变性高效液相色谱分析肺炎支原体基因类型研究

目的 运用变性高效液相色谱（denaturing high-performance liquid chromatography,DHPLC）技术检测肺炎支原体（mycoplasma pneumoniae,MP）基因型,了解儿童MP流行状况.方法 对我院300例患儿鼻咽吸出物进行表型确证试验,采用PCR对标准菌株和MP表型阳性的临床菌株进行PCR扩增,扩增产物经DHPLC分析,通过与标准菌株色谱峰比对进行临床菌株基因分型.结果 300例患儿咽拭子经过24h培养后呈阳性110例.用特异引物扩增MP-129、MP-FH标准株和标本,分别得到2 280 bp和2580bp基因片段.110株临床菌株经DHPLC分析检测出107株是P1-Ⅰ型,均是1b亚型,3株是P1-Ⅱ型,均是2a亚型.结论 运用DHPLC技术检测出本院儿童发生MP感染以P1-Ⅰ、1b亚型为主.     

220例住院肺炎患儿肺炎支原体的分子检测及其基因分型研究

目的：通过检测社区获得性肺炎患儿支气管肺泡灌洗液了解肺炎支原体（MP）感染情况及基因分型特点。方法：采用聚合酶链反应（PCR）方法对220例住院肺炎患儿支气管肺泡灌洗液进行MP检测,用荧光定量PCR方法验证结果的准确性;将检测阳性标本用Hae Ⅱ 和Hae Ⅲ 两种限制性内切酶进行酶切反应后与标准株对比,分析阳性标本MP的基因分型。通过随机采样对部分阳性标本扩增产物测序证实基因分型的准确性。结果：220份标本MP的阳性检出率为55.0%（121/220）。MP感染好发于学龄前期和学龄期儿童（63.5%,101/159）,6个月内的婴儿阳性率最低20%（1/5）,无明显性别差异和季节差异。随机抽取的60份MP阳性标本经酶切反应均显示为P1-Ⅰ型,4份测序结果也证实其为P1-Ⅰ型。结论：MP是肺炎患儿的重要病原之一,有明显的年龄特征;流行型以P1-Ⅰ型为主。     

肺炎支原体肺炎患儿肺炎支原体基因分型及对大环内酯类抗生素耐药基因突变的分析

目的分析社区获得性肺炎(CAP)患儿肺炎支原体(MP)感染发生与基因分型以及大环内酯类耐药基因突变分型的流行病学特征, 为儿童肺炎支原体肺炎(MPP)的防治提供依据。方法检测佛山市三水区妇幼保健院2018年1月至2021年12月住院的620例CAP患儿MP感染情况, 对MP阳性者行P1限制性片段长度多态性分析(P1-RFLP), 分析阳性者及P1 Ⅰ型、P1 Ⅱ型在不同年份、性别、年龄、季节中分布的差异。检测MP对大环内酯类药物耐药基因突变情况, 分析23SrRNA结构域A2063G、A2064G突变在不同年份、性别、年龄、季节中分布的差异。结果 620例CAP患儿中, 198例MP阳性, 感染率为31.94%。2018至2021年各年MP阳性率差异不大。女性感染率高于男性。MP感染率随年龄增长逐渐升高, 以学龄前期、学龄期感染率最高。夏秋季感染率明显高于春冬季。198例患儿中, P1-RFLP分型P1-Ⅰ型157例(79.29%), P1-Ⅱ型41例(20.71%)。Ⅰ型、Ⅱ型在不同年份、性别、年龄、季节中分布差异不大。对其中143例患儿行大环内酯类药物耐药检测, 其中125例出现耐...     

一步法多重RT-PCR对儿童急性淋巴细胞白血病常见融合基因的检测及意义

目的 探讨一步法多重RT-PCR 对儿童急性淋巴细胞白血病(ALL)常见4 种融合基因的检测效果。方法 2003 年1 月至2010 年12 月确诊的ALL 患儿76 例,采集所有患儿骨髓标本,提取细胞总RNA,一步法多重RT-PCR 或常规巢式PCR 法对E2A/PBX1、MLL/AF4、BCR/ABL 和TEL/AML1 融合基因进行检测;DNA测序对PCR 结果进行验证。结果 一步法多重RT-PCR 及DNA 测序验证显示76 例ALL 患儿中,TEL/AML1融合基因阳性12 例(产物长度分别为298 bp 9 例,259 bp 3 例), E2A/PBX1 融合基因阳性3 例(产物长度373 bp), BCR/ABL 融合基因阳性1 例(产物长度2124 bp),MLL/AF4 融合基因阳性7 例(产物长度分别为427 bp 1 例,673 bp 6 例),与常规巢式PCR 检出结果一致。结论 一步法多重RT-PCR 可应用于儿童ALL 常见融合基因的检测。     

儿科Panton-Valentine杀白细胞素基因阳性耐甲氧西林金黄色葡萄球菌分子特征及Panton-Valentine杀白细胞素基因多态性和耐药特征

目的研究儿童感染Panton-Valentine杀白细胞素(PVL)基因阳性耐甲氧西林金黄色葡萄球菌(MRSA)分离株分子特征、PVL基因序列变异情况及耐药特征。方法对PVL阳性MRSA感染患儿临床资料进行回顾性调查,分析病例临床特征;采用PCR联合测序对MRSA临床分离株进行多位点序列分型(MLST);并应用分段PCR扩增PVL基因,拼接序列与日本ST30 CA-MRSA菌株NN1的PVL序列进行比对,分析序列变异与STs分型之间的关系,构建系统树对PVL基因的系统发生多样性进行分析;采用琼脂稀释法进行16种抗生素的体外药物敏感试验。结果在73例PVL阳性MRSA感染病例中:皮肤软组织感染35例(47.9%),肺炎27例(37.0%),其他严重感染及健康儿童分离株中也检测到PVL基因;社区感染55例(75.3%)。感染菌株MLST分型共有9种,最常见的3种为ST59、ST910、ST338,分别占53.4%、17.8%、9.6%。PVL序列检测及比对结果:共发现9个核苷酸位点变异,其中3个为非同义突变(16 C→A,62 C→T,527 A→G),其余均为同义突变;有9种不同的单体型序列,各STs分型在9种序列中呈散在分布;所研究的MRSA 71株为H型,仅2株R型。PVL阳性MRSA菌株对红霉素、克林霉素、头孢呋辛耐药率高达94.5%、86.3%和82.2%。结论 PVL阳性MRSA分离株来源广泛,以皮肤软组织感染和肺炎为主;分子分型以ST59型最为常见;lukSF-PV序列高度保守,序列变异主要为H型,R型罕见。     

氨苄西林耐药流感嗜血杆菌的基因分型研究

目的了解2000-2003年北京、上海、广州细菌耐药监测项目中,小于5岁呼吸道感染儿童鼻咽部携带氨苄西林（AMP）耐药流感嗜血杆菌（Haemophilus influenzae,Hi）的分子流行病学情况。方法对上述呼吸道感染儿童鼻咽部分离的899株Hi进行AMP敏感性检测,筛选出74株AMP耐药Hi,采用巢式PCR荚膜分型和玻片凝集法,对AMP耐药菌株进行b型Hi（Haemophilus influenzaetypeb,Hib）检测,并用脉冲电场凝胶电泳（pulsed—field gel electrophoresis,PFGE）和多重PCR两种方法,对AMP耐药菌株进行基因分型。结果74株AMP耐药Hi中,有2株Hib（占2．7％）。PFGE分型74株AMP耐药Hi中有38种基因型,具有克隆传播趋势的有5型,包括41株Hi（占55．4％）。其中菌株数最多的为A型,有18株,占24．3％,以2002年上海地区为主。多重PCR分型结果有31型,多重PCR与PFGE分型结果一致率为63．5％。结论北京、上海、广州三地区四年内小于5岁的呼吸道感染儿童鼻咽部携带的AMP耐药Hi有55．4％的菌株存在克隆传播。     

改进探针标记法斑点杂交在轮状病毒VP7分型中的应用

目的建立一种更加简便的A组人轮状病毒（HRV）核酸斑点杂交VP7分型方法,以便用于对HRV流行情况进行调查。方法在HRV VP7编码基因各G基因型间高度变异而型内高度保守区域设计分型探针,在该区域的两侧相对保守区域设计一对通用引物,利用PCR分别将地高辛标记HRV 5种常见型别（G1～4,G9型）的DNA探针,建立基于VP7的斑点杂交方法;选取经抗原检测和聚丙烯酰胺凝胶电泳（PAGE）检测均为HRV阳性的2006至2008年住院腹泻患儿粪便标本200份,RT-PCR扩增VP7全基因,并对扩增阳性产物应用斑点杂交方法进行G型别分析。结果建立的斑点杂交方法在5种型别探针间无交叉反应,各型探针的检测灵敏度可达到10 pg。200份PAGE阳性标本中162份RT-RCR扩增VP7基因阳性,斑点杂交显示G1型41例(25.3%),G2型2例(1.2%),G3型63例(38.9 %),G9型35例(21.6%),混合感染19例（11.7%）,杂交未分出型2例(1.2%),未检测到G4型HRV。结论本研究所建立的斑点杂交方法敏感度和特异度强,适合在HRV大规模分子流行病学调查时应用。通过该方法的初步应用,发现北京地区婴幼儿HRV除了常见的G1、G2和G3型外,还有G9型感染。     

肺炎支原体P1基因多态性分析现状

在肺炎支原体（MP）感染中，黏附宿主呼吸道上皮细胞和成功定植是感染的关键一步，黏附通过MP表面的P1蛋白介导与宿主细胞的分子受体结合。传统上基于川基因序列的变异将MP分为两型，新近研究表明，在川基因上有2个特异的重复序列，一个重复序列被命名为RepMP4，位于编码区的5’末端，另一个为RepMP2／3，位于3’末端。针对这两个重复序列的PER扩增，结果用限制性内切酶分析，使分型更为细致，共检出8个亚型，这表明在P1基因型中存在着更多的亚型。用基因的变异可能通过重复序列之间的重组出现在MP染色体其他的位置上而发生。通过直接测序法对临床分离株川基因更为广泛的研究可做出更精确的分型，以进一步了解用基因多态性的现状。     

腹泻患儿轮状病毒VP7基因分型与临床的关系

目的研究昆明地区2003年9~12月婴幼儿轮状病毒腹泻分子流行病学及临床特点。方法采用逆转录-聚合酶链法(RT-PCR)扩增婴幼儿腹泻大便样本中编码轮状病毒VP7蛋白的全基因片断,用巢式-聚合酶链反应(nested PCR)对扩增得到的VP7基因进行分型。结果1.昆明地区2003年秋冬季50份标本能定型者32份(占64%)。其中G3型占93.75%(30/32);混合型占6.25%(2/32),均为G1 G3型。本次分型中未见G2及G4型。2.2003年9~12月轮状病毒腹泻临床特点为病程短,脱水程度轻,腹泻次数少,电解质紊乱及酸中毒少见。结论1.2003年9~12月昆明地区轮状病毒流行株以G3型为主,与以往比较出现了流行株的转变。2.G1型感染的临床表现较G3型重。     

肺炎支原体肺炎患儿支气管肺泡灌洗液中肺炎支原体耐药基因和13种病原检测结果分析北大核心CSCD

目的:了解肺炎支原体肺炎(MPP)患儿支气管肺泡灌洗液(BALF)中肺炎支原体(MP)耐药基因和13种呼吸道病原的分布情况。方法:回顾性选择2018年1月至2019年1月北京大学第三医院和北京大学第一医院100例MPP患儿的BALF标本,采用荧光定量PCR法检测MP核酸及其耐药基因,采用多重PCR核酸检测技术测定甲型流感病毒、甲型流感病毒H_(1)N_(1)、甲型流感病毒H_(3)N_(2)、乙型流感病毒、人副流感病毒、腺病毒、人博卡病毒、人鼻病毒、肺炎衣原体、人偏肺病毒、MP、人冠状病毒和呼吸道合胞病毒核酸,采用 χ^(2)检验进行分析。 结果:100例BALF标本,荧光定量PCR法检测MP及耐药基因,83例(83.00%)MP阳性,其中78例(93.98%)耐药,均为23S rRNA结构域V区A2063G的点突变。多重PCR法检测13种呼吸道病原,89例(89.00%)阳性。79例(79.00%)检测到MP,其中74例(74.00%)仅检出MP,5例(5.00%)同时检出MP合并其他病原。10例(10.00%)检测出其他病原。0~4岁组病毒检出率高于>4~6岁组( P=0.042)和>6岁组( P=0.002),差异均有统计学意义。 结论:MPP患儿的BALF标本绝大部分可检测到MP,耐药现象严重,以A2063G的点突变为主。少数BALF标本中可检测到其他呼吸道病原和2种或3种病原。     

Allergic factors associated with the development of asthma and the influence of cetirizine in a double-blind, randomised, placebo-controlled trial: First results of ETA®

There is a common progression known as the allergic march from atopic dermatitis to allergic asthma. Cetirizine has several antiallergic properties that suggest a potential effect on the development of airway inflammation and asthma in infants with atopic dermatitis. Methods. Over a two year period, 817 infants aged one to two years who suffered from atopic dermatitis and with a history of atopic disease in a parent or sibling were included in the ETAC^® (Early Treatment of the Atopic Child) trial, a multi-country, double-blind, randomised, placebo-controlled trial. The infants were treated for 18 months with either cetirizine (0.25mg/ kg b.i.d.) or placebo. The number of infants who developed asthma was compared between the two groups. Clinical and biological assessments including analysis of total and specific IgE antibodies were performed. Results. In the placebo group, the relative risk (RR) for developing asthma was elevated in patients with a raised level of total IgE (≥ 30 kU/I) or specific IgE (≥ 0.35 kUA/I) for grass pollen, house dust mite or cat dander (RR between 1.4 and 1.7). Compared to placebo, cetirizine significantly reduced the incidence of asthma for patients sensitised to grass pollen (RR = 0.5) or to house dust mite (RR = 0.6). However, in the population that included all infants with normal and elevated total or specific IgE (intention-to-treat - ITT), there was no difference between the numbers of infants developing asthma while receiving cetirizine or placebo. The adverse events profile was similar in the two treatment groups. Discussion. Raised total IgE level and raised specific IgE levels to grass pollen, house dust mite or cat dander were predictive of subsequent asthma. Cetirizine halved the number of patients developing asthma in the subgroups sensitised to grass pollen or house dust mite (i.e. 20% of the study population). In view of the proven safety of the drug, we propose this treatment as a primary pharmacological intervention strategy to prevent the development of asthma in specifically sensitised infants with atopic dermatitis.     

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孤独症谱系障碍(autistic-spectrum disorders,ASDs)近年来患病率逐年攀升至1%左右,其症状往往伴随终生,成为严重威胁儿童健康和发展的神经发育性疾患;注意缺陷多动障碍(attention deficit hyperactivity disorder,ADHD)是儿童期最常见的精神障碍,国内报道患病率为4.13%～5.83%,其症状可延续至青少年期,甚至到成年期[1]。这两类精神障碍在成年期的临床表现、共患病、治疗策略和预后与儿童期有哪些不同呢?本文通过回顾相     

EXCITING NEW TREATMENT APPROACHES FOR PATHYPHYSIOLOGIC MECHANISMS OF SICKLE CELL DISEASE

During the past several decades, our understanding of the complex pathophysiology of vasoocclusion associated with sickle cell disease has improved greatly. Interaction of genes, hemoglobin molecules, red cell membrane and metabolic changes, cell-cell interactions and cell-plasma interactions, red cell adhesion to vascular endothelium, activation of coagulation, and vascular reactivity play a role in vaso occlusion. Penicillin prophylaxis of pneumococcal infections and appropriate use of blood transfusions and other supportive measures improved survival of sickle cell patients. Hydroxyurea made a major impact on sickle cell therapy when it was shown to decrease acute painful episodes, acute chest syndrome, and the need for blood transfusion in adults. Significant experience in the use of hydroxyurea has been accumulated in older children. The benefits and risks of hydroxyurea for younger children and long-term risks in all patients will be evaluated in future investigations. Other promising therapies include butyrate compounds, clotrimazole, magnesium supplementation, poloxamer 188, antiadhesion agents, anticoagulant approaches, and nitric oxide. Hemopoietic transplantation remains the only curative therapy. However, several transgenic mouse models are available for studies of gene therapy or other treatment approaches on biochemical, cellular, and pathologic effects of mutant genes.     

Infiltrations of Epstein-Barr virus-carrying cells in granular lymphocyte-proliferative disorders corresponding to chronic active Epstein-Barr virus infection

A 21-year-old man with granular lymphocyte-proliferative disorders (GLPD) associated with chronic active Epstein-Barr virus (EBV) infection is described. Chromosomal analyses revealed several clonal abnormalities and two of them were mainly repetitious. High copy numbers of monoclonal EBV genome were also detected in the proliferative large granular lymphocytes (LGLs), indicating the monoclonal expansion of EBV-infected LGLs. The patient had an indolent course for several years, and there was no evidence of infiltrations of his bone marrow until the end stage. At autopsy, microscopic studies revealed marked infiltrations of LGL in the liver and spleen, and the infiltrating cells were NK-cell immunophenotype. The infiltrated LGLs showed latency I.     

LUTEINIZING HORMONE RECEPTOR MUTATIONS IN DISORDERS OF SEXUAL DEVELOPMENT AND CANCER

Human male sexual development is regulated by chorionic gonadotropin (CG) and luteinizing hormone (LH). Aberrant sexual development caused by both activating and inactivating mutations of the human luteinizing hormone receptor (LHR) have been described. All known activating mutations of the LHR are missense mutations caused by single base substitution. The most common activating mutation is the replacement of Asp-578 by Gly due to the substitution of A by G at nucleotide position 1733. All activating mutations are present in exon 11 which encodes the transmembrane domain of the receptor. Constitutive activity of the LHR causes LH releasing hormone-independent precocious puberty in boys and the autosomal dominant disorder familial male-limited precocious puberty (FMPP). Both germline and somatic activating mutations of the LHR have been found in patients with testicular tumors. Activating mutations have no effect on females. The molecular genetics of the inactivating mutations of the LHR are more variable and include single base substitution, partial gene deletion, and insertion. These mutations are not localized and are present in both the extracellular and transmembrane domain of the receptor. Inactivation of the LHR gives rise to the autosomal recessive disorder Leydig cell hypoplasia (LCH) and male hypogonadism or male pseudohermaphroditism. Severity of the clinical phenotype in LCH patients correlates with the amount of residual activity of the mutated receptor. Females are less affected by inactivating mutation of the LHR. Symptoms caused by homozygous inactivating mutation of the LHR include polycystic ovaries and primary amenorrhea.     

Resurgence of measles in Singapore: profile of hospital cases

OBJECTIVE: To ascertain the profile of cases of measles seen at a general hospital during a recent outbreak that occurred despite a measles vaccination program. METHODOLOGY: A retrospective study from January 1991 to March 1998. All patients with measles (ICD code 055. 9) seen at the emergency unit or as inpatients were included. RESULTS: There were 87 cases identified. The diagnosis was clinical in all and proven serologically in 71%. Eighty-five per cent of the cases occurred between January 1997 and March 1998. There was a bi-modal age distribution with peaks in the very young (相似文献   

Personality profiles and heart rate variability (vagal tone) in children with recurrent abdominal pain

The aim of the study was to explore psychological factors and autonomic activity in children with recurrent abdominal pain and to compare them with those in a control group of healthy children. The Personality Inventory for Children was used for assessment of developmental, emotional and psychosocial factors in 25 children with recurrent abdominal pain (age, 7-15 y). Parasympathetic and sympathetic functions in these children and in 23 healthy control subjects (age, 7-13 y) were also investigated, non-invasively using a computerized polygraph. Vagal tone (parasympathetic function) was indexed by calculation of respiratory sinus arrhythmia in beats/min. Skin conductance (sympathetic function) was recorded by the constant current method. On the Personality Inventory for Children, 16 patients had high scores on somatic concern. Several patients had scores in the clinical range for depression, withdrawal and anxiety, but the mean scores for these personality profile scales were well within the normal range of healthy children. Interestingly, there was a spike on the L (Lie)-scale for most of the patients and 15 patients had scores above or close to the clinical cut-off value. As compared with the scores in healthy children, vagal tone and sympathetic tone were normal. Conclusion: Many children with recurrent abdominal pain have scores in the clinical range for depression, withdrawal, anxiety and L-scale indicating coping problems, denial and a trend towards somatic concern that may contribute to the evolution of abdominal pain. Autonomic nerve activity was not disturbed in these children.     

Overcoming surfactant inhibition with polymers

Inhibition of the function of pulmonary surfactant in the alveolar space is an important element of the pathophysiology of many lung diseases, including meconium aspiration syndrome, pneumonia and acute respiratory distress syndrome. The known mechanisms by which surfactant dysfunction occurs are (a) competitive inhibition of phospholipid entry into the surface monolayer (e.g. by plasma proteins), and (b) infiltration and destabilization of the surface film by extraneous lipids (e.g. meconium-derived free fatty acids). Recent data suggest that addition of non-ionic polymers such as dextran and polyethylene glycol to surfactant mixtures may significantly improve resistance to inhibition. Polymers have been found to neutralize the effects of several different inhibitors, and can produce near-complete restoration of surfactant function. The anti-inhibitory properties of polymers, and their possible role as an adjunct to surfactant therapy, deserve further exploration.     

Understanding infant formula

The World Health organisation recommends breast feeding infants for the first six months of life. When this breast feeding does not occur either through parental choice or medical need, infant formulas will be required. There is a bewildering array of formulas on the UK market for many different requirements. When faced with an unsettled infant many parents (and healthcare professionals) will experiment with the infant formula available and then attend the paediatric clinic looking for help and advice. It is therefore essential that paediatricians understand what milks are available and what the key differences between different products are. This review attempts to provide a simple guide through many of the formulations currently available in the UK; and offers advice for the dietary management of the child with extra calorie requirements, infants with cow's milk protein allergy, gastro oesophageal reflux disease, apparent unresolved hunger and infantile colic. Whatever the underlying condition, there is likely to be an infant formula that is suitable in this generation of ever expanding formulations.