Atorvastatin: In-Vivo Synergy with Metronidazole as Anti-Blastocystis Therapy.

Blastocystis is an enteric Straminopile in tropical, subtropical and developing countries. Metronidazole has been a chemotheraputic for blastocystosis. Failures in its regimens were reported and necessitate new studies searching for alternative therapeutic agents. Aim of current study is to investigate potential effects of Atorvastatin (AVA) compared to the conventional chemotherapeutic MTZ in experimentally Blastocystis-infected mice. Anti-Blastocystis efficacy of AVA was evaluated parasitologically, histopathologically and by transmission electron microscopy using MTZ (10 mg/kg) as a control. Therapeutic efficacy of AVA was apparently dose-dependent. Regimens of AVA (20 and 40 mg/kg) proved effective against Blastocystis infections with high reduction in Blastocystis shedding (93.4-97.9%) compared to MTZ (79.3%). The highest reductions (98.1% and 99.4%) were recorded in groups of combination treatments AVA 20-40 mg/kg and MTZ 10 mg/kg. Blastocystis was nearly eradicated by the 20th day post infection. Genotype analysis revealed that genotype I was most susceptible, genotype III was less. Histopathologic and ultrastructural studies revealed apoptotic changes in Blastocystis and significant improvement of intestinal histopathological changes more remarkable in combinational therapy groups. Thus, the present study offers AVA as a potential candidate for Blastocystis therapy combined with MTZ.

Genotypic Identification of Cystoisospora in Immunocompromised Patients Using Tm-Variation Analysis.

Cystoisospora is responsible for morbidity in immunocompromised patients. PCR is sensitive for diagnosing Cystoisospora; however, it needs reevaluation for differential molecular diagnosis of cystoisosporiasis. We aimed at evaluating melting curve analysis (MCA) after real-time PCR (qPCR) in diagnosis and genotyping of Cystoisospora as an alternative to conventional PCR. We included 293 diarrheic stool samples of patients attending the Department of Clinical Oncology and Nuclear Medicine of Cairo University Hospitals, Egypt. Samples were subjected to microscopy, nested PCR (nPCR), and qPCR targeting the internal transcribed spacer 2 region (ITS2) of the ribosomal RNA (r RNA) gene followed by melting temperatures (Tms) analysis and comparing the results to PCR-RFLP banding patterns. Using microscopy and ITS2-nPCR, 3.1% and 5.8% of cases were Cystoisospora positive, respectively, while 10.9% were positive using qPCR. Genotyping of Cystoisospora by qPCR-MCA revealed 2 genotypes. These genotypes matched with 2 distinct melting peaks with specified Tms at 85.8°C and 88.6°C, which indicated genetic variation among Cystoisospora isolates in Egypt. Genotype II proved to be more prevalent (65.6%). HIV-related Kaposi sarcoma and leukemic patients harbored both genotypes with a tendency to genotype II. Genotype I was more prevalent in lymphomas and mammary gland tumors while colorectal and hepatocellular tumors harbored genotype II suggesting that this genotype might be responsible for the development of cystoisosporiasis in immunocompromised patients. Direct reliable identification and differentiation of Cystoisospora species could be established using qPCR-Tms analysis which is useful for rapid detection and screening of Cystoisospora genotypes principally in high risk groups.

P selectins and immunological profiles in HCV and Schistosoma mansoni induced chronic liver disease.

BACKGROUND: Hepatitis C virus (HCV) and Schistosoma mansoni are major causes of chronic liver disease (CLD) in which immune alteration is common. Recent studies suggested that certain platelets and lymphocytes activation markers may have an impact on progression of CLD. This study aimed to evaluate the potential of platelets and lymphocytes activation molecules expression on the pathogenesis of CLD in distinct or concomitant chronic HCV and schistosomiasis mansoni infections. METHODS: The study populations were divided into group-I: patients with chronic schistosomiasis mansoni, group-II: HCV patients without cirrhosis, group-III: patients with combined liver diseases without cirrhosis, group-IV: patients with chronic HCV and liver cirrhosis and group-V: Age and sex matched healthy individuals as normal controls. All groups were subjected to full clinical evaluation, ELISA anti-HCV antibodies screening, parasitological examination for diagnosing S. mansoni and flow cytometry for lymphocyte (CD3, CD4, CD8, CD19, CD22, & CD56) and platelets activation (CD41, CD42 & CD62P (P- selectins)) markers. RESULTS: The platelet count was significantly decreased in HCV and/or S. mansoni patients. The total T-lymphocytes and T-helper cells were significantly reduced, while T-cytotoxics were increased. The patients possessed a significantly higher platelets activation marker; CD62P (P-selectins) and higher mean fluorescent intensity (MFI) positivity. There were considerable correlations between platelets count and both of CD62P and MFI. CONCLUSION: Our Findings suggest an increased expression of certain platelets and lymphocytes activation markers in chronic HCV and S. mansoni induced CLD that may have a role in disease progression.

Molecular identification of giardia intestinalis in patients with dyspepsia.

BACKGROUND/AIMS: Giardia intestinalis triggers symptoms of functional dyspepsia. The aim of this study was to distinguish genotypes of G. intestinalis isolated from dyspeptic patients to evaluate their correlation with dyspeptic symptoms. METHODS: In total, 120 dyspeptic subjects were investigated by upper endoscopy, including gastric and duodenal biopsies for histopathological examination, and parasitological examination of their stools and duodenal aspirates was performed. The patients were classified into five groups: group I (G. intestinalis) included 19 patients, group II (Helicobacter pylori) included 36 patients, group III (coeliac disease) included 3 patients, group IV (mixed G. intestinalis and H. pylori infection) included 4 patients, and group V (unexplained aetiology) included 58 patients. Genotyping of G. intestinalis was performed for groups I and IV using PCR-RFLP. The urease test was performed for H. pylori. Serum anti-gliadin, anti-endomysial and anti-transglutaminase antibody estimation was performed for the diagnosis of coeliac disease. RESULTS: Genotype A of G. intestinalis was detected in the stool samples of 68.42% (13/19) and the duodenal aspirates of 42.1% (8/19) of dyspeptic patients harbouring the parasite. Genotype B was detected in 31.58% (6/19) of cases in stool samples and in 3 cases in duodenal aspirates. CONCLUSIONS: H. pylori, G. intestinalis and coeliac disease are common causes of dyspepsia. G. intestinalis genotype A demonstrated a greater association with dyspeptic symptoms.

The effect of praziquantel and Carica papaya seeds on Hymenolepis nana infection in mice using scanning electron microscope.

Hymenolepis nana (H. nana) is the most common tapeworm infection worldwide. It is more prevalent in warm climates where sanitation is poor, particularly among children. The effect and mechanism of action of praziquantel (PZQ), given at a dose of 25-mg/kg BW, and Carica papaya dried seed crude aqueous extract (CAE), given at a dose of 1.2-g/kg BW, were assessed on H. nana worms in experimentally infected mice. Tegumental changes were studied using the scanning electron microscope (SEM) and different parasitological parameters were observed. Each group of infected mice was divided into two subgroups. The first subgroup received either treatment before the 4th day after infection to investigate their effects on the cysticercoid stage. The other subgroup received treatments after the development of the adult stage, confirmed by eggs detection in stool. Both PZQ and C. papaya dried seed CAE resulted in a significant reduction of worm burden, total egg output and viable egg count. Marked tegumental changes were evident in adult worms treated with either treatment including shrinkage of the scolex and neck region with rostellar edema and complete loss of its hooks. However, all previous effects were exerted more rapidly in the case of PZQ treatment. They both significantly reduced cysticercoid stage size. Nevertheless, C. papaya outstand PZQ in having a deforming effect on adults arising from treated cysticercoids. It was concluded that C. papaya has significant anti-cestodal properties that enable its seed extract to be a very effective alternative to PZQ against H. nana.

Therapeutic potential of myrrh and ivermectin against experimental Trichinella spiralis infection in mice.

Trichinosis is a parasitic zoonosis caused by the nematode Trichinella spiralis. Anthelmintics are used to eliminate intestinal adults as well as tissue-migrating and encysted larvae. This study aimed to investigate the effects of ivermectin and myrrh obtained from the aloe-gum resin of Commiphora molmol on experimental trichinosis. Ninety albino mice were orally infected with 300 T. spiralis larvae. Drugs were tested against adult worms at day 0 and day 5 and against encysted larvae on day 15 and day 35 post-infection (PI). Mature worms and encysted larvae were counted in addition to histopathological examination of muscle specimens. Serum aspartate aminotransferase (AST), alanine aminotransferase (ALT), total protein, albumin, globulin, urea, and creatinine values were estimated. Significant reductions in mean worm numbers were detected in ivermectin treated mice at day 0 and day 5 PI achieving efficacies of 98.5% and 80.0%, while efficacies of myrrh in treated mice were 80.7% and 51.5%, respectively. At days 15 and 35 post-infection, ivermectin induced significant reduction in encysted larval counts achieving efficacies of 76.5% and 54.0%, respectively, while myrrh efficacies were 76.6% and 35.0%, respectively. AST, ALT, urea, and creatinine levels were reduced, while total proteins were increased in response to both treatments compared to their values in the infected non-treated mice. Ivermectin use for controlling T. spiralis could be continued. Myrrh was effective and could be a promising drug against the Egyptian strains of T. spiralis with results nearly comparable to ivermectin.

Antigen Recognition Patterns of Intestinal Capillariasis Using Immunoblot-Based Serodiagnosis.

BACKGROUND: Diagnosis of intestinal capillariasis depending on microscopic detection of parasitic stages is of low sensitivity, especially in cases with low worm burden. There is a necessity to develop sensitive and specific diagnostic tools of capillariasis for early treatment to avoid complications. Western blot (WB) technique showed promising results for antigen recognition patterns in several parasitic infections. AIM OF THE STUDY: This study is directed to identify and evaluate relevant proteins of intestinal capillariasis crude worm antigens using WB immunodiagnosis. MATERIALS AND METHODS: Capillaria crude worm antigens were extracted and analyzed using SDS-PAGE. Sixty serum samples belonging to 3 groups (20 individuals each) were included; Group I (shedding Capillaria in feces), Group II (infected with other parasites) and Group III (negative parasitological results). Reactivity of the resulting bands of Capillaria with serum samples was analyzed using WB technique. RESULTS: Thirty-two immunoreactive bands were detected in WB analysis representing recognition of proteins with molecular weights (MW) varying from 19 to 110 kDa. Immunodominant proteins of 23.5, 31, 36.5, 40.5 and 44 kDa were recognized, respectively, in 35%, 30%, 85%, 95% and 75% of sera from patients with confirmed capillariasis and in 30%, 25%, 35%, 25%, and 20% of sera from those infected with other parasitic infections. One serum sample from group III gave reaction with 31 kDa band. CONCLUSION: Immunodiagnosis of intestinal capillariasis using WB proved that 23.5, 31, 36.5, 40.5 and 44 kDa bands could be considered useful tools for diagnosis of capillariasis.

Molecular prevalence and estimated risk of cutaneous leishmaniasis in Libya.

BACKGROUND/PURPOSE: Cutanoeus leishmaniasis (CL) is an endemic disease in the Mediterranean area including Libya. The aim of the present study is to detect the prevalent Leishmania species obtained from smeared cutaneous lesions in addition to studying the diverse sociodemographic risk factors of the reported cases from different provinces of Libya. METHODS: A total of 250 archived microscopic slides from clinically suspected cases of CL attending the leishmaniasis clinic in the Dermatology Department, Tripoli Central Hospital, Tripoli, Libya, were microscopically examined. Leishmania-DNA was amplified using combined polymerase chain reaction (PCR) targeting kinetoplast-DNA (kDNA) and ribosomal internal transcribed spacer 1 (ITS1)-DNA with restriction fragment length polymorphism analysis for direct Leishmania species identification. RESULTS: Using kDNA and ITS1-PCR, 22.5% and 20% of cases were positive, respectively. Only 14.4% of cases were positive using microscopy. Nominating ITS1-PCR as the reference standard, kDNA-PCR assay was highly sensitive while microscopy was 100% specific but of limited sensitivity (72%) with a substantial agreement and an overall accuracy of 94.4%. Leishmania major and Leishmania tropica were the predominant species reported from the north-western provinces including Tripoli, Zintan, and Gharyan with their related subprovinces; Asabaa, Mizdan, Alkawasem, and Alorban. CL prevailed more among men and residents of rural areas. House wives and students were the most affected professions. Children were the least affected, while the middle-aged were the most affected age group. CONCLUSION: L. major and L. tropica are the predominant species in the north-western regions of Libya. ITS1-PCR-restriction fragment length polymorphism assay offered a sensitive, specific, and faster diagnostic method especially with negative parasitologic examination.

Nematodes ultrastructure: complex systems and processes.

Nematode worms are among the most ubiquitous organisms on earth. They include free-living forms as well as parasites of plants, insects, humans and other animals. Recently, there has been an explosion of interest in nematode biology, including the area of nematode ultrastructure. Nematodes are round with a body cavity. They have one way guts with a mouth at one end and an anus at the other. They have a pseudocoelom that is lined on one side with mesoderm and on the other side with endoderm. It appears that the cuticle is a very complex and evolutionarily plastic feature with important functions involving protection, body movement and maintaining shape. They only have longitudinal muscles so; they seem to thrash back and forth. While nematodes have digestive, reproductive, nervous and excretory systems, they do not have discrete circulatory or respiratory systems. Nematodes use chemosensory and mechanosensory neurons embedded in the cuticle to orient and respond to a wide range of environmental stimuli. Adults are made up of roughly 1000 somatic cells and hundreds of those cells are typically associated with the reproductive systems. Nematodes ultrastructure seeks to provide studies which enable their use as models for diverse biological processes including; human diseases, immunity, host-parasitic interactions and the expression of phylogenomics. The latter has, however, not been brought into a single inclusive entity. Consequently, in the current review we tried to provide a comprehensive approach to the current knowledge available for nematodes ultrastructures.

SOCIO-DEMOGRAPHIC RISK FACTORS OF SCHISTOSOMIASIS MANSONI IN PATIENTS WITH GASTROINTESTINAL SYMPTOMS: A SEROPREVALANCE STUDY IN EGYPT.

Limited data is available on the epidemiologic status of schistosomiasis mansoni in Egypt. The present work aimed to explore the seroepidemiological status of Schistosoma inansoni infection in Egypt by screening inhabitants of different Egyptian govemorates and its correla- tion with morbid symptoms and risk factors. Health questionnaires and indirect haemagglutination test (IHAT) were performed upon a cross-sectional study of 1788 individuals from 22 govemorates. Socio-demographic varjables included sex, age, residence and.canal water contact. A multivariate regressi6n model was used to assess associations betWeen S.mansoni infection and socio-demographic variables; S.mansoni significant titre ≥ 1:160 was detected in 43% of samples. S. mansoni showed the highest prevalence in Al-Fayoum (15.2%), Kafr EI-Sheikh (11.2%) then Assiut (10.9%) while the least positiveresults were from Matrouh (0.2%). This may be the first indication to emerging foci in Cairo, Luxor, Aswan, Suez, Port Said and the Red Sea Govemorates. Anti-S.mansoni antibodies were least de- tected at 1 1-2Oys while they were the highest at 41-SOys, the highest titres (1/1280) were achieved by the age group 31 -4Oys.Male gender was a risk factor as 48.2% of males were IHAT +ve. Contacting canal water tends to be advantageous for schistosomiasis mansoni as 72.6% had a histoxy of canal contact and 96.7% of them achieved the highest titre. The alteration in the actual prevalence of schistosomiasis mansoni in Egypt with emergence of new foci including Cairo, Luxor, Aswan, Mersa-Matrouh and the north-eastern province alongside Suez Canal that may be explained by the associated socioepidemiologic risk factors.

Genomic instability in complicated and uncomplicated Egyptian schistosomiasis haematobium patients.

BACKGROUND: Exploration of genetic changes during active Schistosoma infection is important for anticipation and prevention of chronic sequelae. This study aimed to explore the genomic instability in chromosomal and cellular kinetics in Egyptians suffering from uncomplicated active schistosomiasis haematobium infection in addition to chronic schistosomiasis haematobium cases complicated by bilharzial-associated bladder cancer (BAC). RESULTS: This study was conducted on 46 schistosomiasis haemotobium cases, 22 were active (Viable S. haematobium eggs in urine samples as detected by microscopy) and 24 were chronic complicated with bladder cancer. Three cytogenetic techniques were applied; the first was quantitative nuclear-morphocytometry by means of which the Feulgen-stained nuclei were analyzed for parameters including shape, size, integrated optical-density and nuclear area. The second was Fluorescent In-Situ Hybridization (FISH) for specific p53gene-locus of chromosome 17 and the third technique was karyotyping. Concerning chronic complicated cases, the mean ± SD of DNA-content in urinary bladder tissue sections was 3.18 ± 0.65. Five samples (20.83%) of bladder tissue sections of chronic complicated cases showed diploid nuclei, 6 urinary bladder tissue samples (25%) were tetraploid, while 13 bladder samples (54.16%) were aneuploid. Epithelial cells of urine samples demonstrated aneuploidy (mean ± SD = 3.74 ± 0.36).Nuclear contents showed high proliferative DNA index in all urinary epithelial cells. In the acute uncomplicated group, nuclear-DNA of urinary epithelial cells was found diploid with mean nuclear-DNA content of 2.2 ± 0.16SD. Half of these diploid smears had a high proliferation index. The difference between nuclear DNA-contents in acute and chronic cases was significant (P = 0.0001). FISH technique for specific p53gene-locus and karyotyping were done on urinary bladder tissue specimens and peripheral blood monocytes of 8 chronic cases respectively. Three samples (37.5%) with invasive BAC had a deletion of the p53 gene. Karyotyping showed three cases out of the 8 chronic schistosomiasis haematobium patients with chromosomal fragmentations. CONCLUSIONS: DNA morphometry was valuable in detection of gross genetic changes in urothelial tissues. It is an important prognostic factor in established schistosomiasis haematobium induced bladder malignancy. It has the great advantage of being applicable on urine cells making it suitable for the prediction of a tendency towards genetic instability in active schistosomiasis haematobium patients.

The pathogenic role of different Blastocystis hominis genotypes isolated from patients with irritable bowel syndrome.

BACKGROUND AND STUDY AIMS: The pathogenic role of Blastocystis hominis is still regarded by some as controversial. Studies have been in progress for years to evaluate the role of blastocystosis in irritable bowel syndrome (IBS) and demonstrated that faecal carriage of B. hominis was frequent in these patients. This study attempted to distinguish different genotypes of B. hominis isolates obtained from patients with IBS and to evaluate their pathogenic potentials. PATIENTS AND METHODS: One hundred subjects (51 patients with IBS and 49 asymptomatic infected subjects) harbouring B. hominis were investigated by a direct smear examination and in vitro culture of stool samples followed by genotyping of B. hominis by PCR using STS primers. Sigmoidoscopy was done in all subjects and biopsies were taken and subjected to histopathologic examination. RESULTS: Genotyping proved that only four genotypes of B. hominis were identified. In patients with IBS, genotypes III, I, and IV were detected (28, 15 and 14 isolates, respectively). On the other hand, genotypes III, IV, and II were identified in asymptomatic infected individuals (21, 19 and 13 isolates, respectively). The degrees of chronic inflammatory changes in sigmoidoscopic biopsies caused by B. hominis genotypes among IBS patients revealed that severe inflammation was present mainly in patients harboring genotype I isolates (4/15) (26.66%), while genotype III caused severe inflammation only in 9.09%. Genotype II isolates were not detected in IBS cases. Asymptomatic infected individuals harboring genotypes II, III and IV exhibited mild to moderate inflammatory changes. Genotype I isolates were not detected in asymptomatic infected group. The correlation between different B. hominis genotypes and degree of inflammation was statistically insignificant. CONCLUSION: Genotype I was the most pathogenic genotype of B. hominis isolates in patients with IBS while genotype II was not detected among those patients. Also, our results suggest the presence of pathogenic and non-pathogenic strains among genotypes III and IV.