌性脑膜炎组患儿血清PCT在经过抗生素治疗后较入院时明显下降,两者比较差异有统计学意义（P〈0．05）。并且在细菌组急性期脑脊液PCT与血清PCT存在正相直线相关性。结论血清降钙素原检测在小儿早期中枢神经系统感染鉴别诊断中有重要应用价值。     

脑脊液检查在早产儿及足月儿细菌性脑膜炎的诊断价值

目的:评价脑脊液检查在早产儿及足月儿细菌性脑膜炎诊断中的价值。方法:选取2014年6月1日至2016年12月31日上海市儿童医院新生儿科收治的行腰椎穿刺检查的447例新生儿,回顾性分析新生儿的一般资料、脑脊液常规生化、培养等指标,根据胎龄将患儿分为早产儿167例与足月儿280例,再根据有无患发细菌性脑膜炎分为早产儿细菌性脑膜炎27例(早产儿观察组)、早产儿非细菌性脑膜炎140例(早产儿对照组)、足月儿细菌性脑膜炎38例(足月儿观察组)、足月儿非细菌性脑膜炎242例(足月儿对照组),采用受试者工作特征(ROC)曲线评估蛋白定量、白细胞计数、葡萄糖对早产儿及足月儿细菌性脑膜炎的诊断价值。结果:与同组对照组相比,足月儿观察组和早产儿观察组蛋白定量和白细胞计数均明显升高,而葡萄糖含量显著下降,且差异均具有统计学意义(P0.05);本研究65例细菌性脑膜炎患儿脑脊液培养分离出11株细菌(16.9%)。足月儿脑脊液白细胞计数、蛋白定量以及葡萄糖诊断细菌性脑膜炎的ROC曲线下面积分别为0.995、0.846、0.703。早产儿脑脊液白细胞计数、蛋白定量以及葡萄糖诊断细菌性脑膜炎ROC曲线下面积分别为0.970、0.711、0.705。结论:脑脊液白细胞计数、蛋白定量在足月儿和早产儿细菌性脑膜炎中具有较高的诊断价值。     

电化学方法在李斯特氏菌毒力基因检测中的应用

单核李斯特氏菌(Listeria monocytogenes, LM)是食源性李斯特氏病的病源菌, 该病可引起败血病、脑膜炎、流产等。李斯特氏菌的毒力因子listeriolysin O (LLO)是引发李斯特氏病的主要原因。文章使用一种特殊的电化学方法从样品中检测编码LLO的hlyA基因。该方法以化合物Nhydroxysulfosuccinimide (NHS) 和 N-(3-dimethylamion) propyl- N'-ethyl carbodiimidehydrochloride (EDC) 作为激活剂, 使单链DNA探针结合到金电极表面组成工作电极, 以[Co(phen)₃](ClO₄)₃ 作为指示剂来检测循环伏安曲线(Cyclic voltammetry , CV), 通过CV峰值的变化来估算hlyA基因的含量, 从而确定LM的污染情况。这种新颖的电化学方法用于免标记的目标DNA的杂交检测, 具有快速和方便的特点。     

055产单核细胞李斯特菌胃肠炎

产单核细胞李斯特菌通常被认为是一种食物源性的致病菌，在细胞免疫缺陷的病人，包括新生儿、孕妇、老年人以及移植术后免疫缺陷患者中引起菌血症和脑膜脑炎。该菌在健康人群中可以引起急性、自限性、发热性胃肠炎。目前至少有7次由于产单核细胞李斯特菌引起的食物源性胃肠炎的暴发流行。规模最大的一次发生在1997年，受累人数达1566人。     

临床标本中一组黄杆菌的分离和鉴定

本文报道从5例肺炎病人痰液,1例化脓性脑膜炎病人血及脑脊液检出的7株黄杆菌,经裹型特征和遗传型特征的系统鉴定,4株为黄杆菌Ⅱ群菌,2株为大比目鱼黄杆菌,另一株为脑膜脓毒黄杆菌。测试了7株菌的药物敏感性和动物致病性。讨论了黄杆菌与临床感染的关系。     

毒力基因调控蛋白PrfA促进单核细胞增生李斯特菌生物被膜的形成

单核细胞增生李斯特菌(Listeria monocytogenes,LM)是重要的革兰氏阳性食源性致病菌,易在食品以及各种食品加工、运输和保藏设备的接触面形成生物被膜,从而具有更强的抗逆性而难以彻底清除,因此成为食品卫生安全的重要隐患.PrfA是LM毒力基因转录表达的重要调控因子,通过比较研究LM野生株(EGD和EGDe)、PrfA缺失株(EGDAprfA和EGDeAprfA)、无害李斯特菌(Listeria innocua,LI),携带组成性表达PrfA蛋白的重组无害李斯特菌(LI-pERL3-prfA*)以及重组单核细胞增生李斯特菌(EGDeΔprfA-pERL3-prfA*)生物被膜形成能力的差异,探讨LM重要的毒力调控蛋白PrfA对生物被膜形成的影响.实验结果显示:LM野生株具有较强的生物被膜形成能力,而LI形成生物被膜的能力最弱;PrfA的缺失能降低LM生物被膜的形成能力;组成性高量表达PrfA蛋白可以回复EGDeΔprfA的生物被膜形成能力,但对LI没有增强作用.以上实验结果表明:PrfA在LM生物被膜形成中具有重要的促进作用.     

单增李斯特菌溶血素O的功能研究进展

单核细胞增生李斯特菌(Listeria monocytogenes, Lm,简称单增李斯特菌)是一种普遍存在的革兰阳性食源性病原体,可引起人类和一些动物的李斯特菌病。侵袭性李斯特菌病通常很严重,临床上表现为自然流产、败血症和脑膜脑炎,也可表现为发热性胃肠炎综合症。成孔蛋白单增李斯特菌溶血素O(Listeriolysin O,LLO,由hly基因编码)是一种重要的毒力因子,属于胆固醇依赖性细胞溶解素(cholesterol-dependent cytolysins,CDC)毒素,其通过膜穿孔机制介导Lm从吞噬体逃逸并引起李斯特菌病。最近的研究表明LLO除了主要的膜穿孔作用,还存在其他功能,在Lm感染过程中扮演了重要的角色。从LLO的功能和作用机制等方面综述了近些年对该毒素的研究进展,以便更好地理解单增李斯特菌的感染机制,为防治李斯特病的相关研究提供参考。     

单核细胞增生李斯特菌PrfA蛋白转录调控毒力基因表达的分子机制

单核细胞增生李斯特菌(Listeria monocytogenes LM)属于典型的细胞内寄生革兰氏阳性菌,是WHO公布的四大食源性致病菌之一.LM不仅是人畜共患传染病李斯特菌病(listeriosis)的主要病原菌,也是研究胞内感染和细胞介导的免疫应答的模式细菌.绝大多数LM毒力基因的转录表达受到PrfA蛋白的调控.本文简单介绍了LM侵染宿主细胞必需的毒力基因及其产物;重点对毒力基因调节蛋白PrfA的结构和功能,PrfA调节毒力基因表达的主要方式最新进展进行了综述和讨论.     

Prevention of allograft tolerance by bacterial infection with Listeria monocytogenes

Exposure to certain viruses and parasites has been shown to prevent the induction of transplantation tolerance in mice via the generation of cross-reactive memory T cell responses or the induction of bystander activation. Bacterial infections are common in the perioperative period of solid organ allograft recipients in the clinic, and correlations between bacterial infections and acute allograft rejection have been reported. However, whether bacterial infections at the time of transplantation have any effect on the generation of transplantation tolerance remains to be established. We used the Gram-positive intracellular bacterium Listeria monocytogenes (LM) as a model pathogen because its effects on immune responses are well described. Perioperative LM infection prevented cardiac and skin allograft acceptance induced by anti-CD154 and donor-specific transfusion in mice. LM-mediated rejection was not due to the generation of cross-reactive T cells and was largely independent of signaling via MyD88, an adaptor for most TLRs, IL-1, and IL-18. Instead, transplant rejection following LM infection was dependent on the expression of the phagosome-lysing pore former listeriolysin O and on type I IFN receptor signaling. Our results indicate that bacterial exposure at the time of transplantation can antagonize tolerogenic regimens by enhancing alloantigen-specific immune responses independently of the generation of cross-reactive memory T cells.     

单增李斯特菌感染胎盘相关毒力因子的研究进展

单核细胞增生李斯特氏菌(Listeria monocytogenes,LM)是一种可引起李斯特菌病的食源性致病菌。由于妊娠相关免疫缺陷和LM对非吞噬细胞独特的细胞内感染能力,孕妇是LM的主要目标人群。LM可穿过胎盘屏障,对胎儿造成重大伤害,包括早产、流产甚至死产。胎盘特异性毒力因子的作用对LM感染期间穿过胎盘屏障并感染胎儿尤为重要。文中介绍了国内外近年在孕妇中发生LM感染的事件,详细讨论了LM垂直传播以及在胎盘定殖机制方面的研究进展,着重讨论并分析了LM与感染胎盘相关毒力因子的最新发现,以期为今后防控LM的胎盘感染并保障食品安全提供参考。     

TNF is important for pathogen control and limits brain damage in murine cerebral listeriosis

Cerebral listeriosis is a life-threatening disease. However, little is known about the bacterial virulence factors responsible for the severe course of disease and the factors of the immune system contributing to the control of Listeria monocytogenes (LM) or even to the damage of the brain. To analyze the importance of the actA gene of LM, which mediates cell-to-cell spread of intracellular LM, the function of TNF in murine cerebral listeriosis was studied. C57BL/6 mice survived an intracerebral (i.c.) infection with actA-deficient LM, but succumbed to infection with wild-type (WT) LM. Upon infection with actA-deficient LM, macrophages and microglial cells rapidly, and later LM-specific CD4 and CD8 T cells, produced TNF. In contrast to WT mice, TNF-deficient animals succumbed to the infection within 4 days due to failure of control of LM. Histology identified a more severe meningoencephalitis, brain edema, and neuronal damage, but a reduced inducible NO synthase expression in TNF-deficient mice. Reciprocal bone marrow chimeras between WT and TNF-deficient mice revealed that hematogenously derived TNF was essential for survival, whereas TNF produced by brain-resident cells was less important. Death of TNF-deficient mice could be prevented by LM-specific T cells induced by an active immunization before i.c. infection. However, brain pathology and inflammation of immunized TNF-deficient mice were still more severe. In conclusion, these findings identify a crucial role of TNF for the i.c. control of LM and survival of cerebral listeriosis, whereas TNF was not responsible for the destruction of brain tissue.     

In Vitro Antibacterial Activity of Phlorotannins from Edible Brown Algae, <Emphasis Type="Italic">Eisenia bicyclis</Emphasis> Against Streptomycin-Resistant <Emphasis Type="Italic">Listeria monocytogenes</Emphasis>

Listeria monocytogenes (LM) is an important food borne pathogen responsible for listeriosis. Further, LM is an etiological agent associated with life threatening conditions like meningitis and encephalitis. Biofilm forming and drug resistant LM may potentially become difficult to treat infections and hence effective controlling measures are required to prevent LM infections. In view of this, the present study evaluated an anti-listerial potential of edible brown seaweed, Eisenia bicyclis, by disc diffusion and micro-dilution methods. The results of the present study suggested that the anti-listerial activity of various phlorotannins isolated form E. bicyclis were in the range of 16–256 µg/ml. Among the phlorotannins isolated, fucofuroeckol-A (FAA) exhibited the highest anti-listerial potential (MIC range 16–32 µg/ml) against LM strains tested. Further, in checker board synergy assays, FFA-streptomycin combination exhibited significant synergy (fractional inhibitory concentration index, ∑FIC < 0.5) against aminoglycoside resistant clinical strains of LM. The results of the present study suggested the potential use of edible seaweed E. bicyclis as a source of natural phlorotannins to control food borne pathogenic infections.     

Cutting edge: protective cell-mediated immunity to Listeria monocytogenes in the absence of myeloid differentiation factor 88

In addition to their role in triggering innate immune responses, Toll-like receptors are proposed to play a key role in linking the innate and adaptive arms of the immune response. The majority of cellular responses downstream of Toll-like receptors are mediated through the adapter molecule myeloid differentiation factor 88 (MyD88), and mice with a targeted deletion of MyD88 are highly susceptible to bacterial infections, including primary infection with Listeria monocytogenes (LM). In contrast, herein we demonstrate that MyD88-deficient mice have only a modest impairment in their LM-specific CD4 T cell response, and no impairment in their CD8 T cell response following infection with ActA-deficient LM. Furthermore, CD8 T cells from immunized MyD88-deficient mice protected naive recipient mice following adoptive splenocyte transfer, and immunized MyD88-deficient mice were protected from infection with wild-type LM. These results indicate that adaptive immune responses can be generated and provide protective immunity in the absence of MyD88.     

Membrane fusion induced by the catalytic activity of a phospholipase C/sphingomyelinase from Listeria monocytogenes

Listeria monocytogenes is a bacterium responsible for localized and generalized infections in humans and animals. It has the ability to spread from the cytoplasm of an infected cell to neighboring cells without becoming exposed to the extracellular space. The bacterium secretes a phospholipase C (PLC(LM)) that is active on glycerophospholipids, e.g., phosphatidylcholine, and on sphingomyelin; thus, PLC(LM) should be described more appropriately as a phospholipase C/sphingomyelinase. We have obtained PLC(LM) free from a frequent contaminant, listeriolysin O, using an improved purification procedure. PLC(LM) has been assayed on large unilamellar liposomes of defined lipid composition. The enzyme is activated by K(+) and Mg(2+), and readily degrades phospholipids in bilayer form, in the absence of detergents. Enzyme activity is accompanied by important changes in the structure of the phospholipid vesicles, namely, vesicle aggregation, intervesicular mixing of lipids, and mixing of aqueous contents, with very low leakage of vesicular contents. The data are interpreted as indicative of PLC(LM)-induced vesicle fusion. This is confirmed by the demonstration of intervesicular mixing of inner monolayer lipids, using a novel procedure. The observation of PLC(LM)-induced membrane fusion suggests a mechanism for the cell-to-cell propagation of the bacterium, which requires disruption of a double-membrane vacuole.     

Cross-reactive antigen is required to prevent erosion of established T cell memory and tumor immunity: a heterologous bacterial model of attrition

Induction and maintenance of T cell memory is critical for the control of intracellular pathogens and tumors. Memory T cells seem to require few "maintenance signals," though often such studies are done in the absence of competing immune challenges. Conversely, although attrition of CD8(+) T cell memory has been characterized in heterologous viral models, this is not the case for bacterial infections. In this study, we demonstrate attrition of T cell responses to the intracellular pathogen Listeria monocytogenes (LM) following an immune challenge with a second intracellular bacterium, Mycobacterium bovis (bacillus Calmette-Guérin, BCG). Mice immunized with either LM or recombinant LM (expressing OVA; LM-OVA), develop a potent T cell memory response. This is reflected by peptide-specific CTL, IFN-gamma production, and frequency of IFN-gamma-secreting T cells to native or recombinant LM Ags. However, when the LM-infected mice are subsequently challenged with BCG, there is a marked reduction in the LM-specific T cell responses. These reductions are directly attributable to the effects on CD4(+) and CD8(+) T cells and the data are consistent with a loss of LM-specific T cells, not anergy. Attrition of the Ag (OVA)-specific T cell response is prevented when LM-OVA-immunized mice are challenged with a subsequent heterologous pathogen (BCG) expressing OVA, demonstrating memory T cell dependence on Ag. Although the reduction of the LM-specific T cell response did not impair protection against a subsequent LM rechallenge, for the first time, we show that T cell attrition can result in the reduction of Ag-specific antitumor (B16-OVA) immunity previously established with LM-OVA immunization.     

Resistance to tumor growth mediated by Listeria monocytogenes. Destruction of experimental malignant melanoma by LM-activated peritoneal and lymphoid cells.

A murine experimental model of nonspecific tumor destruction mediated by cells activated by Listeria monocytogenes (LM) is described. B16 melanoma growth is prevented or suppressed in the syngeneic host when tumor cells are inoculated in contact with viable LM. In vitro, cultured B16 cells are destroyed by LM immune peritoneal or splenic cells in the presence of the bacterial antigen(s). Activation of LM immune cells in vitro is immunologically specific. Replacement of LM by sheep red blood cells or bovine serum albumin in the in vitro cultures aborts the cytotoxic effect. Further, no tumor cell killing is obtained when thioglycollate-induced or normal peritoneal cells are substituted for LM immune cells in the in vitro cultures. Normal spleen cells in the presence of LM are weakly cytotoxic for B16 cells. Normal peritoneal cells plus LM or LM alone are not. Elimination of thymus derived "T" cells by anti-theta C3H or rabbit anti-mouse brain serum (RAMB) abrogated the cytotoxic effect. Therefore, LM-induced tumor destruction probably occurs through nonspecific mechanism(s) consequent to activation of host "T" cells by specific immune reactivity to LM antigen(s).     

Characterization of a Listeria monocytogenes protein interfering with Rab5a

Listeria monocytogenes (LM) phagocytic strategy implies recruitment and inhibition of Rab5a. Here, we identify a Listeria protein that binds to Rab5a and is responsible for Rab5a recruitment to phagosomes and impairment of the GDP/GTP exchange activity. This protein was identified as a glyceraldehyde-3-phosphate dehydrogenase (GAPDH) from Listeria (p40 protein, Lmo 2459). The p40 protein was found within the phagosomal membrane. Analysis of the sequence of LM p40 protein revealed two enzymatic domains: the nicotinamide adenine dinucleotide (NAD)-binding domain at the N-terminal and the C-terminal glycolytic domain. The putative ADP-ribosylating ability of this Listeria protein located in the N-terminal domain was examined and showed some similarities to the activity and Rab5a inhibition exerted by Pseudomonas aeruginosa ExoS onto endosome–endosome fusion. Listeria p40 caused Rab5a-specific ADP ribosylation and blocked Rab5a-exchange factor (Vps9) and GDI interaction and function, explaining the inhibition observed in Rab5a-mediated phagosome–endosome fusion. Meanwhile, ExoS impaired Rab5-early endosomal antigen 1 (EEA1) interaction and showed a wider Rab specificity. Listeria GAPDH might be the first intracellular gram-positive enzyme targeted to Rab proteins with ADP-ribosylating ability and a putative novel virulence factor.