First report on natural Leishmania infection of Phlebotomus sergenti due Leishmania tropica by high resolution melting curve method in Southeastern Iran

Objective:To identify the Leishmania species in infected sand flies by Real-time PCR coupled with HRM analysis.Methods:Real-time PCR coupled with HRM analysis targeting the first internal transcribed spacer(ITS1)of nuclear ribosomal DNA as the genetic marker was used to identify and distinguish Lelthmania species in sand flies specimens.Results:Three out of 115females of Phlebotomus sergenti(P.sergenti)(2.6%)were positive to Leishmania tropica(L.tropica).Conclusions:This is the first report on P.sergenti as the main and proven vector of anthroponitic cutaneous leishmaniasis in Dehbakri County using Real-time PCR coupled with HUM analysis.This method is rapid,sensitive and specific for diagnosing of parasites in infected Sand flies and ideal for large scale genotyping projects.

Tioxolone niosomes exert antileishmanial effects on Leishmania tropica by promoting promastigote apoptosis and immunomodulation

Objective: To explore the antileishmanial effect of tioxolone and its niosomal form against Leishmania tropica. Methods: Tioxolone niosomes were prepared by the hydration method and were evaluated for morphology, size, release study, and encapsulation efficiency. The cytotoxicity of tioxolone and its niosomal form was measured by MTT assay, leishmanicidal activity against promastigote and amastigote by MTT assay, apoptosis by flow cytometry, IL-12, IL-10 and metacaspase gene expression levels by q-PCR. Results: Span/Tween 40 and Span/Tween 60 niosomes had good physical stability as depicted in their size distribution curves and high encapsulation efficiency(>99%). The release profile of the entrapped compounds showed Fickian’s model of tioxolone delivery based on diffusion through lipid bilayers. With the IC50 value for amastigote as(24.5±2.1) μg/mL and selectivity index as 10.5, the Span/Tween 60 niosome(NT2) had a superior effect to other drugs. The CC50 value and IC50 of promastigote value for NT2 were(257.5±24.5) μg/mL and(164.8±20.6) μg/mL, respectively. The flow cytometric analysis showed that tioxolone and niosomal forms induced apoptosis of Leishmania tropica promastigotes in a dose-dependent manner. NT2 increased the expression level of IL-12 and metacaspase genes and decreased the expression level of the IL-10 gene.Conclusions: Niosomes of tioxolone play an immunomodulatory role in increasing Th1 cytokine profile and inhibiting the Th2 cytokine profile. It could be used for treatment of anthroponotic cutaneous leishmaniasis.

Leishmania tropica: The comparison of two frequently-used methods of parasite load assay in vaccinated mice

Objective:To compare limiting dilution assay and real-time PCR methods in Leishmania tropica parasite load measurement in vaccinated mice.Methods:BALB/c mice were vaccinated by Leishmania tropica soluble Leishmania antigen or recombinant Leishmania tropica stressinducible protein-1 with/without adjuvant.After three vaccinations,mice were challenged by Leishmania tropica promastigotes.Two months after challenge,the draining lymph nodes of mice footpad were removed and parasite load was assayed by limiting dilution assay and real-time PCR methods.Limiting dilution assay was done by diluting tissue samples to extinction in a biphasic medium.For real-time PCR,DNA of the lymph nodes was extracted,equal dilutions of each sample were prepared and hot-start real-time PCR was done using appropriate primers.The data of the two methods were compared by appropriate statistical methods.Results:Both methods were able to measure different levels of parasite load in vaccinated/unvaccinated mice.In addition,wherever parasite load of a group was estimated high(or low)by one method,the estimated parasite load by another method was the same,although statistically significant differences were found between some groups.Conclusions:Both methods lead to approximately similar results in terms of differentiating parasite load of the experimental groups.However,due to the lower errors and faster process,the real-time PCR method is preferred.

Role of higher levels of post-challenge antibodies in protective vaccination against Leishmania tropica infection of BALB/c mice

Objective:To study the role of antibodies in protection against Leishmania tropica(L.tropica)infection in the experimental model of BALB/c mice.Methods:BALB/c mice were vaccinated against L.tropica by soluble Leishmania antigen or recombinant L.tropica stress-inducible protein-1(LtSTI1)of L.tropica,and against Leishmania major(L.major)by soluble Leishmania antigen.Monophosphoryl lipid A was used as an adjuvant.The L.tropica-or L.major-vaccinated mice were challenged by L.tropica or L.major,respectively.The levels of anti-Leishmania antibodies(IgG1 and IgG2 a)were determined after vaccination and after challenge.Results:All vaccinated groups caused a higher antibody response in comparison with the control group.The L.major-vaccinated group showed lower IgG1 response than the control group after the challenge.Conversely,in L.tropica-vaccinated mice,the levels of antibodies were higher than the control group.Moreover,the group receiving rLtSTI1 and monophosphoryl lipid A showed higher levels of antibodies than those of the rLtSTI1 group.In vaccinated mice,antibody responses against L.tropica remained high until 16 weeks after the challenge.Conclusions:The higher levels of post-challenge antibodies are associated with protective vaccination against L.tropica infection of BALB/c mice.Our findings provide new insight into the association of antibody with vaccine-induced protective immunity against L.tropica infection.More studies are needed to clarify the role of antibody in protection against L.tropica.

Effects of different colours in the visible region on <i>Leishmania Tropica</i>

The aim of this study was to investigate the wavelength-dependency of chromotherapy effects on cutaneous Leishmaniasis parasite growth. Chromotherapy uses visible range radiations to improve healing;however, its effects on parasite are not well understood. Leishmania Tropica was irradiated using seven (7) different wavelengths of visible region. Optical density was observed, which showed that red colour (644 nm) wavelength inhibited the growth of parasite while other colour wavelengths also affected the growth of parasite. It is, therefore, suggested that as red colour inhibits the growth of parasite so patients suffering from l.tropica can be treated with the application of red colour.

Effect of Vigna radiata, Tamarix ramosissima and Carthamus lanatus extracts on Leishmania major and Leishmania tropica: An in vitro study

Objective:Current therapy strategies of leishmaniasis have some problems such as high cost,toxicity and side effects.Plant extracts can be a source of drugs to control leishmaniasis.In this study,the effect of hydroalcoholic and chloroformic extracts of Vigna radiata,Tamarix ramosissima,and Carthamus lanatus on Leishmania major and L.tropica was studied.Methods:The plant samples were collected from west of Iran and their extracts were prepared.Antipromastigote activity assay of all extracts was done using tetrazolium-dye assay.Results:Only high concentrations of V.radiata and C.lanatus were able to inhibit Leishmania,while both high and low concentrations of T.ramosissima had antileishmanial effect.No difference was observed between hydroalcoholic with chloroformic extract of each plant.Conclusion:Altogether,the results revealed the antileishmanial activity of T.ramosissima extracts against L.major and L.tropica,indicating its potential as an antileishmanial agent.

中国南部高等真菌的热带亲缘

中国南部 ,特别是北回归线以南的地区 ,高等真菌区系与世界热带真菌区系有着广泛的联系。该地区高等真菌具有明显的热带性质 ,主要有泛热带成分、热带亚洲 -热带非洲和热带亚洲的成分 ,尽管在该区高等真菌区系中也有相当比重的温带成分。该区系中某些属种可能有古南大陆起源的背景 ,另一些区系成分与热带亚洲的有密切的亲缘关系。

凹纹胡蜂与黑尾胡蜂蛋白氨基酸分析研究

为弄清胡蜂蛋白氨基酸(特别是人体必需氨基酸)的组成,探索利用胡蜂作食品,作者对凹纹胡蜂Vespa velutina auraria Smith与黑尾胡蜂Vespa tropica ducalisSmith成虫、蛹、幼虫、巢室、巢壳的氨基酸成分和含量进行了分析,结果报道如下.

两种捕食螨对紫跳虫的捕食功能研究

研究热带吸螨(Bdella tropicaAtyeo)和分吸螨(Bdella distinctaBaker and Balogh)对紫跳虫(Hypogastrura communisFolsom)的捕食功能反应,结果表明:两种捕食螨均能捕食紫跳虫,且热带吸螨对紫跳虫的瞬时发现率(a=0.7132)高于分吸螨(a=0.5839),但猎物处理时间(Th=0.3269 d)长于分吸螨(Th=0.1938 d),因此理论最大捕食量(Namax.=3.06)也低于分吸螨(N=5.16),参数a/Th说明分吸螨对紫跳虫的捕食功能比热带吸螨高。

热带吸螨对二斑叶螨的捕食作用

试验确定了热带吸螨能够捕食二斑叶螨,能生长发育繁殖.在20-35℃温度范围内,热带吸螨对二斑叶螨的功能反应均符合HollingⅡ型的圆盘方程,在20-30℃温度范围内,随着温度的上升,热带吸螨雌成螨对二斑叶螨雌成螨的捕食量增加,处理猎物时间缩短,单位时间攻击率增大,控制能力也增强;与30±1℃相比,在35±1℃时,单位时间攻击率虽有增强,但日最大捕食量明显下降,处理猎物时间延长,控制能力显著下降.

热带水草红椒的组培快速繁殖及色彩显现研究

研究了热带水草红椒(Cryptocorynewendtiitropica)的组培快速繁殖及其色彩显现。结果表明,适宜的芽增殖培养基为MS+BA3mg/L+NAA0.5mg/L,而适宜的生根培养基为1/2MS的无激素培养基。在此生根培养基中,植株上部叶能显现出其独特的红褐色彩,其呈现的颜色主要取决于花青素含量。类胡萝卜素含量和Chlb/Chla比值的提高有利于其色彩的显现。添加1mg/L的GA3可促进植株的生长。

凹纹胡蜂与黑尾胡蜂生物学初步研究(膜翅目:胡蜂科)

本文首次介绍了凹纹胡蜂Vespa velutina auraria Smith与黑尾胡蜂V.tropica ducalis Smith在实验室内饲养和自然条件下的生物学习性观察情况。结果表明,两种胡蜂在室内温度14—25℃,相对湿度43—89％条件下完成一代发育所需时间为33—53天,其中卵期9—15天,幼虫期10—18天,蛹期15一20天,全年可发生三代。工蜂寿命为24—142天,雄蜂4—60天。 凹纹胡蜂可在人工控制条件下越冬和自行营巢。 两种胡蜂的主要天敌是蜂蝙科Stylopidae和寄绳科Tachinidae昆虫。

热带头梗霉在中国NL351杨树上的首次发现

为研究NL351杨树上的真菌种类,对其杨木边材进行了组织分离,通过纯化培养,得到了内生菌热带头梗霉(Cephaliophora tropica)。采用形态特征和分子生物技术相结合的方法对其热带头梗霉进行了确凿鉴定,并确认分布于中国的NL351杨树是热带头梗霉的新寄主,此菌在杨树上发现属于首次报道。

First report on molecular characterization of Leishmania species from cutaneous leishmaniasis patients in southern Khyber Pakhtunkhwa province of Pakistan

Objective: To report presence of Leishmania major in Khyber Pakhtunkhwa of Pakistan, where cutaneous leishmaniasis(CL) is endemic and was thought to be caused by Leishmania tropica only. Methods: Biopsy samples from 432 CL suspected patients were collected from 3 southern districts of Khyber Pakhtunkhwa during years 2011–2016. Microscopy on Giemsa stained slides were done followed by amplification of the ribosomal internal transcribed spacer 1 gene. Results: Leishmania amastigotes were detected by microscopy in 308 of 432 samples(71.3%) while 374 out of 432 samples(86.6%) were positive by ribosomal internal transcribed spacer 1 PCR. Subsequent restriction fragment length polymorphism confirmed Leishmania tropica in 351 and Leishmania major in 6 biopsy samples. Conclusions: This study is the first molecular characterization of Leishmania species in southern Khyber Pakhtunkhwa. It confirmed the previous assumptions that anthroponotic CL is the major CL form present in Khyber Pakhtunkhwa province. Furthermore, this is the first report of Leishmania major from a classical anthroponotic CL endemic focus identified in rural areas of Kohat district in southern Khyber Pakhtunkhwa.

First molecular identification of Leishmania species in a new endemic area of cutaneous leishmaniasis in Lorestan,Iran

Objective:To identify Leishman{u using PCR.Methods:This studs was conducted from April2009 to March 2011 in order to identify Leishmania species in a new endemic area of CL in Lorestan.Iran.Samples were taken from 62 patients that referred to the health centers in different cities of Lorestan province,the presence of Leishmcania was confirmed using direct smear and then grown in NNN media and mass cultured in RPM!1640 medium supplemented with 10%heat-inactivated fetal bovine serum.DNA was extracted from cultured promastigotes and used in P15-PCR.Results:45(72.6%)samples out of 62 showed a hand in the range of 485 hp and 17(27.4%)with a hand in the range of 626 hp which were similar to standard strains of Leichmania tropica(L.tropical and Leishnrania major(L.major),respectively.50(65.80%)of samples were collected from people with no history of travel in at least a year prior to the onset which shows that indigenous source of infection.Conclusions:Since the vector and reservoir of the two species are different.so precise and extensive control and prevention methods should be designed and earned out.

黑尾胡蜂毒液抗菌肽的分离纯化及功能研究

由于病原微生物对众多传统抗生素药物的耐药性问题日益突显,因此寻找新型的抗菌药物显得尤为紧迫,多肽类制剂因其具有广谱的抗菌活性,已成为克服病原微生物耐药性的潜在候选药物.从黑尾胡蜂毒液中分离并鉴定了一种具有抗菌活性的多肽,命名为VT-1.VT-1对临床致病菌均表现出广谱的抗菌活性.此外,VT-1对人红细胞显示出微弱的溶血活性.

一株十二烷基硫酸钠高效降解菌的分离鉴定、降解特性及代谢途径研究

为获得更丰富的十二烷基硫酸钠(sodium dodecyl sulfate,SDS)降解菌资源,从本实验室筛选保存的复合菌系SDS1中,分离出1株以SDS为唯一碳源生长的高效降解菌D2,结合形态学观察、生理生化特征及16S rRNA序列分析,结果显示D2为帕拉伯克霍尔德菌(Paraburkholderia tropica)。实验结果证实,在温度30℃、pH 7、盐度0.1%(W/W)、附加氮源硝酸钠+氯化铵的条件下培养48 h,D2对初始浓度1 200 mg/L的SDS降解率达到98.3%。采用离子色谱法、气相色谱法、液相色谱法对D2在SDS降解过程中的主要代谢产物进行检测与鉴定,初步推测出其对SDS的降解途径可能为SDS→硫酸盐+1-十二烷醇→十二烷醛→十二烷酸→乙酸→CO+HO。本研究可为含SDS表面活性剂废水的高效降解处理提供微生物资源。

红椒生根培养基配方的优化

以红椒不定芽为材料,1/2 MS培养基,在(25±1)℃,30μmol·m-2·s-1,10 h·d-1光照条件下,采用正交试验设计方法研究IBA、NAA、6-BA组合对椒草生根的影响,优化生根培养基配方。结果表明:6-BA是影响红椒生根的主要因子,组合1/2 MS+IBA 0.5 mg·L-1+NAA 1.0 mg·L-1+6-BA 0.5 mg·L-1较好,既有利于生根,又有较高的增殖率,生根率100%,每丛根数11.90条,根长1.02 cm,增殖率2.8934,每丛小苗11.58株,苗高1.48 cm。

曲腿龟甲轮虫和热带龟甲轮虫的mtDNA COI基因和rDNA ITS序列差异以及几种龟甲轮虫的系统发生关系

从芜湖市镜湖和汀棠湖中分别采得曲腿龟甲轮虫(Keratella valga)和热带龟甲轮虫(K.tropica)个体,实验室内对其进行克隆培养.运用常规的分子生物学技术,对各30个曲腿龟甲轮虫克隆和热带龟甲轮虫克隆分别进行了mtDNA COI基因和rDNA ITS的扩增、测序和序列分析.以萼花臂尾轮虫轮虫(Brachionus calyciflorus)为外群,结合Genbank上已登录的几条其它龟甲轮虫COI基因和ITS序列,以及测序所得的60个COI基因和ITS序列构建系统发生树(ML、MP、NJ和贝叶斯树).结果表明,曲腿龟甲轮虫和热带龟甲轮虫间COI基因序列差异百分比为24.9%-26.5%,平均为25.7%.曲腿龟甲轮虫个体间序列差异百分比为0-2%,热带龟甲轮虫个体间序列差异百分比为0-1.5%.两种轮虫间ITS序列差异百分比为6.1%-7.6%,平均为7.25%.曲腿龟甲轮虫个体间序列差异百分比为0-1.1%,热带龟甲轮虫个体间序列差异百分比为0-0.7%.基于COI基因序列和ITS序列构建的四个系统树均支持曲腿龟甲轮虫单倍型和热带龟甲轮虫单倍型各自成支.

热带地区月季扦插漂浮育苗技术

漂浮育苗是无土育苗式工厂化育苗的一种形式,它具有生根快、生长迅速、病虫害(尤其是土壤病虫害)少、嫁接成活率高的优势,且能获得整齐一致的壮苗、缩短育苗周期、节约成本。从漂浮槽的建立与消毒、水与基质的准备、营养液配制、装盘、扦插、生长条件、炼苗及移栽等方面介绍了热带地区月季扦插漂浮育苗技术,保证热带地区切花月季优质种苗供应。