旋毛虫肌幼虫排泄分泌抗原诊断早期旋毛虫病的研究

目的评价旋毛虫肌幼虫排泄分泌(excretory-secretory,ES)抗原诊断早期旋毛虫病的价值。方法应用旋毛虫肌幼虫ES抗原Western blot对旋毛虫感染后6~11d的小鼠血清及19d的早期旋毛虫病人血清进行检测,并与感染后35d的晚期旋毛虫病人和其他寄生虫病人血清的检测结果进行比较。结果 Western blot分析显示,肌幼虫ES蛋白中的2条蛋白带(41.5、55kDa)可被旋毛虫感染后7~11d的小鼠血清识别,6条蛋白带(41.5、44.1、45、55、61和65.2kDa)能被早期和晚期旋毛虫病人血清识别,阳性反应率均为100%(15/15);这6条蛋白带与裂头蚴病人和健康人血清无交叉反应,但与其他寄生虫病(血吸虫病、并殖吸虫病、华支睾吸虫病、棘球蚴病及囊尾蚴病)患者血清具有明显的交叉反应(19.12%~38.23%)。结论旋毛虫肌幼虫ES抗原中的41.5kDa~65.2kDa蛋白带可与旋毛虫感染早期血清反应,但与其他蠕虫病患者有明显的交叉反应。     

Troponin dosage in a patient with asymptomatic myocarditis due to trichinellosis

Human trichinellosis is a potentially severe parasitic disease occurring after ingestion of undercooked meat infected with Trichinella sp. larvae. We report the case of a patient who ate an undercooked bear meat hunted in Canada; he presented with the usual symptoms of trichinellosis (i.e, facial oedema, myalgias and fever) complicated with an asymptomatic myocarditis. Myocarditis is a rare, but potentially lethal complication of trichinellosis. Myocarditis should be screened systematically even when specific symptoms are missing; dosage of troponin serum is a simple and reliable mean for such screening.     

A major trichinellosis outbreak suggesting a high endemicity of Trichinella infection in northern Laos

Trichinellosis is an important and under-recognized food-borne zoonosis in Southeast Asia. After 30 years of no reports, a small outbreak was described in Central Lao PDR (Laos) in 2003. Here we report a large outbreak of at least 650 estimated patients in Udomxay (northern Laos) in June 2005. Trichinella ELISA assays on serum from 133 patients and Western blot assays on 16 patients were positive in 67.6% and 81.2%, respectively. No deaths were recorded. Consumption of uncooked or fermented pork at funeral and wedding ceremonies was the main source of infection. Larvae of Trichinella spiralis were found in 1 of 11 local pigs not involved in this outbreak. The results suggest that trichinellosis may be an under-recognized but important endemic disease in Laos and reinforces the need to urgently implement veterinary and educational programs.     

旋毛虫肌幼虫p45cDNA的克隆及鉴定

目的本研究根据GenBank中旋毛虫45kDa抗原基因的序列设计引物,采用PCR技术从旋毛虫肌幼虫cDNA文库中扩增靶基因p45cDNA,并克隆到pMD-18T载体,转化至大肠埃希氏菌NovaBlue后测序分析。获得827bp的p45cD-NA。该序列与GenBank中的旋毛虫p45基因序列相比共有1个核苷酸发生改变,二者的同源性为99%,其编码蛋白质由268个氨基酸组成,与45kDa抗原的同源性为99%。将p45cDNA克隆到原核表达载体pET28a并转化至表达菌BL21star（DE3）后,经IPTG诱导表达出约30kDa的重组蛋白。Western-blot检测表明,p45重组蛋白可以被旋毛虫感染猪血清识别,具有良好的抗原性。     

抗旋毛虫肌肉期幼虫排泄分泌物中46-58kDa蛋白IgM和IgG抗体的研究

目的寻求简便、快速、可靠的方法,用于旋毛虫病的诊断及疗效考核。方法建立检测抗旋毛虫肌肉期幼虫排泄分泌物(TsL1-ES)中46-58kDa蛋白抗体的斑点金免疫渗滤试验(DIGFA),并对患者在治疗前和治疗后血清中特异抗体水平的动态变化进行研究。结果旋毛虫病患者治疗前血清中特异IgM和IgG抗体的检出率分别为94.83%(55/58)和91.38%(53/58);正常人血清(100份)、囊虫病(25份)、血吸虫病(20份)、肺吸虫病(20份)及蛔虫病(22份)病人血清均未检出该两类特异性抗体。52例两类抗体均阳性的旋毛虫病患者经有效药物治疗后3个月、6个月、1年、1.5年连续采血,经DIGFA检测连续观察的结果表明,IgM抗体出现较早,阴转较为明显和迅速,治疗后3月的阴转率达46.15%(24/52),1年的阴转率可达88.46%(46/52);而IgG抗体则出现较晚,阴转缓慢,治疗后1.5年的阴转率仅为23.08%(12/52)。结论DIGFA为检测抗TsL1-ES中46-58kDa蛋白抗体的有效方法,检测IgM抗体可用于诊断及疗效考核,而IgG抗体的检测不宜用来判断疗效。     

Imported trichinellosis with severe myositis--report of a case

A 38-year-old Japanese male who had traveled in China from September 13 to October 5, 1997, developed fever and severe conjunctivitis from October 20. After he was hospitalized in Kyoto City Hospital for persistent high fever on October 29, he developed muscular weakness and dysphagia which continued for two weeks. An electromyogram showed a myogenic pattern, and laboratory findings showed significant elevation of serum enzyme levels of muscle origin: CPK, 3,095 IU/l; aldorase, 195 IU/l; myoglobin, 7,570 ng/ml, and myoglobinuria, 94,700 ng/ml. The WBC was 10,800/microliter with 45% eosinophils. Muscular biopsy showed degeneration of muscle fibers with infiltration of macrophages and lymphocytes. On further inquiry, it was revealed that the patient had eaten smoked bear meat in China on September 30, three weeks prior to the onset of symptoms. A dot-ELISA serologic test for parasites was positive for Trichinella. Further, a coiled 1.2 mm long Trichinella larve was recovered from approximately 100 mg of frozen biopsied muscle by an enzyme digestion method. Mebendazole was given to the patient at a dosage of 200 mg/day for seven days. CPK levels were normalized within 3 days of the beginning of the treatment, and he was discharged without any symptoms. Physicians must be aware of trichinellosis and should include it in their differential diagnosis when examining patients with myositis and eosinophilia of unknown origin.     

Establishment and characterization of a cholangiocarcinoma cell line (RMCCA-1) from a Thai patient

AIM: To establish and characterize a new cell line derived from peripheral cholangiocarcinoma of a Thai patient.METHODS: The peripheral cholangiocarcinoma specimen surgically obtained from the patient was aseptically processed by washing and mincing before culturing in Ham's F12 medium containing 10% fetal bovine serum. After 3 mo, when the cell line has become homogeneous and stabilized, several features were investigated, including growth characteristics,immunofluorescence staining for cytokeratins, expression of tumor markers, chromosomal analysis by G-banding and multicolour fluorescence in situ hybridization (mFISH), in vitro migration and invasion characteristics.RESULTS: The RMCCA-1 cell line has been established.These cells proliferated as a monolayer with a population doubling time of 48 h. Immunofluorescence staining showed positive staining for human cytokeratin 7 and 19 verifying the biliary epithelial origin. RMCCA-1secreted carbohydrate antigen 19-9 (CA19-9), but insignificant levels of carcinoembryonic antigen (CEA)and α-fetoprotein (AFP). Chromosome analysis identified aneuploidy karyotypes with a modal chromosome number of 59. RMCCA-1 exhibited a low level of in vitro invasiveness, but a high degree of motility. The cell line exhibited a significant number of chromosomal aberrations as shown by mFISH and G-banding methods.CONCLUSION: A new cell line derived from peripheral cholangiocarcinoma of a Thai patient has been established. This cell line shows a low level of in vitro invasiveness, but a high degree of motility. It will serve as a valuable tool for further studies on tumor biology,molecular pathogenesis, metastatic mechanism and response to therapeutic drugs of cholangiocarcinoma.     

Establishment and characterization of a cholangiocarcinoma cell line （RMCCA-1） from a Thai patient

INTRODUCTIONCholangiocarcinoma is a highly malignant epithelial neoplasm that arises within the intrahepatic and extrahepatic biliary tract[1].The pathogenesis of this disease has been strongly associated with chronic inflammation and cellular injury with…     

An outbreak of trichinellosis due to consumption of bear meat infected with Trichinella nativa,in 2 northern Saskatchewan communities

In June 2000, bear meat infected with Trichinella nativa was consumed by 78 individuals in 2 northern Saskatchewan communities. Interviews and blood collections were performed on exposed individuals at the onset of the outbreak and 7 weeks later. All exposed individuals were treated with mebendazole or albendazole, and symptomatic patients received prednisone. Confirmed cases were more likely to have consumed dried meat, rather than boiled meat (P<.001). Seventy-four percent of patients completed the recommended therapy, and 87% of patients who were followed up in August 2000 reported complete resolution of symptoms. This outbreak of trichinellosis was caused by consumption of inadequately cooked bear meat contaminated with T. nativa. Apart from clinical symptomatology, blood counts, creatine kinase levels, serology test results, and analysis of the remaining bear meat helped establish the diagnosis. Treatment with antiparasitic drugs and prednisone was beneficial in limiting the severity and duration of the illness.     

Human trichinellosis due to Trichinella britovi in southern France after consumption of frozen wild boar meat

Six patients were infected with Trichinella britovi in southern France following consumption of frozen wild boar meat, which had been frozen at -35 degrees C for 7 days. Microscopic examination of a sample of frozen wild boar muscle revealed the presence of rare encapsulated Trichinella larvae, identified as T. britovi. People eating wild boar must follow individual prophylactic rules such as efficient cooking of meat (at least 65 degrees C at the core for 1 minute) as recommended by the International Commission on Trichinellosis, or freezing exceeding four weeks at -20 degrees C.     

成人重症肺炎患者EB病毒和人鼻病毒混合感染的病原学分子鉴定

目的对1例有禽类接触史的成人重症肺炎患者进行病原学研究,确定其病原体。方法采集患者的下呼吸道标本,应用荧光PCR方法检测禽流感H5N1和SARS病毒及甲型H1N1和季节性流感病毒;应用多重PCR方法检测12种常见呼吸道病毒;应用普通PCR方法检测EB病毒和巨细胞病毒,扩增阳性的病毒基因片段测序后,与GenBank数据库参考序列进行同源性分析。结果荧光PCR法检测禽流感病毒H5N1病毒和SARS病毒核酸阴性,甲型H1N1和季节性流感病毒核酸阴性;多重PCR检测人鼻病毒、EB病毒均阳性;普通PCR检测EB病毒核酸阳性。基因序列比对显示,EB病毒核苷酸序列与GenBank数据库中参考序列同源性为99.5%～100%;人鼻病毒的核苷酸序列与GenBank数据库中A组鼻病毒(73血清型)同源性最高,为94.7%。结论该例重症肺炎患者为EB病毒和A组(73血清型)人鼻病毒混合感染,排除禽流感H5N1和SARS感染。     

A SPINK1 gene mutation in a Thai patient with fibrocalculous pancreatic diabetes

Fibrocalculous pancreatitis diabetes (FCPD), a late stage of tropical chronic pancreatitis (TCP), is classified as a secondary cause of diabetes mellitus resulting from pancreatic exocrine dysfunction. The distinctive features of FCPD and TCP are young age at onset, presence of large intraductal pancreatic calculi, and reported mainly in tropical developing countries. Their etiology is still obscure, but the autodigestion due to aberrant intraductal activation of zymogens by trypsin is thought to be a primary common event. Recently, mutations in SPINKI gene encoding a pancreatic secretory trypsin inhibitor have been reported in association with an increased risk of pancreatitis. We describe a heterozygous mutation, IVS3+2 T>C, of SPINK1 gene in a young Thai female patient with typical presentation of FCPD. To our knowledge, this is the first report of the SPINK1 gene mutation in a FCPD patient in Southeast Asia.     

旋毛虫亲肌肉抗原的基因克隆和序列分析

目的克隆一个新的旋毛虫基因即亲肌肉抗原(myophilin)并分析其序列同源性,为旋毛虫病疫苗的研制奠定基础。方法检索GenBank旋毛虫基因组数据库,获得旋毛虫亲肌肉抗原的cDNA序列,利用Primer5.0软件设计引物(上、下游引物分别含NdeⅠ和XhoⅠ酶切位点)。收集云南株旋毛虫肌幼虫,提取总RNA,并以此为模板进行RTPCR。将PCR产物进行2%琼脂糖凝胶电泳,目的基因切胶回收后与克隆载体pMD-19T分别用NdeⅠ、XhoⅠ双酶切,酶切产物以3︰1的比例与pMD-19T连接,然后转化到DH5a大肠埃希菌感受态细胞中,用Amp抗性培养基培养,阳性克隆提质粒后做PCR及双酶切鉴定,阳性菌液进行测序,测序结果与检索基因核苷酸序列通过DNAMAN软件比较,分析其同源性。结果 RT-PCR扩增出旋毛虫亲肌肉抗原全长基因序列,大小为579bp;亲肌肉抗原基因重组质粒经双酶切得到的片段大小分别为579、2 300和300bp,与预期值相符;该基因序列与检索基因核苷酸序列相比,同源性为100%。结论成功克隆出旋毛虫亲肌肉抗原基因,为旋毛虫病疫苗的研制奠定了基础。     

Molecular identification of lice from pre-Columbian mummies

BACKGROUND: Three distinctly different lineages of head and body lice are known to parasitize humans. One lineage includes head and body lice and is currently worldwide in distribution (type A). The other 2 (types B and C) include only head lice and are geographically restricted. It was hypothesized that head louse phylotypes were exchanged only recently, after European exploration and colonization (after Columbus). METHODS: To determine which louse type or types were found in the Americas before European colonization, we used polymerase chain reaction in 2 laboratories to amplify DNA from 2 genes (Cytb and Cox1) belonging to 1000-year-old lice collected from Peruvian mummies. RESULTS: Only the worldwide type (type A) was found. Therefore, this phylotype was worldwide before European colonization, as type A lice were common in Europe, Africa, and Asia. CONCLUSIONS: The findings of this study show that several phylotypes of head lice have coexisted for centuries in humans and support the claim that type A lice were present in the Americas before the time of Columbus.