结核分枝杆菌Rv3428c重组蛋白表达与纯化及其单克隆抗体的制备

目的 制备并鉴定Rv3428c的单克隆抗体，为结核分枝杆菌快速特异性抗原检测奠定基础。方法 同源重组得到pET28a-Rv3428c表达载体，转入E.coli BL21(DE3)后进行IPTG诱导表达，用Ni-NTA亲和层析纯化目的蛋白。重组蛋白免疫小鼠后进行细胞融合，间接ELISA、Western Blot筛选阳性杂交瘤细胞系，以筛选获得的阳性杂交瘤细胞诱导小鼠产生腹水，蛋白A亲和层析法纯化抗体，BCA法测定浓度，ELISA和SDS-PAGE进行单克隆抗体的鉴定及亚型和效价的测定。结果 成功构建pET28a-Rv3428c表达载体、表达并纯化出目的蛋白。Rv3428c蛋白免疫小鼠使小鼠产生致敏B淋巴细胞，在聚乙二醇作用下，淋巴细胞可与骨髓瘤细胞发生融合，形成杂交瘤细胞。筛选得到的阳性杂交瘤细胞株，通过体内诱生法制备获得Rv3428c单克隆抗体。该单抗属于IgG2a亚类，κ型轻链，效价约为1∶128 000,浓度为4 mg/ml。结论 本研究制备并纯化了Rv3428c重组蛋白，获得了抗Rv3428c单克隆抗体，为Rv3428c用于结核分枝杆菌快速特异性抗原检测奠定基础。     

人IL-12基因表达对含信号肽mtb8.4基因疫苗的免疫效应

[目的]观察结核分枝杆菌含信号肽Mtb8.4(MS)基因疫苗与人白细胞介素12(hIL-12)表达质粒联合免疫小鼠所诱导的细胞免疫应答。[方法]15只C57BL/6N小鼠随机分为MS基因疫苗 hIL-12质粒组(联合免疫组)、MS基因疫苗组、卡介苗(BCG)组、空载体组和PBS组,基因疫苗、空载体和PBS经肌内注射法免疫各组小鼠,每隔3周免疫1次,共免疫3次,BCG组经尾部皮下注射1×106 CFU BCG免疫1次。采用酶联免疫吸附法(ELISA)检测小鼠脾细胞培养上清中细胞因子水平;采用乳酸脱氢酶(LDH)释放法检测免疫小鼠细胞毒性T细胞(CTL)杀伤活性。[结果]联合免疫组能诱导较强的抗原特异性Th1型细胞免疫应答,免疫小鼠脾细胞培养上清液IFN-γ和IL-2水平(分别为(1546.18±36.23)pg/ml、(681.64±40.88)pg/ml),显著高于MS基因疫苗组,与BCG组相当,IL-4分泌减少,特异性CTL杀伤活性增强。[结论]hIL-12表达质粒与MS基因疫苗联合免疫后,能够增强MS基因疫苗所诱导的细胞免疫应答。     

疫苗候选抗原HBHA通过诱导IL-17发挥抗结核免疫保护效应

为探讨疫苗候选抗原—结核分枝杆菌肝素结合血凝素（HBHA）通过黏膜接种诱导的保护性免疫效应。动物实验均使用6~8周龄雌性C57BL/6小鼠。30只小鼠接受不同的免疫接种策略,采用随机数字表法分为对照组、早期分泌抗原靶蛋白-6（ESAT-6）滴鼻组、HBHA滴鼻组、BCG初免PBS对照组、BCG初免HBHA加强组,每组6...     

新型肺结核三抗原组分疫苗的开发和小鼠免疫原性研究

目的构建高效表达结核分枝杆菌(Mycobacterium tuberculosis,MTB)ESAT6、rv3407、Rpf B三个显性抗原基因(3 antigens,3Ag)的重组人5型腺病毒(Adenovirus 5,Ad5)载体疫苗,在小鼠模型中初步完成药效学研究。方法用Ad EasyTMXL腺病毒载体系统构建重组质粒p Ad5-3Ag,经Pac I线性化后转染AD293细胞获得表达3Ag融合蛋白的Ad5-TPA-3Ag;采用Western blot检测腺病毒载体中融合蛋白的表达;将纯化的病毒颗粒以肌肉注射和滴鼻两种方式免疫小鼠,采用ELISA检测血清中抗体效价,对病毒的免疫剂量进行优化。结果含3Ag基因的重组腺病毒载体构建成功。包装的病毒Ad5-TPA-3Ag以8×107IFU的剂量免疫小鼠42d时各抗原的效价最高,ESAT6肌肉注射和滴鼻的最高效价分别为4.54和4.87;rv3407肌肉注射和滴鼻的最高效价分别为3.41和3.60;Rpf B肌肉注射和滴鼻的最高效价为3.63和2.42。结论 Ad5-TPA-3Ag在小鼠模型中可以诱导良好的免疫应答,可能成为一种新型的针对MTB潜伏感染的候选疫苗。     

结核分枝杆菌MPT64蛋白多克隆抗体的制备

目的利用原核表达系统获取结核分枝杆菌MPT64蛋白,制备并鉴定MPT64蛋白多克隆抗体。方法提取结核分枝杆菌H37Rv株基因组DNA并以其为模板,PCR扩增获取目的基因MPT64,并插入蛋白表达载体PET-26b(+)。将纯化得到的MPT64蛋白免疫新西兰兔,最终制备得到MPT64多克隆抗体,通过免疫印迹法鉴定该抗体的特异性。结果成功获取结核分枝杆菌MPT64基因及其重组蛋白,分子量约为27 k D左右。免疫印迹实验证实,以该重组蛋白为免疫原制备得到的多克隆抗体可特异性识别来自于结核分枝杆菌MPT64基因重组腺病毒疫苗所表达的MPT64蛋白。结论获得高特异性的结核分枝杆菌MPT64蛋白多克隆抗体,有助于结核分枝杆菌疫苗的研发。     

内含子A提高分枝杆菌热休克蛋白65DNA疫苗的免疫原性

目的 探讨以内含子A增强外源抗原在真核细胞中的表达效率及其对DNA疫苗在小鼠中免疫原性的影响.方法 以分枝杆菌热休克蛋白65 (Hsp65)为模式抗原,将其分别克隆入含有内含子A和不含内含子A的真核表达载体pCMV4.0和pVAX1,将表达Hsp65的不同重组质粒瞬时转染人胚肾上皮细胞系293T并免疫接种BALB/c小鼠,应用ELISA分析其所诱导产生的抗原特异性抗体水平及亚类,并以酶联免疫斑点试验(ELISPOT)和胞内细胞因子染色技术(ICS)分析其诱导产生的细胞免疫应答反应.结果 含有内含子A的重组表达质粒pCMV4.0hsp65较之不含内含子A的pVAX1 hsp65在293T细胞中的抗原表达量更高,免疫小鼠6周后诱导产生的抗-Hsp65特异性总IgG抗体水平(3.76 ±0.23对3.15 ±0.22,P＜0.01)和IgG2a/IgG1比值(4.08 ±0.04对2.23±0.12,P＜0.01)也更高,差异有统计学意义;分泌γ-干扰素(IFN-γ)的CD4+T淋巴细胞频率[(2.0±0.058)％对(1.5±0.087)％,t=4.804,P＜0.01]和CD8+T淋巴细胞频率[(0.6±0.058)％对(1.0±0.115)％,t=3.098,P＜0.05]增加,差异有统计学意义,提示其诱导产生了增强的Th1型免疫应答.结论 内含子A可增强分枝杆菌Hsp65在真核细胞中的表达效率并可提高DNA疫苗在小鼠中的免疫原性.     

抗结核分枝杆菌早期分泌性抗原靶蛋白6的新型单克隆抗体的制备及其临床应用

目的制备抗结核分枝杆菌早期分泌性抗原靶蛋白6(ESAT-6)单克隆抗体,以应用于多种检测技术和临床研究。方法设计并合成结核分枝杆菌ESAT-6特定抗原肽序列,免疫BALB/c小鼠,取其脾细胞和小鼠骨髓瘤细胞SP2/0融合,经酶联免疫吸附试验(ELISA)检测、多次有限稀释亚克隆建立能稳定分泌抗结核分枝杆菌特异抗原肽单克隆抗体的杂交瘤细胞系,纯化后行免疫球蛋白亚型分析,并鉴定分析该单克隆抗体在蛋白质免疫印迹法、免疫沉淀法和酶联免疫法(ELISA)检测中的应用效果。收集结核分枝杆菌培养上清液样本38份,非结核分枝杆菌培养上清液样本20份,结核性胸水样本32份,非结核性胸水样本24份以及健康对照血清样本20份。结果单克隆抗体免疫球蛋白亚型分析显示抗体类型为IgM、κ型,并可应用于Western blotting和免疫沉淀法。ELISA定量检测显示该特异性抗体检测结核分枝杆菌的敏感度为95.7%(67/70),特异度为100%(64/64)。结论 ESAT-6单克隆抗体用于结核分枝杆菌检测有较高的敏感性和特异性,可用于结核病的早期诊断。     

抗苗勒管激素的单克隆抗体制备与鉴定

目的表达高纯度的抗苗勒管激素蛋白,以该蛋白作为抗原制备单克隆抗体,并鉴定抗体亚型,为进一步开发优质的抗苗勒管激素诊断试剂盒做技术储备。方法在表达载体pET-28a(+)质粒中接入经密码子优化的抗苗勒管激素基因(AMH)片段,转化于BL21细菌中,异丙基硫代-β-D-半乳糖苷(IPTG)诱导重组蛋白大量表达,对上清中可溶性蛋白进行纯化。使用SDS-PAGE分析纯化蛋白的纯度。经纯化的抗原蛋白3次皮下免疫小鼠,挑选产生高效价的抗体的小鼠,取其脾细胞,制备悬液。经细胞杂交技术与骨髓瘤细胞进行融合,筛选能稳定持久分泌抗AMH抗体的杂交瘤细胞株,并制备腹水抗体。通过亲和层析技术、BCA蛋白检测方法等测定手段,纯化抗体、检测抗体浓度。结果筛选到2株能持久分泌抗AMH蛋白单克隆抗体的杂交瘤细胞株,分别为克隆细胞株3F6和4E7,分泌抗体均为IgG1亚型。结论筛选出稳定分泌高纯度的抗苗勒管激素抗体的2株杂交瘤细胞,并制备单克隆抗体。下一步将其应用于AMH检测试剂盒的开发。     

菊糖作为结核分枝杆菌和人乳头瘤病毒疫苗佐剂的免疫原性研究

目的研究菊糖的免疫佐剂效果,探讨菊糖与结核分枝杆菌(Mtb)ESAT-CFP10融合蛋白EC、人乳头瘤病毒16型E6-E7融合蛋白E6E7联合免疫小鼠的免疫应答特征。方法将菊糖(Inulin)分别与E6E7、EC共价结合制备E6E7-I、EC-I疫苗,采用E6E7-I、EC-I免疫小鼠,通过比较小鼠血清中抗原特异性IgG滴度,脾淋巴细胞抗原特异性IFN-γ、TNF-α分泌水平和CD4~+T、CD8~+T亚群分布,评价菊糖的疫苗佐剂功能。结果 (1)第7、14、21 d,菊糖结合的EC-I、E6E7-I组抗原特异性抗体IgG滴度分别高于单独抗原EC组、E6E7组(P0.05),且未检出抗菊糖抗体。(2)EC-I、E6E7-I能分别诱导出高于EC组、E6E7的脾淋巴细胞IFN-γ、TNF-α分泌水平(P0.05)。(3)EC-I组、EC组、PBS组的方差分析结果表明,EC-I组CD8~+T、CD4~+T细胞亚群所占百分比高于EC组(P0.05;P=0.017);E6E7-I组的CD8~+T、CD4~+T细胞亚群所占百分比高于E6E7-I组,分别(P=0.007;0.016)。结论菊糖佐剂共价结合E6E7、EC蛋白,能有效激活机体的抗原特异性体液免疫和细胞免疫,且佐剂自身免疫原性低,提示菊糖作为HPV16型宫颈癌疫苗、TB疫苗佐剂具有良好的佐剂效应。     

结核分枝杆菌Lsr2蛋白原核重组表达及其多克隆抗体制备

目的 利用原核系统获得重组结核分枝杆菌Lsr2蛋白,制备抗Lsr2多克隆抗体。 方法 以临床标准株 H37Rv DNA为模板,合成引物PCR扩增Lsr2核酸序列,插入原核表达载体pET28a,转化大肠杆菌Rosetta(DE3),经IPTG诱导表达,柱上复性纯化后,重组蛋白免疫新西兰大白兔,制备并纯化抗Lsr2多克隆抗体,采用间接ELISA和Western-blotting实验对抗体进行验证。 结果 成功构建pET28a-Lsr2表达质粒,重组蛋白Lsr2经SDS-PAGE电泳鉴定分子量为14 kD。亲和层析及柱上复性后,纯化的重组蛋白纯度在95%左右。制备的抗Lsr2多克隆抗体效价在1:5.5×10⁶以上,并能特异性识别纯化的重组蛋白和结核分枝杆菌菌体蛋白。 结论 成功在原核系统内表达并纯化了重组结核分枝杆菌Lsr2蛋白,制备了高效价的抗Lsr2多克隆抗体,为进一步研究Lsr2蛋白的功能,筛选其下游相互作用蛋白以及相关分子机制研究奠定了基础。     

Clinical and laboratory evidence of the importance of the tick D. marginatus as a vector of B. burgdorferi in some areas of sporadic Lyme disease in Bulgaria

56 adult ticks D. marginatus and 38 adult ticks H. punctata were sampled by the flagging method, transferred to the laboratory, dissected and tested for infection with Lyme disease spirochetes. 3 (7.9%) of the adult H. punctata and 2 (3.57%) of the adult D. marginatus were infected with B. burgdorferi (Bb). This report also presents the case of a patient, who developed Lyme disease symptoms after he had been bitten by a D. marginatus tick. The clinical diagnosis was confirmed by serological testing and by a biopsy, taken from the patient's skin lesion. The skin biopsy was examined under an electron microscope and Bb was found in the sections of the deeper strata of the dermis in two structural forms: (a) cylindrical bodies (protoplasm cylinder) with circular ends covered with a three-layered membrane; (b) granules, situated among the collagenous fibres either closely adhered to them or covered with a membrane. The result of the study demonstrates that in single cases in some ecosystems, ticks D. marginatus might be implicated in Bb transmission to humans as secondary vectors.     

Absorption,metabolism and elimination of N,N-dimethylformamide in humans

Summary Excretion of N,N-dimethylformamide (DMF) and DMF metabolites N-hydroxymethyl-N-methylformamide (MF), N-hydroxymethyl-formamide (F) and N-acetyl-S-(N-methylcarbamoyl)cysteine (AMCC) has been monitored in the urine of volunteers during and after their 8-h exposure to DMF vapour at a concentration of 10, 30 and 60 mg · m^–3. The pulmonary ventilation in these experiments was typically about 101 · min^–1 and the retention in the respiratory tract was 90%. After exposure to 30 mg DMF · m^–3, the yield of compound determined in the urine represented 0.3% (DMF), 22.3% (MF), 13.2% (F) and 13.4% (AMCC) of the dose absorbed via the respiratory tract. The excretion curves of the particular compounds attained their maximum 6–8h (DMF), 6–8h (MF), 8–14h (F) and 24–34h (AMCC) after the start of the exposure. The half-times of excretion were approximately 2, 4, 7 and 23 h respectively. In contrast to slow elimination of AMCC after exposure to DMF, AMCC was eliminated rapidly after AMCC intake. This discrepancy could be explained by rate-limiting reversible protein binding of a reactive metabolic intermediate of DMF, possibly methylisocyanate.     

The impact of needle-exchange programs on the spread of HIV among injection drug users: A simulation study

Objective. To determine the impact of the implementation of a needle-exchange program (NEP) on the spread of human immunodeficiency virus (HIV) in an injection drug user (IDU) community. We conducted a Monte Carlo simulation study of a theoretical population of 10,000 IDUs. The population was followed monthly from 1984 to 2000. HIV was assumed to be transmitted only by needle sharing. The NEP was introduced in 1989 and evaluated over a period of 11 years. The impacts of the proportion of the population attending the NEP, the risk level of IDUs attending the NEP, the reduction in needle-sharing frequency, and the number of new needle-sharing partners acquired at the NEP on prevalence and incidence of HIV were determined. Increasing the proportion of the population who always attend the NEP and eliminating needle-sharing incidents among IDUs who always attended the NEP were the most effective ways of reducing the spread of HIV. Attracting high-risk users instead of lower risk users to the NEP also reduced the spread of HIV, but to a lesser extent. NEPs are effective at reducing the spread of HIV; even under the worst case scenario of low risk users more likely to attend the NEP, one additional partner per month as a result of attending the NEP, and poor NEP attendance, the estimated prevalence was still less than that from the scenario without an NEP. Under our model, NEPs were shown to reduce the spread of HIV significantly. Efforts should be focused on getting as many IDUs as possible to become regular NEP attenders and stop sharing needles rather than partially reducing the frequency of sharing by a larger number of IDUs.     

Re-assessment of the influence of polymorphisms of phase-II metabolic enzymes on renal cell cancer risk of trichloroethylene-exposed workers

Problem Individual differences in susceptibility to trichloroethylene-induced nephrocarcinogenicity may be conferred by genetic polymorphisms of glutathione S-transferases (GST), because enzymes of this group are pivotal for the metabolic activation of trichloroethylene. Because of a potential involvement of N-acetylation in the detoxication of reactive trichloroethylene metabolite(s) to N-acetyl-cysteine derivatives, polymorphisms of the NAT2 gene may also be relevant. Methods The primary collective used for a re-investigation of these questions was that of a hospital-based case-control study by Brüning et al. (Am J Ind Med 43:274–285, 2003) of 134 renal cell cancer cases (20 cases exposed to trichloroethylene) and 401 matched controls. Genetic polymorphisms of GSTT1, GSTM1, GSTP1 and NAT2 were studied. Additional control collectives of non-diseased persons were used for comparison of allele frequencies. Results No genetic influences on the development of renal cancer due to trichloroethylene were apparent, related to the deletion polymorphisms of GSTT1 and GSTM1, as well as to the NAT2 rapid/slow acetylator states. However, renal cell cancer cases displayed a somewhat higher proportion of the homozygous GSTP1 313A wild type (GSTP1*A), although this was not statistically significant (χ² test: P = 0.1071, when using only the original controls of Brüning et al. (2003); P = 0.0781 with inclusion of the additional controls). Conclusion The re-investigation does not confirm the working hypothesis of an influence of the deletion polymorphisms of the glutathione S-transferases GSTT1 and GSTM1 on renal cell cancer development due to high occupational exposures to trichloroethylene.     

Screening of the municipal water system of La Plata, Argentina, for human intestinal parasites

The La Plata River, though severely contaminated by intestinal parasites through the discharge of tons of crude fecal material from a main sewage channel, nevertheless provides drinking water to two-thirds of La Plata, Argentina, after conventional purification at a processing plant. With intestinal parasitosis being endemic here, we investigated the importance of this water in transmitting such pathogens to the city's populace by means of standard methodology for sample acquisition and processing involving filter-concentration of waterborne particulates. Of 14 tap-water samples collected from the distribution network, 12 pertained to four zones (A-D) within the city center; while the remaining 2 were obtained near the processing plant, 15 kilometers outside the city. Although parasites were found within the samples derived from the four urban zones, none were detected in the specimens obtained near the plant. The four downtown areas differed from each other as to the quantity and nature of the parasites present in their water: whereas zones A and B registered similar lower levels of contaminants, C and D exhibited higher values significantly different from the former two and from each other. Given an average parasite count/l citywide of 0.38 and a probability of encountering a parasite within 11 of water of 0.32, the municipal network is seen to contribute to the transmission of intestinal parasites. A routine system of water-quality control is therefore needed throughout the city along with the establishment of infrastructures for locating and eliminating peripheral sources of contamination.     

Seroepidemiology of rickettsial infections in Morocco

The prevalence of antibodies reactive withRickettsia conorii, Rickettsia typhi, Coxiella burnetii andEhrlichia chaffeensis was investigated using indirect immunofluorescence (IFA) test on human sera obtained from 300 blood donors in Casablanca and 126 sera obtained from clinical laboratories in Fez. In sera from Casablanca, antibodies reactive at titers >=1: 32 were found againstR. conorii (7%), andR. typhi (1.7%), but not againstE. chaffeensis. In the sera from Fez, antibodies were also detected againstR. conorii (5.6%),R. typhi (4%), but not againstE. chaffeensis. By Western immunoblotting, seroprevalence forR. conorii was in Casablanca and 4.8% in Fez. Antibodies reactive at titers >=1:50 againstC. burnetii (phase II) were present in sera from Casablanca (1%) and Fez (18.3%).Abbreviations IFA Immunofluorescence assay - MSF Mediterranean spotted fever - PBS Phosphate-buffered saline     

Percutaneous absorption of N,N-dimethylformamide in humans

Summary Skin penetration fo N,N-dimethylformamide (DMF) liquid or vapour was studied in volunteers. Exposure to liquid DMF was performed in two ways: in a dipping experiment, one hand was dipped up to the wrist in DMF for 2–20 min, while in a patch experiment, 2 mmol DMF was applied to the skin and allowed to be absorbed completely. The period of exposure to DMF vapour (50 mg · m^–3) was 4 h. The DMF metabolites N-hydroxymethyl-N-methylformamide (MF), N-hydroxymethylformamide (F), and N-acetyl-S-(N-methylcarbamoyl)cysteine (AMCC) were monitored in the urine. Liquid DMF was absorbed through the skin at a rate of 9.4 mg · cm^–2 · h^–1. Percutaneous absorption of DMF vapour depended strongly on ambient temperature and humidity and accounted for 13%–36% of totally excreted MF. The results suggest that skin absorption of liquid DMF is likely to contribute to occupational exposure substantially more than penetration of DMF vapour. The yield of metabolites after transdermal DMF absorption was only half of that seen after pulmonary absorption. Elimination of MF and F but not that of AMCC was delayed, which supports the contention that AMCC should be used instead of MF as the most suitable biomarker of DMF in cases where percutaneous intake can occur.     

A vulnerable population in a time of crisis: Drug users and the attacks on the World Trade Center

In this article, we present preliminary findings from a qualitative study focused on the impact of the World Trade Center attacks on New York City residents who are current or former users of heroin, crack, and other forms of cocaine. In it, we present data describing their responses to and feelings about the attacks, changes in drug use after the attacks, and factors affecting changes in use. Our analysis is based on 57 open-ended interviews conducted between October 2001 and February 2002. The majority of study participants reported that the attacks had a significant emotional impact on them, causing anxiety, sadness, and anger. Several described practical impacts as well, including significant reductions in income. On September 11th and the weeks and months that followed, several participants who had been actively using did increase their use of heroin, crack, and/or other forms of cocaine. Reductions in use were, however, as common over time as were increases. There was some relapse among former users, but this was limited to those who had stopped using drugs within the 6 months immediately preceding the attacks. A diverse set of factors interacted to control use. For some participants, these factors were internal, relating to their individual motivations and drug use experiences. Other participants were essentially forced to limit use by marked reductions in income. For others, access to health and social service professionals, as well as drug treatment, proved to be key. Department of Family Medicine and Community Health at Montefiore Medical Center     

Selenium content of Portuguese unifloral honeys

The selenium contents of a total of 62 unifloral honeys from Erica spp., Castanea sativa, Eucalyptus spp., Lavandula stoechas, Citrus spp. and Echium plantagineum honeys collected in Portugal were determined by fluorometry after reaction with 2,3-diaminonaphthalene. The selenium levels of the honey samples studied were low, ranging from <1.0 to 2.91 μg/100 g fresh weight. The honeys from Erica spp., C. sativa and E. plantagineum presented the highest selenium values from all the honeys studied (median values 1.69, 1.51 and 1.51 μg/100 g fresh weight), and the honeys from Eucalyptus spp., L. stoechas and Citrus spp. presented the lowest values (median values 1.33, 1.28 and 1.20 μg/100 g fresh weight). The selenium content of Erica spp., was significantly higher than that observed for the Eucalyptus spp., L. stoechas and Citrus spp. and the selenium level of the Eucalyptus spp., was also significantly lower than that observed C. sativa and E. plantagineum honeys. From these results it can be concluded that honey is a marginal source of Se for the Portuguese population.     

Monitoring of Cryptosporidium and Giardia in the Vantaa river basin, southern Finland

We compared two sampling methods to assess the contamination of the Vantaa river basin by Giardia cysts and Cryptosporidium oocysts: 10-1 grab samples, the common river mussel Anadonta piscinalis, were analysed for concentration of (oo)cysts from river water. The samples were collected 2-5 times in autumn 2001 from four wastewater treatment plants and four river water sites located downstream of the plants, and six times from raw water of a drinking water plant using the river as water source. The presence of Giardia and Cryptosporidium was analysed by IF microscopy and PCR. Both cysts and oocysts were detected at all sampling sites, but oocysts were more common than cysts in river water samples. In contrast, cysts were more common in A. piscinalis. Most Cryptosporidium-positive samples were of genotype 2 and Giardia were assemblage B. In river water, MPN of Escherichia coli did not correlate to the presence of (oo)cysts. In conclusion, low (oo)cyst counts were regularly identified in the Vantaa river basin which is contaminated by discharges of treated wastewater of human origin. In general, both methods to appropriate to detect (oo)cysts, but grab samples yielded more positive results. Grab sampling is also more practical and less expensive than analysis of A. piscinalis.