Detection of chronic toxoplasmosis in the brain of mice using loop-mediated isothermal amplification (LAMP) and conventional PCR.

BACKGROUND: Toxoplasma gondii is a common protozoan parasite that infects approximately one-third of the world's population. It is a disease with multiple manifestations. In immunocompetent individuals, symptoms are mild and flu-like, whereas, in immunocompromised patients, it often results in severe morbidity and mortality. Thus, studies for developing a simple, rapid diagnostic tool for early detection of Toxoplasma are emerging. Molecular diagnosis is highly accurate and helpful in congenitally infected and immunocompromised patients. The loop-mediated isothermal amplification (LAMP) technique was invented to improve nucleic acid amplification efficacy in terms of sensitivity and specificity. AIM OF THE STUDY: The study aimed to validate a LAMP protocol for detecting Toxoplasma DNA in the brain homogenates from mice experimentally infected with Toxoplasma's ME-49 (cyst-forming type II) strain in comparison to PCR. METHODS: In this study, the target DNA fragment was the Toxoplasma 529-bp, repeated 200-300 copies/genome. The sensitivity of both LAMP and conventional PCR techniques was estimated in brain homogenates in experimental mice at eight weeks post-infection and compared to the histopathology data. RESULTS: The LAMP reaction showed positive results in 18 of the 26 examined samples of brain homogenates. PCR showed the characteristic 529-bp band in 15 of the 26 examined samples. CONCLUSION: The LAMP showed a higher sensitivity over PCR in detecting Toxoplasma infection in brain homogenates of infected mice.

The Idiosyncratic Efficacy of Spironolactone-Loaded PLGA Nanoparticles Against Murine Intestinal Schistosomiasis.

Background: Schistosomiasis is a chronic debilitating parasitic disease accompanied with severe mortality rates. Although praziquantel (PZQ) acts as the sole drug for the management of this disease, it has many limitations that restrict the use of this treatment approach. Repurposing of spironolactone (SPL) and nanomedicine represents a promising approach to improve anti-schistosomal therapy. We have developed SPL-loaded poly(lactic-co-glycolic acid) (PLGA) nanoparticles (NPs) to enhance the solubility, efficacy, and drug delivery and hence decrease the frequency of administration, which is of great clinical value. Methods: The physico-chemical assessment was performed starting with particle size analysis and confirmed using TEM, FT-IR, DSC, and XRD. The antischistosomal effect of the SPL-loaded PLGA NPs against Schistosoma mansoni (S. mansoni)-induced infection in mice was also estimated. Results: Our results manifested that the optimized prepared NPs had particle size of 238.00 ± 7.21 nm, and the zeta potential was -19.66 ± 0.98 nm, effective encapsulation 90.43±8.81%. Other physico-chemical features emphasized that nanoparticles were completely encapsulated inside the polymer matrix. The in vitro dissolution studies revealed that SPL-loaded PLGA NPs showed sustained biphasic release pattern and followed Korsmeyer-Peppas kinetics corresponding to Fickian diffusion (n<0.45). The used regimen was efficient against S. mansoni infection and induced significant reduction in spleen, liver indices, and total worm count (ρ<0.05). Besides, when targeting the adult stages, it induced decline in the hepatic egg load and the small intestinal egg load by 57.75% and 54.17%, respectively, when compared to the control group. SPL-loaded PLGA NPs caused extensive damage to adult worms on tegument and suckers, leading to the death of the parasites in less time, plus marked improvement in liver pathology. Conclusion: Collectively, these findings provided proof-of-evidence that the developed SPL-loaded PLGA NPs could be potentially used as a promising candidate for new antischistosomal drug development.

Intestinal Giardiasis in Children Undergoing Upper Endoscopy for Unexplained Gastrointestinal Symptoms: Implication for Diagnosis.

BACKGROUND: Giardia lamblia is a flagellated protozoan causing diarrheal outbreaks worldwide. Microscopic stool examination is widely used. We conducted this study to explore intestinal giardiasis in children undergoing upper endoscopy for unexplained gastrointestinal symptoms. METHODS: The study included 160 children undergoing upper endoscopy for unexplained gastrointestinal symptoms (patients) and 90 children as controls. We collected stool samples for microscopic examination and ELISA coproantigen detection from all participants. We examined duodenal biopsies for patients. RESULTS: In patients, stool examination revealed Giardia in 23.8% and coproantigen detection was positive in 37.5%. Endoscopic duodenal biopsies revealed Giardia trophozoites in 5% of patients, in addition to various pathological changes. CONCLUSION: Giardiasis was significantly higher (P = 0.001) in children with unexplained gastrointestinal complaints than the controls. Diagnosis by coproantigen detection was superior to microscopic stool examination, with a sensitivity of 90.9%. Duodenal biopsies examination confirmed the infection in fewer cases but added other diagnostic information.

Association between Toxoplasma gondii infection and coronary atherosclerosis.

BACKGROUND: Toxoplasma gondii is a worldwide protozoon that can infect all nucleated vertebrate cells. Little information is available about the association between T. gondii infection and coronary atherosclerosis. METHODS: A total of 320 cases were enrolled (160 patients with coronary atherosclerosis and 160 non-atherosclerotic individuals). Blood samples were collected to measure anti-T. gondii immunoglobulin G (IgG) antibodies using enzyme-linked immunosorbent assay (ELISA) and serum lipid profile. Coronary angiogram was also performed. RESULTS: The seroprevalence of anti-Toxoplasma antibodies in atherosclerotic and non-atherosclerotic individuals was 63.1% and 46.2%, respectively, with higher levels of anti-T. gondii IgG in atherosclerotic patients. Consumption of contaminated water, unwashed fruits and vegetables and raw meat and contact with soil were significant risk factors for Toxoplasma infection. Significant differences were detected in serum levels of low-density lipoproteins, triglycerides and cholesterol between both groups. Positive correlations were detected between ELISA titres and serum levels of low-density lipoproteins, triglycerides and cholesterol, disease severity and the number of affected vessels. Male gender and contact with soil had a significant association with positive T. gondii serology in atherosclerotic patients. CONCLUSIONS: Patients with coronary atherosclerosis have a high prevalence of T. gondii infection. More studies are crucial to elucidate the mechanisms underlying the effects of chronic toxoplasmosis on coronary atherosclerosis.

Trichomonas vaginalis serostatus and prostate cancer risk in Egypt: a case-control study.

Trichomonas vaginalis is one of the most common non-viral sexually transmitted infections (STIs) that has been associated with prostate cancer in some countries. This study aims to investigate if T. vaginalis infection can be a risk factor for prostate cancer in Egypt and its possible relationship with cancer prognostic factors and overall survival. Serum samples were collected from a total of 445 age-matched males; 126 with prostate cancer, 108 with bladder cancer, 91 with different types of cancers, and 120 healthy controls, and then analyzed by ELISA for detection of anti-Trichomonas IgG and prostate-specific antigen (PSA). The results revealed that only 8.3% of controls were seropositive for trichomoniasis, compared with 19% of prostate cancer patients (P = 0.015). There were positive associations between the levels of PSA and tumor stage with T. vaginalis IgG optical density scores among the seropositive cases (P < 0.001 and < 0.05, respectively). However, no significant correlations were detected between seropositivity of T. vaginalis and other prognostic factors or overall survival in those patients. In conclusion, chronic T. vaginalis infection may be associated with prostate cancer, but it does not seem that this STI aggravates the cancer status.

In vivo studies of the effect of PPQ-6, a quinoline-based agent against Schistosoma mansoni in mice.

Schistosomiasis is still a public health problem. Praziquantel is the only drug available for treatment of all forms of human schistosomiasis. Although praziquantel is an effective drug against all species of human schistosomes, concerns about resistance have been raised, especially in endemic areas. A hybrid compound containing several pharmacophore within a single molecule is a promising strategy. Here, we described the anti-schistosomal effect of 4-(2-Chloroquinolin-3-yl)-2-oxo-6-(p-tolyl)-1,2-dihydropyridine-3-carbonitrile (PPQ-6), a hybrid drug based on quinoline and pyridine. PPQ-6 was given as two regimens (20 or 40 mg/kg). In both regimens, PPQ-6 significantly reduced liver and spleen indices, nitric oxide production, tissue egg load, hepatic granuloma size and count, immature eggs and total worm burden especially females. Our findings suggested that PPQ-6 is a promising anti-schistosomal agent; however more research is needed to elucidate its mechanism of action and report its activity on juvenile schistosomes and other species of human schistosomes.

Loop mediated isothermal amplification (LAMP) of Toxoplasma DNA from dried blood spots.

INTRODUCTION: The cosmopolitan protozoan Toxoplasma gondii is a major parasite of warm-blooded animals including man. Early and accurate diagnosis is a must for proper treatment that prevents life threatening sequels. Loop-mediated isothermal amplification (LAMP) is a novel technique that can amplify DNA with high sensitivity and specificity under isothermal conditions. AIM OF THE STUDY: To validate a LAMP-specific protocol for detection of Toxoplasma DNA using dried blood spots (DBS) from mice experimentally infected with the cystogenic Toxoplasma ME-49 strain. METHODS: In this study, the target DNA fragment was the Toxoplasma 529-bp repeat element that exists in 200-300 copies per T. gondii genome. The sensitivity of both LAMP and conventional PCR techniques was estimated in DBS samples from experimental mice at 1-week and 8-weeks post-infection. RESULTS: Out of 20 blood samples gathered on Whatman filter paper from mice at 1-week post-infection, 18 and 16 were positive by LAMP and conventional PCR, respectively. Neither techniques detected parasite DNA in blood at 8th week of infection. CONCLUSION: Dried blood spots are easy source of material for molecular studies. LAMP assay proved higher sensitivity than the conventional PCR in detecting parasitemia in early infection with the cystogenic Toxoplasma strain.

Effect of omega-3 fatty acids administered as monotherapy or combined with artemether on experimental Schistosoma mansoni infection.

Schistosomiasis is on the top list of endemic diseases in sub-Saharan Africa. Praziquantel is the drug of choice for treatment of human schistosomiasis. Yet, the sole dependence on the drug raises concerns about the potential for increased drug resistance, which would subsequently result in searching for alternative preventive chemotherapy options, ideally among natural compounds. Therefore, we conducted this work to assess the effect of omega-3 polyunsaturated fatty acids [(ω-3) PUFAs] monotherapy or combined therapy with artemether (ART) against Schistosoma mansoni infection in a mouse model. A total of 42 mice were divided into 4 groups and infected with 50 ± 5 S. mansoni cercariae for 10 weeks. Mice were treated orally with either (ω-3) PUFAs as 273 mg/ kg, 4 times/ week throughout the experiment, ART as a single dose of 400 mg/ kg, 3 weeks post-infection, or combined ART + (ω-3) PUFAs using the same respective treatment regimen, while infected untreated mice were served as controls. The study explored that combined administration of (ω-3) PUFAs and ART has the best schistosomicidal efficacy as it significantly reduced liver and spleen indices, worm count, egg burdens, and granulomas count as well as diameter. Besides, the combined regimen was associated with a significant decrease in both hepatic nitric oxide and serum interleukin-4 level. The results highlighted the possibility of using (ω-3) PUFA combined with ART as a novel anti-schistosomal combination therapy. However, further researches should be conducted to clarify the possible synergistic mechanism/s between the two natural compounds.