Human leukocyte antigen class-Ⅱ DRB1 alleles and Giardia lamblia infection in children: A case-control study

Objective:To compare the genotype frequencies of HLA class-ⅡDRB1 alleles in Giardia(G.)lamblia-infected children.Methods:A total of 490 Egyptian children aged 2-16 years were subjected to microscopic stool examination to detect G.lamblia infection,and to exclude other intestinal pathogens.On the basis of their microscopic findings,a group of 80 children were chosen as giardiasis cases,another 80 children were confirmed as Giardia free control group by immunochromatographic test,and the remaining children were excluded.Both giardiasis and control groups were then subjected to blood examination to identify their genetic type of HLA-DRB1 alleles.Results:HLA class-ⅡDRB1*03:01 and DRB1*13:01 alleles were significantly associated with G.lamblia infection(P<0.001 for each variable).On the other hand,HLA class-ⅡDRB1*04:02,DRB1*10:01,DRB1*14:01 and DRB1*15:01 alleles were significantly demonstrated in Giardia free children.However,other HLA-DRB1 alleles did not show any significant association with giardiasis.Conclusions:HLA class-ⅡDRB1*03,DRB1*13,DRB1*04,DRB1*10,DRB1*14 and DRB1*15 alleles may be involved in the establishment of host immune response to G.lamblia infection.

人源蓝氏贾第虫(Giardia lamblia)病毒部分基因的克隆及序列分析

目的 从我国蓝氏贾第虫 (Giardialamblia)中寻找贾第虫病毒。方法 对人源贾第鞭毛虫北京株进行了体外纯培养 ,将其总核酸电泳 ,并用DNA酶和RNA酶处理。根据已发表的蓝氏贾第虫病毒基因序列合成一对引物并进行RT PCR ,将产物回收后连接到Pmd18 T载体上进行克隆并测序 ,通过BLAST对GenBank进行同源性搜索。结果 在贾第虫总核酸电泳图谱上观察到一个分子量约 7 0kb的片段。该核酸不能被DNA酶 (10 0 μg/ml)降解。但可被RNA酶 (10 μg/ml)降解。经RT PCR扩增后得到 1条预计 85 6bp的片段 ,所得序列与蓝氏贾第虫病毒基因同源性为 98%。结论 在我国人源贾第鞭毛虫北京株中发现贾第虫病毒 。

In vivo study of pomegranate(Punica granatum) peel extract efficacy against Giardia lamblia in infected experimental mice

Objective: To investigate the efficacy of pomegranate(Punica granatum) peel extract as an alternative treatment on the white laboratory mice against giardiasis.Methods: Experimental animals were divided into five groups, including Group A:control(infected untreated), Group B: infected and fed with pectin 7 days before infection, Group C: infected and fed with pectin starting from 7th day of infection,Group D: infected and fed with pomegranate peel extract 7 days before infection, and Group E: infected and fed with pomegranate peel extract starting from 7th day of infection.Results: Results from this study revealed that the prevention rate in the experimental groups reached approximately 50% by the 10 th day of using pomegranate peel extract.Moreover, stool cyst counts of groups showed a significant reduction in the shedding of cysts approximately 75.6% by day 20 post-infection. ELISA test showed a reduction in Giardia antigen in the stools of the experimental groups which received pomegranate peel extract. The cure rate of these groups was approximately 97.4% by 28 th day of infection.Conclusions: Our present findings indicated that the pomegranate peel extract proved to be valuable in prevention and treatment of Giardia lamblia infection. Further studies are required to determine the effective dose of pomegranate peel extract against Giardia lamblia infection.

贾第虫研究──Ⅰ．西蒙氏贾第虫（Giardia simoni）滋养体和包囊对实验动物的感染

作者从自然感染西蒙氏贾第虫（Ｇｉａｒｄｉａｓｉｍｏｎｉ）的金黄地鼠肠道内和粪便中分离、收集滋养体和包囊，分别经口感染大鼠、小鼠、豚鼠和兔子。结果表明兔子和脉鼠均不感染，大鼠、小鼠均不同程度地感染该虫，且４周龄大鼠比８周龄大鼠更易感（Ｐ＜０．０５），而在小鼠中成年鼠与老年鼠对西蒙氏贾第虫的敏感性没有差异（Ｐ＞０．０５）。滋养体在感染动物的肠道内主要分布于十二指肠前段，中前段和中段。同时观察到结肠和直肠内有包囊，这表明该虫在大鼠、小鼠体内完成了其生活史。作者还对感染前后的滋养体作了蛋白银和铁苏木精染色比较，二者在形态上完全相同。

Construction and characterization of a full-lengh cDNA library from non-fresh Giardia lamblia

Objective:To construct rapidly a full-length cDNA library from nanogram amounts total RIMA of Giardia lamblia(G.lamblia) trophozoites stocked in RNA stabilization reagent.Methods:Total RNA of Giardia was extracted using Trizol reagent.A full-length cDNA library of G.lamblia trophozoites was constructed by a long-distance PCR(LD-PCR) method.The recombinant rate and the coverage rate of full-length clones of the library were evaluated.The inserted fragments were identified and sequenced by PCR amplification.Results:The titer of cDNA library was 3.85×10~7 pfu/mL.The length of inserted fragments ranged from 0.4 to 2.5 kb,and the recombination efficiency accounted for 100%(20/20).The coverage rate of full-length clones is high(17/20). Conclusions:The RNA stabilization reagent may be used to fix the cells and prevent the RNA in cells even though delivered under normal atmospheric temperature.The long-distance PCR can be used to construct a full-length cDNA library rapidly and it needs less RNA than the traditional method from mRNA.

Giardia duodenalis infection among rural communities in Yemen:A community-based assessment of the prevalence and associated risk factors

Objective: To investigate the prevalence and risk factors of Giardia infection among rural communities among rural communities in Yemen.Methods: In this cross-sectional study, a total of 605 stool samples were collected and screened for the presence of Giardia duodenalis(G.duodenalis) cysts and/or trophozoites by using three different diagnostic methods: direct smear, formalin-ether sedimentation,and trichrome staining.A pre-tested questionnaire was used to collect information on the demographic, socioeconomic, behavioural and environmental characteristics of the participants.Results: Overall, 28.1%(170/605) of the participants were infected by G.duodenalis.The prevalence was significantly higher among male participants compared to female(P = 0.034); however, it was not significant among different age groups(P > 0.05).Univariate and multivariate analyses identified four variables as the significant key risk factors of Giardia infection among the sampled communities.These are, in addition to being of the male gender, using unsafe water sources for drinking water, not washing hands after defecation, presence of other family members infected with Giardia, and close contact with domestic animals.Conclusions: The study reveals that Giardia infection is still prevalent among rural communities in Yemen.The provision of clean and safe drinking water, proper sanitation,and health education regarding personal hygiene practices, particularly handwashing, as well as identifying and treating infected family members is imperative and these interventions should be considered in a strategy to control intestinal parasites among these communities in order to curtail the transmission and morbidity caused by G.duodenalis.

A New Duplex Reverse Transcription PCR for Simultaneous Detection of Viable Cryptosporidium parvum Oocysts and Giardia duodenalis Cysts

Objective To simultaneously detect viable Cryptosporidium parvum oocysts and Giardia duodenalis cysts for the purpose of reducing time and cost spent. Methods A duplex reverse transcription polymerase chain reaction （RT-PCR） method was newly developed. Results Using duplex RT-PCR method for the hsp70 gene, viable （oo）cyst concentrations of 10^1 and l0^3 （oo）cysts/100 I.tL could be detected for C. parvum and G duodenalis, respectively. However, after heat-shock stimulation the expression of hsp70 mRNAs was detectable at 10^0 and 10^1 （oo）cysts/100 μL concentrations of C. parvum and G. duodenalis, respectively. Thus, the detection sensitivity was significantly increased when the viable （oo）cysts were exposed to heat shock. Conclusion This study describes a new duplex RT-PCR method for hsp70 gene to detect the viable （oo）cysts of the C. parvum and G duodenalis with less time consumed and at a lower cost. This newly developed duplex RT-PCR method may be used to detect these parasites not only in aquatic environments but also in clinical samples.

The Occurrence of Cryptosporidium and Giardia Infections Among Patients Reporting Diarrheal Disease in Chobe District, Botswana

Diarrheal disease is a chronic public health problem in Chobe District, Botswana. Acute diarrheal outbreaks occur annually with a bimodal seasonal pattern coinciding with major hydrological phenomena. Information is lacking regarding the etiology of reoccurring outbreaks. Cryptosporidium and Giardia are recognized as important waterborne causes of diarrheal disease with Cryptosporidium transmission potentially involving zoonotic reservoirs. In Chobe District, municipal water is obtained from the Chobe River after it exits the Chobe National Park where high concentrations of wildlife occur. Using the Giardia/Cryptosporidium QUIK CHEK rapid antigen cartridge test, we evaluate the occurrence of Giardia and Cryptosporidium among patients (n = 153) presenting to medical facilities in Chobe District with diarrhea (August 2011-July 2012). Cryptosporidium was only identified in children less than two years of age (10%, n = 42, 95% CI 3% - 23%), with positive cases occurring during diarrheal outbreak periods. Infections were not identified in adults despite the high level of human immunodeficiency virus infection/acquired immunodeficiency syndrome (HIV/AIDS) in the region (n = 117). Although not significant, cases were predominately identified in the wet season outbreak (p = 0.94, 25%, n = 12, 95% CI 5% - 57%, dry season outbreaks 3%, n = 30, CI 95% 0% - 17%). Giardia infections occurred across age groups and seasons (7%, n = 153, 95% CI 4% - 10%). This is the first published report of human infections with Cryptosporidium and Giardia in this area of Africa. This study suggests that Cryptosporidium may be a potentially important cause of diarrheal disease in children less than 2 years of age in this region. Further research is required to identify pathogen transmission and persistence dynamics and public health implications, particularly the role of HIV/AIDS and vulnerability to waterborne disease.

Introducing nitazoxanide as a promising alternative treatment for symptomatic to metronidazole-resistant giardiasis in clinical isolates

Objective:To identify the frequencies(F) of ferredoxin and nitroreductase mutations were identified on Iranian clinical isolates of Giardia lamblia in order to predict whether the nitazoxanide can be prescribed as suitable drug for symptomatic to metronidazoleresistant giardiasis.Methods:Forty Giardia lamblia isolates as of 38 symptomatic and two metronidazole-resistant patients were collected from Iran.DNAs were extracted and amplified by targeting ferredoxin and Gl NR genes.The amplicons were directly sequenced to determine gene mutations.Results:The various amino acid substitutions(F:20%,Haplotype diversity:0.891,Tajima's D:-0.44013) were identified by analyzing ferredoxin gene in four symptomatic and two resistant isolates.Only,two haplotypes(F:5%,HD:0.345; Tajima's D:0.77815) characterized in metronidazole-resistant isolates of Gl NR,however,no point mutations was found in symptomatic isolates.Conclusions:Non-synonymous mutations of ferredoxin oxidoreductase gene reduce translational regulatory protein's binding affinity which concludes reduction of ferredoxin expression and its activity.This leads to decrease in metronidazole drug delivery into the cells.Mutations in these isolates may lead to their resistance to metronidazole.No to low synonymous mutations of Gl NR demonstrates that nitazoxanide can be prescribed as promising alternative treatment for symptomatic to metronidazole-resistant giardiasis in Iranian clinical isolates.

Impact of seasonality on the prevalence and risk factors of Giardia lamblia infections among the aborigines

Objective: To determine the prevalence and risk factors of Giardia(G.) lamblia infections among the aboriginal community during the wet and dry seasons. Methods: A total of 473 stool samples from the aborigines in Temerloh, Pahang, Malaysia were collected during wet(n=256) and dry seasons(n=217). Smear of all the PVA-preserved stool samples were subjected to Trichrome staining and microscopic examination under 1 000 伊magnification(Nikon eclipse E100) for the detection of G. lamblia. Positivity was recorded based on the presence of G. lamblia in trophozoite and/or cyst forms. Results: The prevalence of giardiasis was 12.10% and 8.29% during the wet and dry season, respectively. Age of less or equal to 15 years old and presence of other family members with G. lamblia infection were found to be the significant risk factors to acquire G. lamblia infections during both seasons. Untreated water supply was the significant risk factor of giardiasis during the dry season. This study highlighted the possibility of anthroponotic transmission of G. lamblia during both seasons and waterborne transmission during the dry season in the aboriginal community. Conclusions: This study suggests that seasonal variation plays an important role in the prevalence and risk factor of G. lamblia infection in the aboriginal community. Therefore, close contact with Giardia-infected family members and water-related activities or usage of untreated water must be avoided to reduce the burden of G. lamblia infection in this community.

Effect of <i>Commiphora molmol</i>(Myrrh) Extract on Mice Infected by <i>Giardia lamblia</i>

Objective: To investigate the efficacy of (Commiphora molmol(Myrrh)) extract as alternative treatment in hamsters against giardiasis. This study was conducted to evaluate the effect of the new compound (Commiphora molmol(Myrrh)), as agent a natural product extracted from the roots of Zingiber officinal, on Giardia lamblia infection as anti-protozoal and to study the parasitological and Histopathological impact of this treatment on the duodenal mucosa of infected hamsters. Fifty hamsters were divided into five groups: Group (A): normal control. Group (B): Giardia lamblia infected and untreated hamsters served as control. Group (C): infected with Giardia lamblia and treated with metronidazole in dose (120 μg/kg body weight twice daily for 7 successive days). Group (D) Giardia lamblia infected and orally treated with Myrrh (500 mg/Kg for 1 week). Group (E): Giardia lamblia infected and orally treated with combination 1/3 dose of (120 μg/kg body weight) metronidazole and Myrrh (250 mg/Kg) for 1 week). After 2 weeks of the treatment, hamsters were sacrificed. The highest percentages of reduction in the number of Giardia lamblia cysts and trophozoites were in the group receiving combined metronidazole and myrrh. Histopathological examination revealed complete healing of intestinal mucosa after the combined treatment, while partial healing of the lining epithelium of the intestine was noticed after metronidazole and either myrrh plant treatment.

Current situation of Giardia infection in Spain: Implications for public health

Giardia is an enteric protozoan that infects a wide range of vertebrate hosts, being considered a major causative agent of gastrointestinal disease in humans in both developing and developed countries. Giardia infection has also a significant impact on livestock health, causing diarrhoea and resulting in significant economic lost. Transmission is either direct, through the faecal-oral route, or indirect, through ingestion of contaminated water or food. In this article, we review current knowledge about the epidemiology of giardiasis in different populations in Spain, including humans, livestock, companion animals, and wildlife. Environmental contamination of surface waters and raw foods by Giardia cysts is also addressed. Special relevance has been given to the data available on the molecular characterization of the Giardia isolates obtained from clinical and environmental samples, as determining thespecies/assemblages and subassemblages involved is essential for accurately identifying the parasite and assessing zoonotic transmission. The public health significance of these findings has also been thoroughly discussed.

Prevalence of Two Gastrointestinal Parasites <i>Entamoeba histolytica</i>and <i>Giardia lamblia</i>within Samarra City, Iraq

The prevalence of two gastrointestinal parasites the Entamoeba histolytica and Giardia lamblia parasites and their impact on some blood parameters, i.e. packed cell volume (PCV), hemoglobin (Hb%) and total protein (TP) of a total 780 patients (children and adults) admitted to Samarra General Hospital were assessed. Samples of fresh feces were collected in normal physiological saline and examined using Olympic microscopes. The frequency of the parasite E. histolytica was 12.8% (46.3% male and 53.6% female). The highest frequency of infection of E. histolytica (13.8%) was found at age group (1 - 5 years old) followed by <1 year old children while the lowest (7.4%) was at ages (>41 years old). The highest rate of infection (33.9%) was found in September and the lowest (2.2%) in January. Similarly, the general infection frequency of the parasite G. lamblia was 3.9% with the highest rate at ages 1 - 20 years old and the lowest rate was 7.3% for >50 years old. The monthly, highest rate of infection (5.2%) was in August and least (2.2%) in January (2.2%). The frequency of total protein (TPD) in the blood relevant to the presence of parasite E. histolytica and G. lamblia was 4.6% and 1%, respectively. It is concluded that the above two parasites are the most common gastrointestinal parasite in Iraq whose pathogenesis to be which is likely to escalate during the summer seasons and at low hygienic services environment. There has been an irrelevance neither to anemia nor total protein deficiency. It is recommended that Ministry of Health in Iraq should not share the global idea of defining the giardiasis as a neglected disease.

Assemblages of <i>Giardia duodenalis</i>Isolates from Dogs by Amplification and Restriction of tpi and <i>β</i>-Giardin Genes and Sequencing of SSU rDNA Gene

Giardia duodenalis exhibits seven assemblages (A-G) that are distributed in different hosts. The A and B assemblages are commonly found in humans and several mammals, while C and D assemblages are typically found in dogs. The purpose of this study was to determine the assemblage of Giardia duodenalis present in the stool samples of ten canines using an assay based on PCR amplification, restriction analysis, and sequencing of the small subunit ribosomal DNA (SSU-rDNA), β-giardin, and triosephosphate isomerase (tpi) genes in order to establish the similarities or differences between the assemblage obtained with each gene. The results indicated that all positive isolates belonged to assemblage A, and specifically to the sub-assemblage A-I. A comparison of the SSU-rDNA gene sequence revealed the presence of three subgroups of assemblage A. These findings highlight the importance of canine transmission of Giardia in Mexico and its genetic plasticity. They also establish a method for additional and more molecularly extensive epidemiological studies to improve sanitation and hygiene in the most affected areas.

Role of Soluble Factors in Immune and Normal Sera in the Control of Giardiasis Due to Giardia Iamblia

Hyperimmune sera (HIS), raised against crude giardia antigen, on in vitro interaction, caused more agglutination of Giardia lamblia trophozoites. Heat inactivated HIS possessed a comparable agglutinating activity as the non-inactivated controls. Non-inactivated normal (unimmunized) serum caused immobilization of Giardia trophozoite, which was checked on heat inactivation. Antibodies in immune sera are mainly responsible for agglutination, whereas the heat labile non-immune components control the mobility of the intestinal parasite.

Prevalence of Giardia duodenalis among dogs seized by the Center for Control of Zoonoses (CCZ) of the city of Lages, Santa Catarina, Brazil

The knowledge of the epidemiology of parasitic infections in stray and domestic animals, especially of its incidence and prevalence, is fundamental to adopting effective prophylactic measures. Stray dogs play an important role in environmental contamination favoring the transmission cycle of zoonotic agents. Among the parasitic infections that affect humans, Giardia duodenalis is the most common intestinal protozoa and was designated as a re-emerging infectious disease. This study aimed to determine the prevalence of G. duodenalis in dogs siezed by the Center for Control of Zoonoses (CCZ) of the city of Lages, Santa Catarina, Brazil using two diagnostic techniques. In 357 stool samples analysed, the prevalence of G. duodenalis cysts was 5.3% (19/357) and 4.8% (17/357) detected by floatation and sedimentation techniques, respecttively. No correspondence between gender and age was found among the methods used for analyzing the infected dogs in this study. Our data suggested that two diagnostic techniques should be used in a complementary way to ensure that false negatives are not neglected.

Infectious causation of chronic disease:Examining the relationship between Giardia lamblia infection and irritable bowel syndrome

AIM: To evaluate whether a higher prevalence of Giardia lamblia infection is associated with an increase in irritable bowel syndrome (IBS) prescriptions at the county level in Michigan. METHODS: The Michigan Disease Surveillance System (MDSS) was used to ascertain both the numbers of Giardia lamblia infections as well as the total number of foodborne illnesses per population by county in Michigan during 2005. This was compared with Blue Cross Blue Shield (BCBS) of Michigan numbers of drug prescriptions for IBS per one thousand members per county in 2005. These data were also analyzed for associations with per capita income by county and the number of refugees entering each county in 2005. RESULTS: There were a total of 786 confirmed cases of Giardia lamblia reported to MDSS in 2005. During the same time period, the number of prescriptions for IBS varied from 0.5 per 1000 members up to 6.0 per 1000 members per month. There was no trend towards higher numbers of IBS prescriptions in the counties with more Giardia lamblia infections. Per capita income was not associated with either IBS prescriptions or Giardiasis. There was a significant linear association between the number of refugees entering each county, and the number of Giardia lamblia cases per 100 000 population. CONCLUSION: In this ecological study, there was no association found between BCBS prescriptions for IBS and Giardia lamblia infections in Michigan counties. Our findings may have been influenced by the disparate number of refugees admitted per county.

Identification, characteristic and phylogenetic analysis of type II DNA topoisomerase gene in Giardia lamblia

The genes encoding type II DNA topoisomerases were investigated in Giardia lamblia genome, and a type IIA gene,GlTop 2 was identified. It is a single copy gene with a 4476 bp long ORF without intron. The deduced amino acid sequence shows strong homology to eukaryotic DNA Top 2. However, some distortions were found, such as six insertions in the ATPase domain and the central domain, a -100 aa longer central domain; a ~200 aa shorter C-terminal domain containing rich charged residues. These features revealed by comparing with Top 2 of the host, human, might be helpful in exploiting drug selectivity for antigiardial therapy. Phylogenetic analysis of eukaryotic enzymes showed that kinetoplastids, plants, fungi, and animals were monophyletic groups, and the animal and fungi lineages shared a more recent common ancestor than either did with the plant lineage; microsporidia grouped with fungi. However, unlike many previous phylogenetic analyses, the ''amitochondriate'' G. lamblia was not the earliest branch but diverged after mitochondriate kinetoplastids in our trees. Both the finding of typical eukaryotic type IIA topoisomerase and the phylogenetic analysis suggest G, lamblia is not possibly as primitive as was regarded before and might diverge after the acquisition of mitochondria. This is consistent with the recent discovery of mitochondrial remnant organelles in G. lamblia.