Protective effect of DNA vaccine with the gene encoding 55kDa antigen fragment against Pneumocystis carinii in mice

Objective:To evaluate the protective effect of DNA vaccine with the gene encoding 55kDa antigen fragment of Pneumocystis carinii(P.carina) against P.carina in mice.Methods:The fragment of the antigen within p55(p55-582) was cloned.Then recombinant plasmid was constructed based on the eukaryotic expression vector pcDNA3.1(+).BALB/c mice were used as experimental models to examine the immunogenicity of pcDNA3.1(+)-p55-582.ELBA and RTPCR were used to evaluate the role of this kind of DNA vaccine.Results:The results of western blot indicated that the recombinant DNA[pcDNA3.1(+)-p55-582]could be expressed correctly and had antigenicity in transfected COS-7 cells.ELBA and RT-PCR showed that pcDNA3.1(+)- p55-582 elicited antibody production,stimulated lymphocyte proliferation and provided partial protection by reducing the P.carina burden.Conclusions:The data demonstrate that pcDNA3.1(+)-p55-582 might be potent vaccination that can afford the partial protection for the immunized animals.

A Study on Using Improved Methenamine-silver Stain to Diagnosis of Pneumocystis carinii

Lung smears of mice and lung sections of rats or human case with Pneumocystis cariniiinfection were stained using the Grocott's modification method of Gomori's methenamine-silver nitratetechnic, in which 5% sodium periodate and 5% chromic acid were used as oxidant respectively. Theoxidation time for the mouse lung smears was 5,15,60 minutes and the oxidation temperature was 20℃.The time of silver impregnation was 90 minutcs and the temperature was 60℃ for the all smearo. Whenthe oxidation time was under 15 minutes. Pneumocystis cariniic cysts showed light or dark brown, and theparenthesis-like structure could clearly be found in part of the cysts. However, if the time of oxidationWas longer, the cysts showed black and secmed to have damaged. In the same batch of the mouse lungsmears oxidated for 5 minutes, the samiples oxidated by sodium periodate showed more the cysts with theparen thesis-like structure than those oxidated by chromic acid.In the rat or patient's lung sectionsoxidated by. sodium periodate, this structure could also be found. The result of the experiment showsthat sodium periodate as an oxidant in the subsequent step of the the silver impregnation is preferable tochromic acid. And then,it is useful to clinical practice that the step of sodium bisulfate can be omittedin the study.

Analysis of 8 chronic kidney disease patients complicated with Pneumocystis carinii pneumonia

Objective To study the clinical characteristics and outcome of Pneumocystis carinii pneumonia(PCP) in patients with chronic kidney diseases.Methods Clinical data of 8 cases of chronic kidney diseases complicated with PCP(excluding renal transplant patients) were examined retrospectively.Results The most common presenting symptoms at admission were fever(100%),cough without or with a little sputum(87.5%),and exertional dyspnea(75%).Beside these,they complained of chest tightness,fatigue,sweating and chills.Six patients(75%) presented with hypoxemia were diagnosed with type 1 respiratory failure during the course of illness.The most common CT feature was bilateral patchy areas of ground-glass opacities.Five patients had peripheral blood lymphocyte count less than 1 ×109/L.Four patients had CD4 cell count less than 200/mm3.Serum LDH level was elevated in 5 patients(582±222.55).Among the 8 patients,2 patients died within 20 days of PCP diagnosis.Conclusion Pneumocystis carinii pneumonia is an opportunistic and serious complication in chronic kidney disease patients treated with immunosuppressants.The disease progression is fast and patients with respiratory failure have a high mortality rate.Early diagnosis and appropriate treatment are important for better prognosis.

Clinical Analysis of 15 Cases of Non-Hodgkin Lymphoma Complicated with Pneumocystis carinii Pneumonia Treated with R-CHOP Regimen

Objective:To investigate the clinical features of R-CHOP regimen in the treatment of non-Hodgkin^lymphoma with Pneumocystis carinii pneumonia(PCP)in order to improve the understanding of PCP and the side effects of Rituxan.Methods:A retrospective analysis of 90 patients with non-Hodgkin’s lymphoma treated with R-CHOP chemotherapy in our hospital from November 2015 to November 2020,of which 15(16.7%)patients,combined with PCP clinical data,including clinical symptoms,physical signs,chest imaging examination and treatment data were used for to analysis and summarization.Results:The clinical features of R-CHOP chemotherapy combined with PCP were fever,cough,and sputum.Some patients had fewer clinical symptoms.Common imaging manifestations were double lung membrane glass shadow,patchy shadow,and flocculent shadow.It can occur in all clinical stages,and the incidence of late stage is high,and there is no clear correlation with bone marrow suppression.Pneumocystis was found in 2 cases of sputum,and the rest of the patients were clinically diagnosed.The main therapeutic drugs are sulfamethoxazole(8/15),compound sulfamethoxazole(6/15),clindamycin(1/15,sulfa drug allergy),and adrenal cortex hormones(4/15).Fourteen cases were cured and 1 case died.Conclusion:The incidence of R-CHOP in advanced non-Hodgkin^lymphoma of PCP is high.Patients with clinical use of R-CHOP chemotherapy will encounter fever,cough,chest computed tomography(CT)film glass shadow,and diffuse patch shadow.Patients should be alert to the possibility of PCP and take sulfonamides as soon as possible for medical treatment.

Pentamidine in Pneumocystis jirovecii prophylaxis in heart transplant recipients

Despite advances in transplantation techniques and the quality of post-transplantation care, opportunistic infections remain an important cause of complications. Pneumocystis jirovecii(P. jirovecii) is an opportunistic organism, represents an important cause of infections in heart transplantation patients. Almost 2% to 10% of patients undergoing cardiac transplantation have Pneumocystis pneumonia. Prophylaxis is essential after surgery. Various prophylaxis regimes had been defined in past and have different advantages. Trimethoprim/sulfamethoxazole(TMP/SMX) has a key role in prophylaxis against P. jirovecii. Generally, although TMP/SMX is well tolerated, serious side effects have also been reported during its use. Pentamidine is an alternative prophylaxis agent when TMP/SMX cannot be tolerated by the patient. Structurally, pentamidine is an aromatic diamidine compound with antiprotozoal activity. Since it is not effectively absorbed from the gastrointestinal tract, it is frequently administered via the intravenous route. Pentamidine can alternatively be administered through inhalation at a monthly dose in heart transplant recipients. Although, the efficiency and safety of this drug is well studied in other types of solid organ transplantations, there are only few data about pentamidine usage in heart transplantation. We sought to evaluate evidence-based assessment of the use of pentamidine against P. jirovecii after heart transplantation.

艾滋病合并耶氏肺孢子菌肺炎和猴痘1例报道

猴痘病毒相关肺部感染较罕见,本文报道1例艾滋病合并耶氏肺孢子菌肺炎(pneumocystis jirovecii pneumonia,PJP)同时合并猴痘的病例。该病例系沈阳市第六人民医院2023年10月收治的40岁男性患者,入院前已确诊HIV阳性,周身皮疹伴发热2个月,干咳2周。皮疹局部可见脓性渗出及少量渗血,周围绕以红晕。CD4^(+)T淋巴细胞6.0×10^(6)/L,猴痘病毒核酸检测阳性。肺部影像学检查显示两肺呈弥漫性分布磨玻璃影,肺内见多发大小不等实性结节灶,边缘可见晕征、毛刺等。肺结节穿刺活检,组织病原微生物检测为猴痘病毒及耶氏肺孢子菌阳性。临床给予抗感染、抗真菌及复方新诺明等治疗,患者发热、呼吸困难持续加重,最终死亡。本文通过复习有关资料,深入探讨了艾滋病合并PJP和猴痘时的临床特点及肺部影像学表现,以期提升临床认识。

PCR检测大鼠卡氏肺孢子虫的研究

目的探讨PCR技术检测大鼠卡氏肺孢子虫的应用价值。方法SD大鼠和Wistar大鼠均随机分成实验组和对照组,实验组每周两次皮下注射醋酸可的松,诱导产生卡氏肺孢子虫;8week后,收集肺组织和支气管肺泡灌洗液(BALF),用PCR技术检测卡氏肺孢子虫DNA,并与Giemsa染色法比较。结果实验组两种大鼠肺组织卡氏肺孢子虫DNA阳性率分别为9643%和100%,BALF阳性率亦分别为9643%和100%,它们之间均无显著性差异(P>005);BALF的PCR阳性检出率显著高于Giemsa病原染色法,肺组织的两种方法检出率无显著性差异。结论PCR是一种检出率较高的方法,可作为早期诊断PCP的常规方法,特别适用于BALF检测卡氏肺孢子虫DNA。

PCR-ELISA检测大鼠卡氏肺孢子虫DNA的研究

目的 建立卡氏肺孢子虫 (P .c )DNA的聚合酶链反应 酶联免疫吸附测定 (PCR ELISA)方法 ,并探讨其应用价值。 方法 实验组患卡氏肺孢子虫肺炎的SD大鼠和Wistar大鼠各 2 8只 ,采用PCR法扩增大鼠肺组织DNA和支气管肺泡灌洗液 (BALF)DNA ,用PCR ELISA检测其扩增产物。 2 8只患病大鼠分别制作肺组织印片及BALF涂片 ,姬姆萨 (Giemsa)染色镜检 10 0个视野中有无P .c包囊 (或滋养体 ) ,与PCR ELISA检测扩增产物结果比较。 结果 两种方法检测大鼠肺组织DNA阳性率及BALFDNA阳性率 ,结果相同 ,均分别为 96 4% (2 7/2 8)和10 0 % (2 8/2 8)。Giemsa染色镜检P .c包囊 (或滋养体 ) ,结果为阳性的大鼠 ,PCR ELISA检测扩增产物结果也均为阳性。阴性对照组 ,两种大鼠的肺组织和BALF各 10份标本 ,均有 1只大鼠阳性。 结论 PCR ELISA检测大鼠卡氏肺孢子虫DNA ,敏感性较高 ,特异性较好 ,操作简便 ,具有实用价值。

用半巢式PCR检测无创性标本诊断卡氏肺孢子虫肺炎的实验研究

目的无创性诊断卡氏肺施子虫肺炎。方法采用一对半引物进行半巢式DHFR－PCR检测实验大鼠无创性标本——支气管液、血清及活检标本——肺及肝脏组织中的卡氏肺孢子虫DNA。结果从59只经病理学检查证实的卡氏肺孢子虫肺炎大鼠中采集到的上述4种标本，卡氏肺孢子虫DNA的半巢式DHFR－PCR检出率分别为72.7％（32／44），37.3％（22／59），94．9％（56／59）和36．4％（8／22），而DHFR－PCR的检出率则仅为25％，13．6％，71．2％和9．1％。12只轻度感染的PCP大鼠的无创性标本中卡氏肺孢子虫DNA的检出率由刚提高至41．7％。正常鼠标本及各种病原体对照的半巢式DHFR－PCR均为阴性。结论用半巢式DHFR－PCR无创性地诊断大鼠卡氏肺孢子虫肺炎具有敏感、特异、省时、省力等优点；本文在血和肝中检出卡氏肺孢子虫DNA，提示在卡氏肺孢子虫肺炎大鼠模型中存在虫血症和肺外卡氏肺孢子虫感染。

卡氏肺孢子虫PCR方法敏感性研究

目的从3种PCR方法中选择出一种检测卡氏肺孢子虫(Pc)敏感性最高的PCR方法,供以后的实验中应用。方法Wistar大鼠30只,皮下注射地塞米松,每周二次,制备PCP动物模型,8周后全部剖杀作大鼠肺印片,六亚甲基四胺银染色。各鼠取肺组织02g,均浆后提取DNA,用三种PCR方法作PCR试验。选取六亚甲基四胺银染色和三种PCR检测均为阳性的大鼠DNA标本20份,对每份标本作对倍稀释1/2、1/4、1/8、1/16……至1/2048,同时用三种PCR方法做扩增试验直到某一稀释度时PCR试验阴性为止,选择出最为敏感的一组PCR方法,并做统计学分析。结果三种方法最低稀释度的均数DHFR-PCT为1/7879±174;TS-PCR为1/4371±207;MT-PCR为1/10975±136。结论三种检测Pc的PCR方法中以MT-PCR方法最为敏感。

PCR技术对大鼠卡氏肺孢子虫DNA的检测

目的 探讨诊断卡氏肺孢子虫感染的最佳方法。方法 建立大鼠动物模型，肺涂片用吉氏染色法查肺孢子虫包囊，支气管灌洗液、肺组织和血液标本，用ＰＣＲ技术进行卡氏肺孢子虫ＤＮＡ的检测，共计实验组大鼠１２ 只及对照组８ 只。结果 实验组大鼠，吉氏染色法卡氏肺孢子虫包囊检出率为６６－７％ ，ＰＣＲ技术支气管灌洗液、肺组织和血液标本，卡氏肺孢子虫ＤＮＡ检出率分别为８１－８ ％ 、５０％ 和５０％ 。比肺印片提前２ 周检出。对照组大鼠，吉氏染色法未检出卡氏肺孢子虫包囊，在支气管灌洗液和血液标本中，ＰＣＲ技术测出了卡氏肺孢子虫ＤＮＡ。结论 在血液标本中ＰＣＲ的成功应用，为卡氏肺孢子虫感染的流行病学调查及人类卡氏肺孢子虫肺炎的诊断，提供了一种有用的、非创伤性的方法。

体外青蒿素衍生物抗卡氏肺孢子虫作用的PCR-SSCP分析

目的 研究双氢青蒿素、青蒿琥酯对卡氏肺孢子虫胸腺嘧啶合成酶基因 (TS)、二氢叶酸还原酶基因 (DHFR)的影响。方法 用不同浓度的双氢青蒿素、青蒿琥酯作用于体外培养的卡氏肺孢子虫。用药 7天后收集培养液提取卡氏肺孢子虫DNA ,分别进行PCR扩增 ,将扩增产物作DNA的单链构象多态性分析 (SSCP)。结果 卡氏肺孢子虫TS基因和DHFR基因的扩增产物分别为 40 3和 2 73bp。经青蒿素衍生物作用后TS基因、DHFR基因在溴化乙锭染色上的强度均有明显减少。继续作SSCP分析发现 ,用药组与不用药组对比DHFR基因DNA条带无明显差别 ,而用TS基因DNA条带中有一条带的位置发生改变 ,提示该基因可能出现变异。结论 SSCP分析结果提示青蒿素衍生物对卡氏肺孢子虫的DHFR基因结构无影响 ,而TS基因则可能出现部分变异。

不同分子检测技术对艾滋病患者合并寄生虫感染在真实世界的应用效果比较

目的对住院艾滋病患者进行合并寄生虫感染的初步检验,比较不同DNA检测技术对艾滋病患者合并寄生虫感染在真实世界的应用效果。方法选择临床诊断为合并弓形虫感染或肺孢子虫感染的16例住院艾滋病患者作为研究对象,以临床诊断为“金标准”进行实验室分子检测。检测方法包括针对不同靶基因的荧光定量聚合酶链反应(qPCR)和环介导等温扩增技术(LAMP);临床样本包括脑脊液、血浆和支气管肺泡灌洗液(BALF);比较采用不同方法和临床样本的检测结果。结果采用Rep-529作为靶基因,LAMP与qPCR诊断艾滋病合并弓形虫感染的总检出率均为53.12%,差异无统计学意义(P>0.05)。对脑脊液样本进行LAMP-529-11和qPCR-529检测的检出率显著高于血浆样本〔90.62%(29/32)比15.62%(5/32),P<0.01〕。采用BALF样本临床诊断合并肺孢子虫肺炎的检出率显著高于血浆样本〔100.00%(32/32)比18.75%(6/32),P<0.01〕。LAMP-18S检测的检出率显著高于LAMP-Cob检测引物组〔65.62%(21/32)比46.87%(15/32),P<0.05〕。LAMP-18S与qPCR-mtSSU的总检出率比较差异无统计学意义〔65.62%(21/32)比53.12%(17/32),P>0.05〕。结论采用Rep-529作为靶基因对艾滋病患者合并弓形虫感染进行检测,qPCR和LAMP方法均可选,建议首选样本为脑脊液;肺孢子虫肺炎检测也可选qPCR和LAMP方法,建议首选样本为BALF。当选择LAMP方法时,建议首选针对18S的引物组。

用PCR诊断不同虫荷量的大鼠卡氏肺孢子虫肺炎的价值

目的 评价在不同虫荷量条件下用 PCR诊断大鼠卡氏肺孢子虫肺炎 (PCP)的价值。 方法 将 PCP大鼠随机分为药物治疗组及未治疗组 ,测定其肺虫荷量 ,收集其支气管灌洗液 (BAL F)和血清标本 ,用 PCR和半套式 (Sn) PCR检测标本中的卡氏肺孢子虫基因 ,比较 PCR和环六亚甲基四胺银 (GMS)染色结果 ,及与病鼠肺虫荷量的关系。 结果 虫荷量高的未治疗组病鼠的 BAL F标本 PCR、Sn PCR及 GMS染色阳性率分别为 10 0 %、10 0 %和 92 .6 % ,差异无显著性 (P>0 .0 5 ) ;而虫荷量低的治疗组的 PCR(88.1% )、Sn PCR(94 .9% )阳性率均显著高于 GMS染色镜检阳性率 (30 .5 % ) (P<0 .0 5 )。肺虫荷量 >5 0 0个 /mg肺的 PCP大鼠 ,血清 PCR阳性率显著高于≤ 5 0 0者 (分别为 4 2 .9%和 1.7% )。 结论 PCR可用于虫株负荷量较低时的 PCP诊断 ,对判断 PCP病情及疗效考核亦有一定的帮助。

卡氏肺孢子虫PCR检测方法研究进展

卡氏肺孢子虫主要引起动物和人的卡氏肺孢子虫肺炎。本文就 PCR检测方法在检测卡氏肺孢子虫中的应用方面进行了综述。

PCR技术用于恶性肿瘤患者并发卡氏肺孢子虫肺炎的诊断

用PCR方法检测恶性肿瘤患者痰液及血清中的卡氏肺孢子虫。结果显承，29例术后接受化疗作有呼吸道症状的恶性肿瘤患者中，16例血清阳性，其中11例疾液阳性。14例接受化疗无呼吸道症状的恶性肿瘤患者及20例正常人其血清均为阴性。本研究表明，用PCR方法检测患者血清中的虫体DNA，安全，敏感性高，特异性强。

用PCR扩增技术检测卡氏肺孢子虫DNA

本文报道了用ＰＣＲ扩增技术检测卡氏肺孢子虫感染大鼠的肺组织、支气管灌洗液和外周血病原体ＤＮＡ，并和肺印片结果进行比较。共检测了实验感染鼠３４只及正常大鼠６只，肺印片阳性者１８例，ＰＣＲ检测肺组织样品出现特异的扩增条带者１９例，检测支气管灌洗液样品出现特异的扩增条带者１６例，外周血样品全部阴性。根据实验结果，认为此法用于检测卡氏肺孢子虫病原体ＤＮＡ有很好的应用前景。

集束化护理在艾滋病并发PCP重症患者中的应用效果

目的探讨集束化护理在艾滋病并发肺孢子菌肺炎(PCP)重症患者中的应用效果。方法采用随机对照方法,将40例AIDS合并PCP重症患者分为试验组(18例)和对照组(22例),比较两组的平均住院天数、7 d好转率及住院费用。结果试验组的平均住院天数、住院费用及7 d好转率均优于对照组,差异有统计学意义(P<0.05)。结论集束化护理能优化护理流程,提高护理人员准确执行治疗护理措施的依从性,还可以缩短患者平均住院日,降低住院费用。

Alteration of surfactant proteins A and D in bronchoalveolar lavage fluid of pneumocystis carinii pneumonia

Objective To understand the interaction between surfactant proteins and pneumocystis carinii pneumonia (PCP),and the impact of corticosteriods on surfactant proteins.Methods We established rat models of PCP and bacterial pneumonia induced by subcutaneous injection of 25mg cortisone acetate.At 8- 12 wk,the bronchoalveolar lavage fluid(BALF)of rats was collected.Total nucleated cells of BALF were counted and differentiated,and the concentrations of surfactant protein A(SP-A)and surfactant protein D(SP-D)were measured by immunoblotting assay.The rats were divided into three immunosuppressive groups and a normal control group.Group I,normal control(n = 6),consisted of healthy SD rats;group Ⅱ,negative control(n = 6),consisted of rats with cortisone acetate injection for over 8 wk without lung infection;group Ⅲ,bacterial pneumonia(n = 11),rats were injected with cortisone acetate over 8 wk that resulted in bacterial pneumonia without other pathogens isolated;and group Ⅳ,PCP(n = 14),rats with injected cortisone acetate for 8 - 12 wk and developed PCP without other pathogens isolated.Results Our results indicated that the total cell count in BALF in the negative control group was lower than that in the normal control group(P ＜ 0.001).During PCP infection,the total cell count and the percentage of polymorphonuclearcytes(PMNs)in BALF were significantly increased(P ＜ 0.01),but were lower than those in the bacterial pneumonia group.The concentration of SP-A of BALF in PCP(45.1 ± 22.1 μg/ml)was significantly increased in comparison with that in the negative control(16.2 ± 9.9 μg/ml,P ＜ 0.05)and bacterial pneumonia groups(6.2 ± 5.6 μg/ml,P ＜ 0.001).We also found that the relative content of SP-D was significantly higher in PCP(24249 ±4780 grey values)than that in the negative control (13 384 ± 2887 grey values,P ＜ 0.001)and that in bacterial pneumonia(11 989 ± 2750 grey values,P＜0.001).SP-A and SP-D were also higher in the moderate to heavy group of PCP than those seen in the mild group(P ＜ 0.01,P ＜ 0.001).SP-A and SP-D were higher in the negative control group than those in the normal control group,but there was no significant difference between the 2 groups.Conclusion These results suggest that the concentrations of SP-A and SP-D in BALF are increased by pneumocystis carinii specific stimulation,but the alteration is not related to the corticosteriod usage.

以合并肺孢子菌肺炎为例探讨新型冠状病毒感染的鉴别诊断及中西医治疗

随着疫情防控形势的调整,新型冠状病毒感染(简称新冠感染)与其他肺部感染性疾病的鉴别诊断尤显重要。肺孢子菌肺炎属真菌感染性肺炎,与新冠病毒同时感染时,其症状和肺部影像学表现常难以区分,易被误诊误治。本篇报道的2例肺孢子菌肺炎合并危重型新冠感染的艾滋病患者,在西医常规治疗的基础上,运用中医清肺解毒、益气养阴等治法,祛邪扶正,帮助减轻患者的症状,延缓病情的进展,促进核酸早日转阴。