Giardiasis Patients Research Articles

EFFECTS OF ALTERNATIVE NATURAL AND NANOTHERAPIES VERSUS METRONIDAZOLE ON GIARDIASIS INFECTED HAMSTERS

Giardia lamblia is one the most common human zoonotic protozoan affecting about 200 million peopleannually, but recurrent symptoms and drug resistance were reported in giardiasis patients. The presentstudy assessed the therapeutic effect of Tomex® and probiotics against giardiasis as well as evaluatedefficacy of chitosan nanoparticles and metronidazole. Sixty healthy male hamsters were dividedinto 6 groups. Each group consists of 10 hamsters as following: GI non-infected hamsters, GII infectedcontrol, GIII, GIV, GV & GVI infected with G. lamblia and treated with tomex, probiotics, chitosannanoparticles (CsNPs), and metronidazole (MTZ) respectively. Hamsters were orally infected with10,000 Giardia cysts. After 3 weeks stool samples were examined for Giardia samples and contentsfor intestinal histopathology and electron microscopy of Giardia trophozoite ultrastructure. The highestreduction in Giardia cyst and trophozoite were in hamsters received MTZ but with natural alternativetherapies chitosan nanoparticles showed a therapeutic effect with highest reduction in cyst andtrophozoite counts (80.59% & 76.55%) respectively and best recovery in intestinal pathology. MTZgave higher reduction rates (91.04% & 90.75%) respectively with normal intestinal histopathology.

Study of lipid profile alteration in the patients infected with Giardia lamblia and compare the results with healthy individuals

This study was include collection of 40 stool samples from people were suffer from steatorrhea and the genral stool examination for these stool samples showed the presence of the intestinal parasite Giardia lamblia the etiologic agent of giardiasis . Blood samples were collected from 60 individual 40 of them were the infected with Giardia lamblia patients and the other 20 were healthy and used as control ,then the serum was obtained from all blood samples in the field of the study and the estimation of total (Cholesterol ,Triglyceride ,High density lipoproteins, Low density lipoproteins and Very low density lipoproteins )was done . the result showed that the cholesterol levels were decreased in giardiasis patients while the other types of lipids were normal in the same patients because Giardia lamblia consumed the cholesterol of the host in the bio synthesis of the cell . because the parasite is unable to synthesize cholesterol by itself.

Giardiasis and Subsequent Irritable Bowel Syndrome: A Longitudinal Cohort Study Using Health Insurance Data

Giardia intestinalis is the most commonly reported human intestinal parasite in the United States. Increased incidence of chronic gastrointestinal complaints has been reported after some giardiasis outbreaks. We examined the relationship between giardiasis diagnosis and irritable bowel syndrome (IBS) diagnosis. We used the 2006-2010 MarketScan commercial insurance database. Persons with at least 1 giardiasis diagnosis were individually matched on age group, sex, and enrollment length in months to 5 persons without a giardiasis diagnosis. Persons diagnosed with IBS before the date of study entry were excluded. We calculated crude incidence rates (IRs) and developed Cox proportional hazards models. The matched cohort included 3935 persons with giardiasis and 19663 persons without giardiasis. One-year incidence of IBS was higher in persons with giardiasis (IR = 37.7/1000 person-years vs 4.4/1000 person-years). The unadjusted hazard ratio was 4.8 (95% confidence interval [CI] = 3.6-6.4), attenuated slightly to 3.9 (95% CI = 2.9-5.4) after adjusting for anxiety, depression, and healthcare utilization. In a large insurance database, individuals diagnosed with giardiasis were more likely to have a subsequent IBS diagnosis, despite accounting for confounders. Future research on risk factors for IBS among giardiasis patients and the pathophysiology of postinfectious IBS is needed.

Prevalence of Giardia duodenalis assemblages and sub-assemblages in symptomatic patients from Damascus city and its suburbs

Giardia duodenalis is one of the most important human enteric parasites worldwide and is endemic throughout the world with a vast range of mammalian hosts. However, there is limited information on the prevalent genetic variability of G. duodenalis in Syria. This study aimed to evaluate the predominance of G. duodenalis assemblages/sub-assemblages causing humans infection in the city of Damascus and its suburbs. 40 symptomatic giardiasis patients were recruited in this study. Fecal samples were genotyped using PCR/RFLP assay targeting the β-giardin and glutamate dehydrogenase (gdh) genes. HaeIII, BspL1 and RsaI restriction enzymes were used to differentiate between G. duodenalis assemblages/sub-assemblages. Our data showed that 65% of isolates were of assemblage A; 45% belonged to sub-assemblage AII and 20% to sub-assemblage AI. Assemblage B was detected in 27.5% of isolates; 12.5% fit in sub-assemblage BIV, 5% fit in sub-assemblage BIII and 10.5% fit in Discordant genotype BIII/BIV. Mixed genotypes (AII+BIII and AI+BIV) were identified in 3 isolates (7.5%). Significant correlation was found between Giardia AII sub-assemblage and weight loss symptom (P-value=0.05) as well as between contact with domestic animals (cats, P-value=0.027). Moreover, a significant correlation was found between sub-assemblage AI and livestock breeding (P-value=0.000). In conclusion genotyping of human Giardia duodenalis isolates suggests anthroponotic transmission for the route of infection in Damascus and its suburbs. Further studies are needed to screen a wide geographic areas in Syria and to estimate the prevalence of G. duodenalis infection in our population.

Ghrelin and melatonin as biomarkers in patients with giardiasis

ABSTRACTGiardia is the most frequently reported intestinal parasite worldwide. The aim of this study was to investigate the ghrelin, melatonin, glucose and cholesterol concentration in male patients infected with Giardia lamblia. We enrolled 66 patients with Giardiasis and the control groups consisted of healthy subjects (n = 30). The results demonstrated that there was a significant decrease (P < 0.05) in ghrelin levels, while the melatonin, glucose and cholesterol levels were significantly increased (P < 0.05) in giardiasis patients as compared to the healthy group. The obtained results suggest that ghrelin and melatonin could serve as biomarkers in patients infected with G. lamblia.

Survey on efficacy of chloroformic extract of Artemisia annua against Giardia lamblia trophozoite and cyst in vitro.

Giardiasis is a parasitic cosmopolitan disease that the rate of infection in developing countries is considerable. This infection directly is associated with poor hygienic conditions, poor water quality control, and overcrowding. Reinfection and drug resistance are two major problems in endemic areas. Recently, researchers are concentrating on herbal drugs as a proper solution. Therefore, the objective of the present study was to survey on efficacy of chloroformic extract of Artemisia annua against Giardia lamblia trophozoite and cyst in vitro. G. lamblia cysts were prepared from faces of giardiasis patients from different hospitals of Mazandaran Medical University. Four concentrations (1, 10, 50 and 100mg/ml) of chloroformic extract of A. annua were utilized for 1, 5, 30, 60 and 180min. Viability of G. lamblia cysts was confirmed by 0.1% Eosin staining. Cyst and trophozoite contact (intermix) of G. lamblia with extract of A. annua with variant concentrations (1, 10, 50 and 100mg/ml) after 1 and 180min caused following cyst and trophozoite elimination rates: (67, 69, 71 and 73%), (65, 67, 67 and 72%), (94, 96, 97 and 99%) and (100, 100, 100 and 100%), respectively. Authors from the current investigation draw a conclusion that chloroformic extract of A. annua has the ability to eliminate G. lamblia cysts and trophozoites in vitro.

Elongation factor 1-alpha is released into the culture medium during growth of Giardia intestinalis trophozoites

The molecular pathogenesis of the intestinal parasite Giardia intestinalis is still not fully understood but excretory–secretory products have been suggested to be important during host–parasite interactions. Here we used SDS–PAGE gels and MALDI–TOF analysis to identify proteins released by Giardia trophozoites during in vitro growth. Serum proteins (mainly bovine serum albumin) in the growth medium, bind to the parasite surface and they are continuously released, which interfere with parasite secretome characterization. However, we identified two released Giardia proteins: elongation factor-1 alpha (EF-1α) and a 58kDa protein, identified as arginine deiminase (ADI). This is the first description of EF-1α as a released/secreted Giardia protein, whereas ADI has been identified in an earlier secretome study. Two genes encoding EF-1α were detected in the Giardia WB genome 35kbp apart with almost identical coding sequences but with different promoter and 3′ regions. Promoter luciferase-fusions showed that both genes are transcribed in trophozoites. The EF-1α protein localizes to the nuclear region in trophozoites but it relocalizes to the cytoplasm during host–cell interaction. Recombinant EF-1α is recognized by serum from giardiasis patients. Our results suggest that released EF-1α protein can be important during Giardia infections.

Symptoms in giardiasis patients

Background: Giardiasis is the most commonly reported gastrointestinal pathogen in United States. Most patients only have small bowel biopsy performed if there are symptoms of malabsorption. It has however been noted that not all patients with giardiasis have malabsorption related symptoms.

Comportamiento de la infección experimental por aislamientos colombianos de Giardia duodenalis en el modelo animal del gerbo (Meriones unguiculatus).

Natural and experimental Giardia infections have been reported from bovines, equines, goats, canines, felines and rodents such as mice, rats and gerbils. The latter have provided successful animal models for Giardia duodenalis and Giardia muris experimental infections. The gerbil model was used to establish the pattern of infection of Colombian Giardia human isolates. Giardia cysts were obtained from stool specimens of symptomatic giardiasis patients by means of sucrose-percoll gradients. Animal inoculation was performed by gastric intubation and injection with 5 x 10(3) Giardia cysts. The course of infection was established by counting cysts every day and trophozoites weekly throughout a period of 30 days. The pattern of cyst excretion was found to be intermittent. Cysts were released during the second and third weeks of infection but not during the first or fourth weeks. The mean minimal number of cysts released per 2-hr collection period was 79 and the mean maximum number was 17,943. Colonization of the small intestine by trophozoites was observed with a mean number ranging from 15,000 to 6,577,778 trophozoites/ml. Gerbils inoculated with G. duodenalis isolates obtained from geographical areas outside Colombia resolved the infection between 86 and 114 days after infection, whereas gerbils infected with Colombian G. duodenalis isolates resolved the infection at 30 days. The gerbil proved to be a good animal model for experimental infection with Colombian isolates of G. duodenalis. Experimental Giardia infection of gerbils permit a sufficient yield of cysts and trophozoites to be used as antigens for the immunization of other animals and to obtain Giardia antibodies that could be used for Giardia antigen detection assays in stool specimens.

Secretory antibodies against Giardia intestinalis in lactating Nicaraguan women

Secretory IgA (sIgA) antibodies are important in the host defence against the intestinal protozoan parasite Giardia intestinalis. However, few antigens have been identified. In this study 100 milk and saliva samples from lactating women, living in an endemic region (León, Nicaragua), were screened for the presence of antibodies against G. intestinalis. Most milk and saliva samples contained anti-Giardia antibodies (59% and 52%, respectively), with a mean sIgA content 50 times higher in milk than in saliva. The positive samples reacted with trophozoite membrane, flagella and cytoplasmic antigens. Western blot analysis showed that milk and saliva anti-Giardia sIgA recognized up to 16 different Giardia proteins in the molecular weight region 20-165 kDa. Two-dimensional Western blotting showed that the major immunoreactive proteins were the same as the immunoreactive proteins identified by serum from acute giardiasis patients in a non-endemic country. The major difference was a stronger reactivity against the variant surface proteins (VSPs) in the milk samples. Milk sIgAs also recognized recombinant Giardia proteins such as alpha-1 giardin, ornithine carbamoyl transferase, VSP-4EX, arginine deaminase and alpha-enolase. These antigens will be important targets in the development of new immunodiagnostic tools and vaccines.

Characterisation of alpha-1 giardin: an immunodominant Giardia lamblia annexin with glycosaminoglycan-binding activity

Alpha-1 giardin is an immunodominant protein in the intestinal protozoan parasite Giardia lamblia. The Triage ® parasite panel, used to detect copro-antigens in stool from giardiasis patients, reacts with an epitope between amino acids 160 and 200 in alpha-1 giardin. This region of the protein is also highly immunogenic during human infections. Alpha-1 giardin is related to annexins and like many other annexins it was shown to be plasma membrane associated. Immunoelectron and immunofluorescence microscopy revealed that some alpha-1 giardin are displayed on the surface of recently excysted cells. Recombinant alpha-1 giardin displayed a Ca 2+-dependent binding to glycosaminoglycans (GAGs), in particular heparan sulphate, a common GAG in the intestinal tract. Recombinant alpha-1 giardin bound to thin sections of human small intestine, a binding which could be inhibited by adding increasing concentrations of sulphated sugars. A surface associated trypsin activated Giardia lectin (taglin) has been suggested to be important for G. lamblia attachment. In this study we show that a monoclonal antibody that inhibits taglin recognises alpha-1 and alpha-2 giardin. Thus, alpha-1 giardin is a highly immunoreactive GAG-binding protein, which may play a key role in the parasite–host interaction. Our results further show a conserved function of annexins from lower to higher eukaryotes.

Secretory Immune Response to Membrane Antigens duringGiardia lambliaInfection in Humans

The secretory immune response in humans infected with Giardia lamblia was studied by using saliva samples and a membrane-rich protein fraction. The membrane fraction, studied by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, showed 24 antigen bands, ranging from 170 to 14 kDa. Saliva samples from giardiasis patients showed a heterogeneous response against the membrane fraction when they were assayed by immunoblotting. Among the antigens recognized by patient saliva samples, those of 170, 105, 92, 66, 32, 29, and 14 kDa stood out. These antigens were not recognized by saliva samples from healthy individuals. They may be of importance in future studies of protection from or diagnosis of G. lamblia infections.

Classification of subgroups of Giardia lamblia based upon ribosomal RNA gene sequence using the polymerase chain reaction

A sensitive and specific polymerase chain reaction-based assay has been developed to detect and analyze polymorphism in the Giardia lamblia 18S ribosomal RNA gene. Efficient amplification required the inclusion of cosolvents (glycerol and dimethyl sulfoxide) in the reaction. Following the optimization of conditions for amplification and subsequent hybridization of amplified product with radiolabeled oligonucleotide probe, a detection limit of less than one organism's worth of DNA was achieved. Thirty-five different G. lamblia strains obtained from various human and animal host types and geographic locations were analyzed by this method. The strains could be divided into 3 groups on the basis of defined nucleotide substitutions within the 183-bp amplified DNA fragment of the 18S ribosomal RNA gene. The groupings based upon the 18S ribosomal RNA gene sequence correlated with groupings previously assigned based upon patterns of surface antigens and restriction enzyme analysis. Analysis of the G. lamblia 18S ribosomal RNA gene sequences present in fecal specimens obtained from giardiasis patients revealed the presence of the different sequence types in these specimens. Some specimens contained more than one sequence type. The identification of subgroups of G. lamblia may facilitate studies of virulence, infectivity, and the epidemiology of giardia infection.

Serum antibody response in children with Giardia lamblia infection and identification of an immunodominant 57‐kilodalton antigen

Giardia lamblia antigens which react with sera from children with G. lamblia infection were investigated by sodium-dodecyl polyacrylamide gel electrophoresis and immunoblotting. Serum IgG, IgM and IgA response to the antigens were immunochemically characterized. Serum antibodies from all giardiasis patients, but none of the controls, was found to react with a 57-kilodalton antigen. The 57 kDa antigen elicited IgG and IgA but not IgM antibodies. The protein nature of the 57 kDa antigen was demonstrated by loss of antibody recognition after trypsin treatment of G. lamblia trophozoites. Subcellular fractionation of G. lamblia trophozoites followed by SDS-PAGE and immunoblotting showed that the 57 kDa antigen was probably not a component of the cytoskeleton.

Identification of Giardia lamblia-specific antigens in infected human and gerbil feces by western immunoblotting

Western immunoblot analysis of aqueous extracts of feces obtained from five giardiasis patients and from experimentally infected gerbils (Meriones unguiculatus) with rabbit antiserum to Giardia lamblia cysts has revealed antigens of three molecular weight groups. A stepladderlike, evenly-spaced set of strongly reactive antigens (darkest at a molecular weight [m.w.] of 55,000 to 70,000) appeared in the gerbil feces from day 4 (first experiment) or day 2 (second experiment) and lasted to about day 7 but disappeared completely by day 8 and did not reappear later. These antigenic bands were seen in gerbils infected with two isolates of G. lamblia. These bands were not revealed when antiserum to trophozoites was used as the probe, nor were they evident in specimens from the patients or in a preparation of sonicated cysts. A second group of antigens, represented by two to three low-m.w. bands of approximately 15,000 to 20,000, was evident in both the blots of gerbil feces after approximately day 8 and the specimens from the giardiasis patients. The third group of antigens revealed by blotting experiments was a high-m.w. band (approximately 110,000) which appeared on a number of days (beginning of day 8 of gerbil infection), but this band was not seen in the human specimens. A clear band corresponding to the previously reported GSA-65 antigen was not seen in either the gerbil or the human samples. Some low- and high-m.w. bands were also detected by antitrophozoite serum in the gerbil samples, but these were weak and unimpressive compared with those visualized using anticyst serum. A monoclonal antibody-based antigen capture enzyme-linked immunosorbent assay revealed that Giardia spp.-specific stool antigen rose suddenly at day 3 of gerbil infection, at the time when fecal cyst numbers began to rise rapidly.

Antigenic variation of Giardia lamblia in the feces of Mongolian gerbils

Enzyme-linked immunoelectrotransfer blot was used to study variations in Giardia lamblia antigens in extracts of feces from infected Mongolian gerbils. A 65-kilodalton antigen was found in feces that contained strain WB (ATCC 30957) cysts and in axenic culture of strains WB and CDC:0284:1 that contained trophozoites. The 65-kilodalton antigen from trophozoites of both strains was membrane associated. A 70-kilodalton antigen was found in feces that contained strain CDC:0284:1 cysts. It was persistent in 16 fecal collections and may be strain specific. Similar variations in antigens may occur in human feces. Coproimmunodiagnostic assays that use monoclonal antibodies will have to include all varieties of G. lamblia antigens present in the feces of giardiasis patients.

In vitro excystation and subsequent axenic growth of Giardia lamblia

Six fresh Giardia lamblia strains were isolated by in vitro excystation and subsequent culturing of excysted parasites in bile-supplemented BI-S-33 medium. The cysts passed in faeces appeared to differ in structure when observed using differential interference contrast microscopy. Sometimes the enclosed organisms were closely applied to the cyst wall; in most stool specimens, however, the parasites were separated from the cyst wall by a space. Cysts with parasites closely applied to the wall were the most viable type, with excystation rates up to 90%. Cysts with organisms detached from the wall displayed low excystation rates, 0–15%. During exposure to the induction solution of hydrochloric acid (pH 2), parasites initially closely applied to the cyst walls completely detached, and after transfer into culture medium vigorous flexing movements of the organisms were observed. Stool samples from 42 symptomatic and asymptomatic giardiasis patients were examined; in 26 of the samples parasites hatched, and 6 new strains were established in axenic culture.

Heterogeneity in the responses of clones of Giardia intestinalis to anti-giardial drugs

Clones of two stocks of Giardia intestinalis have been tested by the [ 3H]thymidine uptake assay to determine their sensitivity to metronidazole, tinidazole, furazolidone and quinacrine. Each stock was not homogeneous with respect to drug sensitivity but was composed of different populations of organisms. Doubling times of lines derived from clones of a stock did not vary significantly. These findings may, in part, account for treatment failures in human giardiasis patients.

Physical and chemical characterization of a Giardia lamblia-specific antigen useful in the coprodiagnosis of giardiasis.

We recently reported the isolation and identification of a Giardia lamblia-specific antigen (GSA 65) that is shed in the stool of giardiasis patients. In the present study, this antigen was affinity purified from sonic extracts of axenically cultured G. lamblia trophozoites and characterized to better understand its biological function and its potential usefulness in the design of coprodiagnostic assays for giardiasis. GSA 65 was resistant to proteolytic digestion with trypsin, chymotrypsin, and protease but was sensitive to treatment with NaIO4 as assessed by Western blotting. This antigen was also stable during prolonged storage at 4 and -20 degrees C in 10% Formalin or distilled H2O as assessed by counterimmunoelectrophoresis. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis and isoelectric focusing gel banding patterns, in conjunction with protein and carbohydrate assays and lectin binding studies, confirmed that this antigen is a highly glycosylated glycoprotein. The resistance of GSA 65 to proteolytic degradation, together with previous immunofluorescence data that indicate the antigen is an integral part of the G. lamblia cyst wall, suggests that this molecule may play a role in maintaining the integrity of the cyst in vivo. The ability of GSA 65 to maintain its antigenic structure under a wide variety of conditions makes it an ideal antigen around which to design sensitive immunodiagnostic assays for giardiasis.

Isolation and identification of a Giardia lamblia-specific stool antigen (GSA 65) useful in coprodiagnosis of giardiasis.

A Giardia lamblia-specific antigen (GSA 65) was isolated from stools of G. lamblia-positive patients by crossed- and line-immunoelectrophoresis and counterimmunoelectrophoresis (CIE) in agarose by using rabbit antiserum prepared against G. lamblia cysts. CIE with rabbit anti-GSA 65 monospecific antiserum revealed that GSA 65 was present in aqueous stool eluates of giardiasis patients and in cysts and trophozoites of the parasite. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of immunoaffinity-purified antigen followed by Western blotting showed that the molecular weight of this molecule was about 65,000. GSA 65 was detectable by CIE in stool eluates of 36 of 40 giardiasis patients but not in eluates of 10 G. lamblia-negative asymptomatic controls. GSA 65 was detected in stool eluates of 2 of 18 individuals with chronic diarrhea who were negative for parasites by microscopic examination. Cross-specificity studies with other genera of parasitic protozoa performed by using CIE and immunofluorescence indicated that GSA 65 was present only in strains of G. lamblia. Based on these findings, GSA 65 may prove to have an important application in the design of sensitive diagnostic tests for giardiasis.