台湾蠛蠓rDNA-ITS序列测定与分析

目的 探讨rDNA-ITS基因序列在吸血蠓分子鉴定中的应用前景。方法 通过PCR扩增获取台湾蠛蠓rDNA-ITS基因,测定分析ITS1和ITS2基因序列,构建系统发育树。结果 台湾蠛蠓ITS1和ITS2片段长度分别为310 bp与360 bp,与汤斯维尔铗蠓的同源性最大,分别为71%和92%。系统发育分析显示,基于ITS1、ITS2基因序列的分子鉴定结果与传统形态学鉴定结果一致。结论 rDNA-ITS基因序列可作为蠓科昆虫分子鉴定的分子标记。     

基于线粒体COI基因序列进行库蠓二囊亚属近似种的分子鉴定

目的 通过采用线粒体COI基因进行库蠓近似种分子鉴定的方法,来弥补传统形态学在种类鉴定中存在的操作繁琐、难度大等不足,从而实现库蠓近似种的快速、准确鉴别。方法 采用DNA测序的方法获得库蠓属二囊亚属的林岛库蠓(C. gaponus)、标翅库蠓(C. insignipennis)、无害库蠓(C. innoxius)、连斑库蠓(C. jacobsoni)、南山库蠓(C. lansangensis)、新竹库蠓(C. liui)、长喙库蠓(C. longirostris)、异域库蠓(C. peregrinus)等8种库蠓近似种的部分线粒体COI序列,并对其进行分子鉴定;基于Kimura 2-parameter模型分析遗传距离,同时应用MEGA 6.0软件以荒川库蠓C. arakawai为外群构建系统发育树。结果 8种库蠓COI序列长度约650 bp;遗传距离在种内和种间具有统计学差异(t=119.452,P<0.05);系统发育树中不同库蠓种类各自构成单系(群),同种类不同地理种群聚为一支。结论 本研究证实了线粒体COI序列可用于进行库蠓近似种的分子鉴定。     

库蠓核糖体DNA内转录间隔区1序列的测定与分析

目的 测定核糖体DNA内转录间隔区1(nuclear ribosomal DNA internal transcribed spacer 1, rDNA-ITS1)序列进行库蠓种类鉴别与系统发育分析,以探究rDNA-ITS1序列在库蠓分子鉴定和系统发育研究方面的适用性。方法 采用DNA扩增、纯化、克隆及测序的方法获得荒川库蠓Culicoides arakawai、凹缘库蠓C. holcus、连斑库蠓C. jacobsoni、印度库蠓C. indianus、霍飞库蠓C. huffi和肩宏库蠓C. humeralis等6种库蠓的rDNA-ITS1序列,基于Kimura 2-parameter公式计算种内和种间遗传距离,应用MEGA 6.06软件分析DNA序列碱基组成并以环纹埃蠓Allohelea annulata为外群构建系统发育树(邻接法和最大似然法)。结果 上述6种库蠓的rDNA-ITS1序列经过Clustal W比对及人工校对和编辑后的长度为352 bp,其中T、C、A、G 4种碱基的含量分别为36.8%、13.0%、30.0%和20.1%,A+T的含量(66.8%)高于C+G的含量(33.1%);K-2p遗传距离在种内和种间具显著差异(t=32.430,P<0.05);系统发育树中不同种类库蠓各自构成单系(群),同种类不同地理种群聚为一支,其与形态学鉴定结果一致。结论 本研究证实了rDNA-ITS1序列可用于进行库蠓及其近似种的分子鉴定和系统发育分析。     

云南省狂犬病病人和犬类感染狂犬病病毒及M基因序列分析

目的 调查云南省部分地区狂犬病病人和疫点犬群携带狂犬病病毒(rabies virus, RABV)状况及RABV膜基质蛋白(matrixprotein, M)基因序列分析,为狂犬病防控提供科学依据。方法 2008-2009年在云南省采集犬脑组织标本606份,狂犬病病人唾液8份,脑脊液1份,用直接免疫荧光试验检测RABV抗原,用RT-PCR检测RABV核酸,对阳性标本进行M基因序列测定和分析。结果 所有标本经检测,RABV抗原和/或核酸阳性16份,其中疫点扑杀的貌似健康犬脑组织3.10%(10/323),狂犬病病人唾液5份、脑脊液1份。狗肉餐馆屠宰犬的脑组织283份均为阴性。云南16株RABV的M基因核苷酸和氨基酸同源性分别为88.5%~100%和85.2%~99.5%。它们与中国人用疫苗株aG核苷酸和氨基酸同源性分别为83.9%~85.7%和82.3%~93.6%;与人用疫苗株CTN181核苷酸和氨基酸同源性分别为99%~99.7%和98.5%~99%。系统进化分析表明,云南16株RABV均属基因Ⅰ型并可分为进化Ⅰ和Ⅱ群并分别与泰国等东南亚国家和相邻省份流行株具有较近的亲缘关系。结论 云南省狂犬病流行与周边省份和东南亚地区的狂犬病传播扩散有一定关系;狂犬病疫点部分貌似健康犬携带RABV并具有传染源意义;云南狂犬病病毒株M基因与我国人用狂犬病疫苗CTN株的同源性和亲缘关系较近,但与aG株同源性存在。     

应用多重PCR法分析西藏察隅疟疾流行区按蚊吸血习性

目的用多重PCR法分析西藏疟疾流行区察隅县常见按蚊的吸血习性,为下一步研究传疟媒介提供参考。方法2011年7~8月选择察隅县不同生态环境的3个自然村(日玛村、塔玛村和京都村),每个村在人房和畜舍选择8个点,采用诱蚊灯全通宵(20∶00至次日08∶00)诱捕法捕捉按蚊,次日清晨收集诱捕的蚊虫,经形态学鉴定蚊种,分析按蚊组成。收集饱血按蚊,分别提取单只蚊胃血的DNA,采用基于不同动物mtDNA-cytb序列差异的多重PCR法鉴定各蚊胃血源,计算人血指数,分析按蚊的吸血习性。结果共捕获按蚊1 442只,经形态学鉴定,多斑按蚊种团占99.6%(1 436/1 442),带足按蚊和腹簇按蚊占0.4%(6/1442)。多斑按蚊种团中,伪威氏按蚊占85.5%(1228/1436),威氏按蚊占14.5%(208/1 436)。用多重PCR检测202只多斑按蚊种团(伪威氏按蚊188只和威氏按蚊14只)的饱血蚊胃血,结果显示,伪威氏按蚊兼吸牛/猪血和人血,人血指数为0.35,威氏按蚊吸食猪血和人血,人血指数为0.29。结论西藏察隅县2种常见按蚊(伪威氏按蚊和威氏按蚊)均兼吸人畜血,伪威氏按蚊的人血指数较高。     

肝母细胞瘤儿童生存状况调查

目的 探究肝母细胞瘤（HB）儿童的生存状况。方法 2011年3月~2015年3月收治的34例HB患儿,采用外科根治手术与辅助化疗联合治疗方案治疗,随访截止时间为2017年1月。结果 本组病理分型主要为胎儿型,肿瘤分期主要为Ⅰ期;随访结束时,8例（23.5%）死亡,平均生存时间为（10.23±2.34）个月;初治9~18个月为死亡集中期;1例患儿血清AFP水平未升高,33例（97.1%）均升高;胎儿型患儿预后较其它病理类型患儿好;Ⅰ/Ⅱ期患儿预后较Ⅲ/Ⅳ期患儿好;CRP升高（大于5mg/L）患儿病死率较正常患儿高,LDH水平升高（大于245U/L）的患儿病死率较正常患儿高（P<0.05）。结论 血清AFP水平、肿瘤病理分型和肿瘤分期对患儿预后具有重要意义。     

乙型肝炎肝硬化患者血清IGF-Ⅰ水平与肠道屏障功能损害的关系分析

目的 研究乙型肝炎肝硬化患者血清胰岛素样生长因子Ⅰ（IGF-Ⅰ）水平与肠道屏障功能损害的关系。方法 2015年1月~2017年1月我院诊治的乙型肝炎肝硬化患者107例,其中Child-Pugh A级35例,Child-Pugh B级42例, Child-Pugh C级30例,另选同期健康志愿者40例。采用酶联免疫吸附法检测血清IGF-Ⅰ水平,采用化学比色法检测血清二胺氧化酶（DAO）,采用改良酶学分光光度法检测血清D-乳酸,采用显色基质鲎实验法检测血清内毒素,采用高压液相色谱分析法检测尿液乳果糖/甘露醇（L/M）排出比。结果 健康人血清IGF-Ⅰ水平为（217.1±40.2） ng/ml,显著高于Child-Pugh A级肝硬化患者的（180.3±33.4） ng/ml、B级的（152.4±26.5） ng/ml或C级的（126.9±21.8） ng/ml（P<0.05）;健康人血清DAO、D-乳酸、内毒素和尿液L/M比值分别为（2.5±0.5） U/ml、（7.1±1.3）μg/ml、（0.4±0.1） EU/ml和（6.3±1.1）%,显著低于Child-Pugh A级肝硬化患者的（3.3±0.7） U/ml、（9.0±1.6） μg/ml、（0.6±0.1） EU/ml和（9.2±1.7）%或B级的（4.6±0.9） U/ml、（11.2±1.9） μg/ml、（0.8±0.1） EU/ml和（12.6±2.3）%或C级的（5.8±1.0） U/ml、（13.4±2.4） μg/ml、（1.1±0.2） EU/ml和（15.7±2.8）%（P<0.05）;乙型肝炎肝硬化患者血清IGF-Ⅰ与DAO、D-乳酸、内毒素和尿液L/M比值均呈负相关（r=-0.845、r=-0.808、r=-0.867、r=-0.839,P<0.01）。结论 乙型肝炎肝硬化患者存在肠道屏障功能损害,检测血清IGF-Ⅰ水平有一定的判断意义。     

CT检查在原发性肝癌患者经肝动脉化疗栓塞术后疗效评估中的应用价值分析

目的 探讨电子计算机断层扫描（CT）在原发性癌（PLC）患者经肝动脉化疗栓塞术（TACE）治疗后疗效评估中的应用价值。 方法 2015年10月~2016年12月行TACE治疗的92例PLC患者,在TACE术前后进行CT平扫及增强扫描,比较肝内病灶密度、强化方式、边界及大小变化,探讨CT检查对术后疗效评估的价值。 结果 在73例肿块型PLC患者,Ⅰ型碘油沉积22例（30.1%）,Ⅱ型40例（54.7%）,Ⅲ型8例（10.9%）,Ⅳ型3例（4.1%）,在19例结节型PLC患者,Ⅰ型碘油沉积14例（73.6%）,Ⅱ型4例（21.0%）,Ⅲ型1例（5.2%）;肿块型PLC患者TACE治疗后总有效率为95.5%（69/73）,结节型为94.7%（18/19）,两组差异无统计学意义（P>0.05）。 结论 CT检查能有效评估PLC患者TACE治疗后的临床疗效,为临床制定进一步的治疗方案提供确切的影像学资料支持。     

儿童肝母细胞瘤CT影像特征分析

目的 总结肝母细胞瘤（HB）儿童特征和腹部CT的表现。方法 2014年1月~2016年11月我院诊治的HB患儿94例,对全部患儿进行腹部CT扫描,获得影像学资料。结果 本组HB患儿Ⅰ期5例,Ⅱ期 44例,Ⅲ期 38例,Ⅳ期7例;肿瘤体积为29.1~3332.5 cm³,其中Ⅰ期大约为199.9±50.2cm³,Ⅱ期 为429.9±89.4 cm³,Ⅲ期为 419.2±112.3cm³,Ⅳ期为1121.0 ±213.5 cm³;肿瘤位于肝右叶45例（47.8%）、左右叶20例（21.3%）、左叶21例（22.3%）和尾叶8例（8.5%）,肿瘤与肝脏分界清楚73例（77.7%）、分界模糊21例（22.3%）,单发肿瘤81例（86.2%）、多发肿瘤13例（13.8%）,影像学表现为点片状50例（53.2%）、团块状33例（35.1%）和多种形态混合状11例（11.7%）;本组发生肿瘤远处转移38例（40.4%）,其中肺转移18例（47.4%）、骨转移12例（31.6%）、胸膜转移4例（10.5%）和淋巴结转移4例（10.5%）。结论 对HB患儿进行腹部CT检查可了解肿瘤的基本特征,早期发现肿瘤转移,有助于帮助决定选择手术方案。     

Short-horned flies (Diptera: Brachycera) are ectoparasites of warm-blooded animals

In the Orenburg Region, the fauna of blood-sucking short-horned flies comprises 23 species, including 12 obligate hematophagi and 11 facultative ones. The collected flies belong to the families Tabanidae and Muscidae. Dominant species, their spread, environmental characteristics, and phenological features have been established. The ratio of various types of blood-sucking flies has been analyzed.     

Investigations on a Plasminogen Activator in Two Blood-Suckers, Rhodnius prolixus Stål and Hirudo medicinalis

I n a previous paper (Hellmann and Hawkins, 1964) we described two anticoagulants and a fibrinolytic activity extractable from an insect, the blood-sucking Hemipteran Rhodnius prolixus . In the present paper we report further experiments to characterize the fibrinolysin and to trace its source. The properties of the two anticoagulants have been described more fully elsewhere (Hellmann and Hawkins, 1965). The fibrinolysin was originally found in the gut of the insect; of four possible sources from which it could reach the gut, the gut wall and the salivary glands have already been shown to be devoid of activity (Hellmann and Hawkins, 1964). The two remaining possibilities, the blood meal and organisms present in the gut, are examined below. The possibility that the fibrinolysin was an exo-product of the symbiotic organism Nocardia rhodnii , which occurs in the gut of R. prolixus (Baines, 1956) was strengthened by the lack of activity found in insects from which this organism had disappeared. By culturing N. rhodnii on artificial media it was possible to examine this idea directly.
Our original assumption that a fibrinolysin might be involved in dealing with the meal taken in by a blood-sucking animal proved well founded in the case of R. prolixus (Hellmann and Hawkins, 1964) and we therefore extended out investigation to the leech Hirudo medicinalis since, if the assumption was generally correct, as one of the larger blood-sucking animals, it might be expected to be a potentially better source of fibrinolysin.     

C2株蓝氏贾第鞭毛虫ILP基因的克隆表达及蛋白结构分析

目的 利用大肠杆菌对C2株蓝氏贾第鞭毛虫ILP蛋白(Impact-like protein)进行克隆表达,运用生物信息学软件进行蛋白结构分析。方法 提取C2株蓝氏贾第鞭毛虫基因组DNA,PCR扩增ILP基因,构建pGM-T重组载体,挑选阳性克隆并进行序列分析;将ILP基因连入原核表达载体pET-28a(+),并转化大肠杆菌Rosetta(DE3),IPTG诱导表达。运用PSIPRED和SWISS-MODEL进行蛋白结构分析。结果 成功构建了原核表达载体pET-28a(+)-ILP,该基因全长831 bp。SDS-PAGE结果显示,目的蛋白条带出现在相对分子量约33 kD的位置,与预期相符。Western blot结果表明,大肠杆菌成功表达了重组蛋白。结论 成功克隆、表达并分析C2株蓝氏贾第鞭毛虫ILP蛋白,为蓝氏贾第鞭毛虫ILP蛋白结构与功能的研究提供了有价值的资料。     

吸血蠓防治研究进展(双翅目:蠓科)

吸血蠓(Blood-sucking midges)是世界上最为微小的有害昆虫类群之一,其不仅刺叮骚扰人畜,还是蓝舌病、赤羽病、施马伦贝格病、土拉弗氏菌病等多种人兽共患病的传播媒介。控制和消灭传播媒介是预防和减缓蠓传疾病暴发流行的有效途径。但吸血蠓体小、分布广、扩散性强、孳生地类型复杂,一般的防治手段很难对其进行有效防治,选择科学、合理、安全的防治措施将有助于提高防治效果。本文综述了近几十年来国内外对吸血蠓的防治对策和经验,为日后开展蠓的防治工作提供理论参考。     

Effect of Gongronema latifolium on gastric emptying in healthy dogs

AIM:To investigate sonographically the effect of Gonogronema latifolium (G.latifolium) on gastric emptying of semi-solid meals in healthy dogs.METHODS:In a randomized,placebo-controlled experiment,twenty-five clinically healthy dogs were randomly allotted into five groups of five dogs in each group.The placebo group served as the control,and the low,moderate and high dose groups ingested the methanolic leaf extract of G.latifolium in capsules at 100 mg/kg,250 mg/kg and 500 mg/kg,respectively,while the prokinetic group ingested 0.5 mg/kg capsules of metoclopramide.After a 12-h fast,each group ingested its treatment capsules 30 min before the administration of a test meal.Measurements of gastric emptying and blood glucoselevels were obtained 30 min before and immediately after the ingestion of the test meal and thereafter every 15 min for 4 h.This was followed by further measurements every 30 min for another 2 h.RESULTS:The gastric emptying times of the placebo,low dose,moderate dose,high dose and prokinetic dose groups were 127.0 ± 8.2 min,135.5 ± 3.7 min,155.5 ± 3.9 min,198.0 ± 5.3 min and 59.0 ± 2.5 min,respectively.Gastric emptying times of the moderate and high dose groups were significantly slower than in the placebo control group (155.5 ± 3.9 min,198.0 ± 5.3 min vs 127.0 ± 8.2 min,P=0.000).No significant difference in gastric emptying between the low dose and placebo control groups was noted (135.5 ± 3.7 min vs 127.0 ± 8.2 min,P=0.072).Gastric emptying of the prokinetic group was significantly faster than that of the control group (59.0 ± 2.5 min vs 127.0 ± 8.2 min,P=0.000).The hypoglycaemic effect of G.latifolium and gastric emptying were inversely related (r=-0.95,P=0.000).CONCLUSION:G.latifolium delays gastric emptying and lowers postprandial blood glucose in healthy dogs.It reduces the postprandial blood glucose by delaying gastric emptying.     

A valid, accurate, office based non-radioactive test for gastric emptying of solids

BACKGROUND: Current breath tests for measurement of gastric emptying of solids are expensive, possibly inaccurate, and require cumbersome calculations. AIMS: We wished to validate a simplified solid gastric emptying test using a [(13)C]Spirulina platensis breath test for accurate results relative to scintigraphy. SUBJECTS: Thirty healthy volunteers. METHODS: We measured gastric emptying of egg containing [(13)C]S platensis and (99m)Tc sulphur colloid by breath (13)CO(2) and scintigraphy over six hours. A generalised linear regression model was used to predict t(1/2) and t(LAG) by scintigraphy from breath (13)CO(2) data. The model was cross validated and normative data calculated for a prepacked [(13)C]meal. RESULTS: Regression models using all breath data over six hours, for the first three hours, and for samples at 75, 90, and 180 minutes ("reduced model") predicted t(1/2) and t(LAG) values similar to scintigraphy (t(LAG) 43 (SD 12) min; t(1/2) 100 (20) min). Standard deviations of differences in t(1/2) and t(LAG) between scintigraphy and the "reduced model" were both 10 minutes. Gastric t(1/2) for the prepacked [(13)C]meal was 91 (15) min (10-90% range: 74-118). CONCLUSION: The [(13)C]S platensis breath test and a simple formula using breath (13)CO(2) at baseline, 90, and 180 minutes measured gastric emptying t(1/2) for solids with results that were comparable with scintigraphy.     

Prospects for fungus identification using CO1 DNA barcodes, with Penicillium as a test case

DNA barcoding systems employ a short, standardized gene region to identify species. A 648-bp segment of mitochondrial cytochrome c oxidase 1 (CO1) is the core barcode region for animals, but its utility has not been tested in fungi. This study began with an examination of patterns of sequence divergences in this gene region for 38 fungal taxa with full CO1 sequences. Because these results suggested that CO1 could be effective in species recognition, we designed primers for a 545-bp fragment of CO1 and generated sequences for multiple strains from 58 species of Penicillium subgenus Penicillium and 12 allied species. Despite the frequent literature reports of introns in fungal mitochondrial genomes, we detected introns in only 2 of 370 Penicillium strains. Representatives from 38 of 58 species formed cohesive assemblages with distinct CO1 sequences, and all cases of sequence sharing involved known species complexes. CO1 sequence divergences averaged 0.06% within species, less than for internal transcribed spacer nrDNA or beta-tubulin sequences (BenA). CO1 divergences between species averaged 5.6%, comparable to internal transcribed spacer, but less than values for BenA (14.4%). Although the latter gene delivered higher taxonomic resolution, the amplification and alignment of CO1 was simpler. The development of a barcoding system for fungi that shares a common gene target with other kingdoms would be a significant advance.     

Infection,Dissemination, and Transmission Potential of North American Culex quinquefasciatus,Culex tarsalis,and Culicoides sonorensis for Oropouche Virus

Oropouche virus (OROV), a vector-borne Orthobunyavirus circulating in South and Central America, causes a febrile illness with high rates of morbidity but with no documented fatalities. Oropouche virus is transmitted by numerous vectors, including multiple genera of mosquitoes and Culicoides biting midges in South America. This study investigated the vector competence of three North American vectors, Culex tarsalis, Culex quinquefasciatus, and Culicoides sonorensis, for OROV. Cohorts of each species were fed an infectious blood meal containing 6.5 log₁₀ PFU/mL OROV and incubated for 10 or 14 days. Culex tarsalis demonstrated infection (3.13%) but not dissemination or transmission potential at 10 days post infection (DPI). At 10 and 14 DPI, Cx. quinquefasciatus demonstrated 9.71% and 19.3% infection, 2.91% and 1.23% dissemination, and 0.97% and 0.82% transmission potential, respectively. Culicoides sonorensis demonstrated 86.63% infection, 83.14% dissemination, and 19.77% transmission potential at 14 DPI. Based on these data, Cx. tarsalis is unlikely to be a competent vector for OROV. Culex quinquefasciatus demonstrated infection, dissemination, and transmission potential, although at relatively low rates. Culicoides sonorensis demonstrated high infection and dissemination but may have a salivary gland barrier to the virus. These data have implications for the spread of OROV in the event of a North American introduction.