Proposed morbidity markers among Schistosoma mansoni patients.

Background: Fecal calprotectin (FC) and fecal occult blood (FOB) were suggested as potential inflammatory markers for assessing intestinal schistosomiasis morbidity that are conventionally detected through invasive methods. Aim and Objectives: The present work aimed to evaluate FC and FOB as morbidity markers of Schistosoma mansoni infection before and after praziquantel treatment. Materials and Methods: A total of 205 stool samples (117 schoolchildren and 88 adults) were collected and examined by Kato Katz. A questionnaire enquiring about diarrhea, history of blood in stool, and abdominal pain was designed and applied. Results: S. mansoni prevalence rates were 20.5% and 11.36% among children and adults, respectively; the majority of cases had light infection intensity. FC and FOB were studied among 25 cured S. mansoni cases (17 children and 8 adults) pre and one-month post treatment. Before treatment, six and four children of moderate and high S. mansoni infection intensity tested positive for FC and FOB, respectively, all turning negative after treatment. FC showed borderline statistical significance before and after treatment among children. However, all adults tested negative for FC and FOB. Conclusion: FC and FOB could be possibly used as morbidity monitoring tools for S. mansoni infection in children with moderate and high infection intensity.

Key cytokines and hematological parameters in patients with uncomplicated falciparum malaria in Hodeidah, Yemen.

Immunity to malaria has a major role in controlling disease and pathogenesis with cytokine production being involved in almost every phase of the immune response. The present study aimed to assess hematological variables and to measure plasma levels of TNFα, IFNγ and IL10, their ratios, and their relation to parasitemia among patients with uncomplicated falciparum malaria in Hodeidah, Yemen. Forty patients with uncomplicated P. falciparum monoinfection and 40 healthy age and sex-matched controls were enrolled in the study. Parasitological diagnosis was confirmed, and parasite density was estimated. Plasma cytokine levels, hematologic parameters, and the presence of gametocytes were determined. Results revealed higher TNFα, IFNγ and IL10 in patients than in controls. A relatively higher IL10 production was demonstrated by the significantly lower TNFα/IL10 and IFNγ/IL10 ratios in patients than in controls. TNFα and IL10 correlated positively with parasite density. Lower Hb levels, RBC, lymphocyte and platelet counts, and higher neutrophil and reticulocyte counts were observed in patients compared to controls. Reticulocyte count was higher and IFNγ level was lower in the presence of gametocytes. Conclusively, uncomplicated falciparum malaria is associated with the ability to regulate the production of pro-inflammatory and anti-inflammatory cytokines. This mediates parasite clearance while simultaneously avoiding severe pathology.

Remarkable histopathological improvement of experimental toxoplasmosis after receiving spiramycin-chitosan nanoparticles formulation.

The present study investigated the anti-Toxoplasma effect of chitosan nanoparticles [CS NPs], spiramycin, spiramycin co-administered with metronidazole and spiramycin-CS NPs formulation on the parasite burden and histopathological changes in the liver, spleen and brain in experimentally infected mice. Seventy male Swiss albino mice were classified into seven equal groups: healthy control (I), infected untreated control (II), infected group receiving CS NPs (III), spiramycin administered infected group (IV), infected group receiving spiramycin-metronidazole (V), infected receiving 400 mg/kg spiramycin-CS NPs (VI) and infected treated with spiramycin-loaded CS NPs 100 mg/kg (VII). All groups were inoculated intraperitoneally with 2500 T. gondii tachyzoites RH strain except the healthy control group. All groups were sacrificed on the 8th day after infection. Density of the parasite and histopathological examination of the liver, spleen and brain of all treated mice revealed reduction in the mean tachyzoites count as well as decreased inflammation, congestion and necrosis within tissue sections. Spiramycin-loaded NPs displayed the highest significant reduction in the pathological insult tailed by spiramycin-metronidazole and CS NPs. In conclusion, spiramycin-loaded CS NPs showed a promising synergistic combination in the treatment of the histopathology caused by toxoplasmosis.

Evaluation of microscopy and PCR for detection of Dientamoeba fragilis.

Introduction: Dientamoeba fragilis (D. fragilis) diagnosis is an intestinal protozoan parasite globally found in rural and urban areas and is attracting a growing interest. Its prevalence in stool varies from 0.2% to more than 19% depending upon the population studied. Materials and Methods: This study was based on the examination of 100 stool samples of randomly referred cases in a rural area in Motobus district, Kafr El-Sheikh governorate, Egypt. Our aim was to investigate the presence of D. fragilis in stool of the examined individuals using conventional polymerase chain reaction (PCR) compared to wet mount and trichrome stain with confirmation of infection by transmission electron microscopy. Results: D. fragilis was detected in 13/100 of the stool samples examined using wet mount smears, while trichrome stain detected 17/100. Conventional PCR diagnosed 41 cases of D. fragilis in the studied group. A very good agreement was found between wet mount and trichrome stain for diagnosing D. fragilis, while there was fair agreement between conventional PCR and both microscopy methods. Transmission electron microscope was performed on pooled positive samples that revealed the internal structures of D. fragilis trophozoite with its characteristic nucleus. Conclusions: PCR technique was superior to microscopy for the detection of D. fragilis. Trichrome stain remains vital for microscopic diagnosis.

A study on positive school children as indicators of schistosomiasis status in their families.

This study aimed to evaluate schistosomiasis positive school children as indicators of infection in their families. Four hundred and sixteen stool samples of children from two primary schools were examined microscopically for Schistosoma mansoni (S. mansoni) after Kato-Katz. Equal numbers from families of schistosomiasis positive and negative school children were examined by Kato-Katz. Circulating cathodic antigen (CCA) cassette test was performed on 100 Kato-Katz negative children to detect missed S. mansoni cases if any. S. mansoni infection rates among the primary school children were 15.3% and 1.6% in high and low prevalence schools respectively. Sixty five percent of school children had light intensity infection. Family members were either negative or exhibited a prevalence rate as low as 5-6% whether related to positive or negative children. A higher infection rate of S. mansoni was detected by the CCA test compared to the Kato-Katz. Conclusively, children can be good indicators of their particular families in areas of limited resources and low endemicity. Efforts are required to permit large scale use of CCA cassette test.

Performance of formalin ethyl acetate, percoll sedimentation and ELISA for the detection of Cryptosporidium infection in asymptomatic children.

This study aimed to assess the performance of formalin ethyl acetate (FEA)/modified Ziehl-Neelsen (MZN), and percoll technique/MZN for the diagnosis of cryptosporidiosis among asymptomatic children compared to ELISA coproantigen. The study was conducted on 100 children in a rural area in Kafr El-Sheikh governorate. Stool samples were collected and examined by the three techniques. Microscopic examination revealed the presence of acid-fast stained oocysts and non-acid fast ghost oocysts. The overall prevalence rate was 7% with an infection intensity of 1-5 oocysts/oil immersion field. FEA/MZN technique showed the highest diagnostic performance (5%) with 71.4% sensitivity and 98% negative predictive value (NPV) compared to the other techniques. ELISA revealed 3% prevalence, 42.9% sensitivity and 96% NPV. Percoll/MZN gave the lowest prevalence, sensitivity and NPV (1%, 14.29% and 93.9% respectively). Agreement fluctuated between moderate and poor regarding FEA/MZN versus ELISA and percoll/MZN versus both techniques. In conclusion, FEA/MZN gave the top diagnostic performance, yet it missed some positive cases. Its combination with ELISA coproantigen might prove beneficial for Cryptosporidium diagnosis. Percoll technique needs more validation by modifying the density gradient, speed of centrifugation, and staining methods.

Intestinal parasites among humans and their livestock animals in a rural community in Gharbia governorate, Egypt.

Our objective was to study the prevalence of intestinal parasitic infections among humans and their contact livestock animals in a rural community in Gharbia governorate, Egypt. The collected human and livestock animal fecal samples were examined by formol ether sedimentation technique and modified Ziehl-Neelsen stain. Human samples were further examined by the Kato-Katz technique. The total prevalence of intestinal parasites in humans was 52.3%; 7.3% helminths and 45% protozoa. Regarding animals, 29% of livestock animals had protozoan parasites and no helminths were detected. Results revealed that risk factors including drinking tap water and attendance of nurseries, affected the rate of parasitic infection among humans. The infection rate of protozoa was significantly higher among humans (1.5 times) as compared to animals.

Intestinal schistosomiasis among preschool and school-aged children in a rural setting near Alexandria: initiative for elimination.

OBJECTIVE: To assess the status of intestinal schistosomiasis among preschool-aged (PSAC) and school-aged children (SAC) and to compare the efficacy of praziquantel (PZQ) in both groups. METHODS: The study was conducted on 400 children; 103 PSAC and 297 SAC. Diagnosis of Schistosoma mansoni was based on triplicate Kato-Katz thick smears from a single stool sample. To identify the missed cases by Kato-Katz, 120 randomly selected negative cases (38 PSAC and 82 SAC) were screened by real-time PCR. All S. mansoni-positive cases by Kato-Katz were treated by crushed PZQ tablets. Four weeks after treatment, the cure rate was assessed by Kato-Katz smears and real-time PCR. RESULTS: The prevalence of S. mansoni with Kato-Katz was 7.8% among PSAC and 7.4% among SAC. Most of children (63.3%) had light-intensity infection. The cure rate was 100% among PSAC by both techniques, and 91%, and 77.2% among SAC by Kato-Katz and real-time PCR, respectively. In the 120 stool samples screened by real-time PCR, S. mansoni prevalence was 25%; 15.8% and 29.3% were among PSAC and SAC respectively. Treated cases showed a lower range of Ct values than untreated cases. Two melting temperature ranges (Tm = 83-87°C and 89-93°C) were recognised among uncured cases which may point to S. mansoni genetic variability. CONCLUSION: Continuous monitoring and inclusion of PSAC in schistosomiasis control programmes are crucial. Real-time PCR and other molecular tools are recommended for evaluation of the true prevalence, assessment of cure and further studies on genetic diversity.

Therapeutic efficacy of mebendazole and artemisinin in different phases of trichinellosis: a comparative experimental study.

The present work aimed at studying the efficacy of mebendazole (MBZ) compared to artemisinin (ART) for the treatment of trichinellosis at various phases of infection. Seventy Swiss albino mice were orally infected by 300 Trichinella spiralis (T. spiralis) larvae. Mice were divided into infected untreated control group and infected groups treated with 50 mg kg-1 MBZ and 300 mg kg-1 ART for three and five consecutive days, respectively, at the enteral phase [2-4 days post infection (PI)], invasive phase (10-12 days PI) and encapsulated phase (28-30 days PI). All mice were sacrificed 35-42 days PI. MBZ and ART revealed a significant decrease in mean larval counts and increase of larval per cent reduction (LR %) when treatment was initiated during the enteral phase compared to the other phases. MBZ showed significantly higher LR % (99.7, 83.95 and 89.65%) than ART (80.58, 67.0 and 79.2%) when administered at the three infection phases. Histopathological study showed a decrease in the number of encysted larvae, their surrounding cellular infiltrates and increased regenerative muscles in all treated mice. In conclusion, ART possesses a substantial anthelmintic activity against T. spiralis infection in mice both at the enteral and encapsulated phases, yet, significantly lower than MBZ.

Detection and Molecular Identification of Cryptosporidium Species Among Children with Malignancies.

BACKGROUND: Cryptosporidiosis represents a major health problem worldwide particularly among children. Its diagnosis is still difficult and demands sensitive methods. In Egypt, there is little documentation of infection among children with malignancies. This work was designed to study the infection rate of Cryptosporidium among children with malignancies, compare the performance of modified Ziehl-Neelsen (MZN) stain with nested polymerase chain reaction (PCR) and identify the species subtypes of positive cases. METHODS: The study was conducted on 100 children with malignancies (leukemia, lymphoma and solid tumors), below 10 years of age, from El-Shatby hospital, Alexandria University. After obtaining the informed consent, their stool samples were collected and examined microscopically following MZN stain for the diagnosis of Cryptosporidium spp. All samples were then subjected to nested PCR. Restriction fragment length polymorphism (RFLP) targeting the Cryptosporidium oocyst wall protein (COWP) gene was applied to positive cases, using restriction enzyme RsaI for digestion of nested PCR products. RESULTS: Out of the 100 examined children, MZN detected higher positive cases compared to nested PCR. Six cases (6%) were diagnosed positive by MZN stain, three of which (3%) were concordantly positive by nested PCR. All positives were among children with acute lymphoblastic leukemia (ALL). Fair agreement was found between the two tests (K = 0.36). Genotyping results revealed that positive samples were of Cryptosporidium parvum (C. parvum) type II. CONCLUSIONS: Low Cryptosporidium infection rate was detected among children with malignancies. MZN diagnosed more positive cases compared to nested PCR. C. parvum type II was the identified species among the examined children. Further optimization of PCR steps is needed.

Comparison among FLOTAC, Kato-Katz and formalin ether concentration techniques for diagnosis of intestinal parasitic infections in school children in an Egyptian rural setting.

The study aimed to compare the diagnostic performance of the Kato-Katz, formalin ether concentration method (FECM) and FLOTAC using Sheather's sugar solution (FS1), saturated sodium chloride (FS2) and zinc sulfate (FS7) for the diagnosis of intestinal parasites among school children, focusing on Schistosoma mansoni. Ninety fecal samples were examined using the above mentioned techniques. The overall infection rate was 87.7%. Concerning protozoa, FLOTAC (FS1 and FS2) and FECM detected nearly equal infection rates (43.3% and 44.4%, respectively) with very good agreement. Kato-Katz diagnosed the highest helminthic infection rate (57.8%) followed by FLOTAC FS7 (44.4%) and FECM showed the lowest helminthic infection rate (27.7%). As for S. mansoni, Kato-Katz showed an infection rate of 38.8% vs FLOTAC (22.2%) and FECM (11.1%). The three techniques detected the same infection rate (11.1%) with egg counts more than 72 eggs/gram of feces. The FLOTAC sensitivity and accuracy for the diagnosis of protozoa were 97% and 99%, respectively. Regarding helminths diagnosis, FLOTAC technique showed higher sensitivity (77%) and accuracy (87%) compared to FECM (48% sensitivity and 70% accuracy). Therefore, FLOTAC can be used synchronously or in replacement to other diagnostic techniques. This can strategically impact future control programmes of intestinal parasitic infections in limited resources settings.

Intestinal Microsporidia Infection in Leukemic Children: Microscopic and Molecular Detection.

BACKGROUND: Microsporidia infection was originally described as an immunocompromised associated pathogen. Limitations to correct microscopic diagnosis of microsporidia include size of the organism presenting a challenge even to a highly competent laboratory expert. OBJECTIVE: The present study aimed to detect microsporidia infection among leukemic children. The performance of modified trichrome stain and PCR in the diagnosis of microsporidia was evaluated with further speciation. METHODS: Stool samples of 100 leukemic children on chemotherapy were examined microscopically for microsporidia. DNA was extracted from all samples. Amplification was performed by conventional and nested PCR. Sequencing of amplified products was performed on unspeciated samples. RESULTS: Microsporidia were detected in 23% of the children by MTS and 29% by PCR. The 29 positive samples were subjected to PCR for speciation. Enterocytozoon bieneusi was found to predominate in 20 cases, Encephalitozoon intestinalis in three cases, two cases had co-infection, and the remaining four samples were not amplified with either E. bieneusi or E. intestinalis specific primers. By DNA sequencing of the unspeciated samples, three samples shared high homology with Encephalitozoon hellem and one sample with Encephalitozoon cuniculi. Referring to PCR as a gold standard, MTS exhibited 72.4% sensitivity and 97.2% specificity with 90% accuracy. Among a number of studied variables, diarrhea and colic were significantly associated with microsporidia infection when diagnosed by either technique. CONCLUSION: The use of sensitive and discriminative molecular tools will contribute to determining the true prevalence of microsporidiosis and possibly their potential transmission source depending on species identification.

Successful treatment of acute experimental toxoplasmosis by spiramycin-loaded chitosan nanoparticles.

Spiramycin-metronidazole and spiramycin-loaded chitosan (CS) nanoparticles (NPs) were tested in comparison with the current spiramycin treatment of T.gondii concerning tissue penetration and blood brain barrier (BBB) passage. Swiss Albino mice were inoculated intraperitoneally with 2500 T. gondii tachyzoites RH strain and were divided into experimental and control groups. The experimental groups orally received CS NPs, spiramycin, spiramycin-metronidazole, spiramycin-loaded CS NPs 400 mg/kg and spiramycin-loaded CS NPs 100 mg/kg. Drug efficacy was assessed by mice survival time, mortality rate, parasite load in different organs and morphological study of the tachyzoites movement by light microscope and the ultra-structure by SEM. The results revealed that the maximum survival time of more than 200 days with no mortality on the sacrifice day (8th) was observed in mice receiving spiramycin-loaded NPs. Spiramycin-loaded NPs showed the highest significant percent reduction of tachyzoites (about 90% reduction) in liver, spleen and brain as compared to the other used drugs denoting successful bypass of BBB. Light microscopy of the treated peritoneal tachyzoites showed sluggish tachyzoites movement while the NPs caused loss of their movement. SEM of the treated tachyzoites were more mutilated and some of them appeared rupturing in those receiving CS NPs and spiramycin-loaded NPs. In conclusion, spiramycin-loaded NPs showed the highest efficiency in the treatment of acute toxoplasmosis. The non-toxic nature and the anti-parasitic effect of both CS and spiramycin make the use of spiramycin-loaded CS NPs a potential material for treatment of human toxoplasmosis.

Assessing the marginal error in diagnosis and cure of Schistosoma mansoni in areas of low endemicity using Percoll and PCR techniques.

OBJECTIVE: The Kato-Katz technique misses a proportion of low-intensity schistosomiasis cases. We set out to estimate the rate of false negative results after Kato-Katz. METHOD: Comparison with the more sensitive Percoll technique and polymerase chain reaction (PCR). RESULTS: Amongst Kato-Katz negative Schistosoma mansoni specimens in a hypoendemic area, Percoll detected 11% positive cases and PCR detected 23%. After treatment, the cure rate was 97% by Kato-Katz, 90% by Percoll and 71% by PCR. Results of Kato-Katz and Percoll showed moderate agreement, all cases positive by Kato-Katz were positive by Percoll. However, PCR showed some negative results amongst cases considered sure positives. CONCLUSION: The Percoll technique seems to be the technique of choice for diagnosis of S. mansoni in low intensity areas and after treatment.