Quercetin inhibits Toxoplasma gondii tachyzoite proliferation and acts synergically with azithromycin.

Quercetin (QUE) is a natural polyphenol known to have numerous pharmacological properties against infectious and non-infectious diseases. Azithromycin (AZ) is an antibiotic that belongs to the azalide class of antimicrobials and an antiparasitic that is known to be effective in combination with clindamycin against pyrimethamine/sulfadiazine-resistant Toxoplasma gondii tachyzoites in clinical settings. Both compounds are known to target protein synthesis and have anti-inflammatory properties. However, little is known about QUE and AZ synergistic interaction against T. gondii growth. Here, we report for the first time the effects of the combination of QUE and AZ on T. gondii growth. The 50% inhibitory concentration (IC50) for QUE at 72 h of interaction was determined to be 0.50 µM, whereas AZ gave an IC50 value of 0.66 µM at 72 h of interaction with parasites. Combination testing of QUE and AZ in a ratio of 2:1 (QUE:AZ) showed an IC50 value of 0.081 µM. Interestingly, a fractional inhibitory index value of 0.28 was observed, indicating a strong synergy. QUE was also found to upregulate the generation of reactive oxygen species and cause dysfunction of the mitochondria membrane of both intracellular and extracellular T. gondii tachyzoites. Overall, the results indicate that QUE is a novel lead capable of synergizing with AZ for inhibiting T. gondii growth and may merit future investigation in vivo for possible combination drug development.

Anti-Toxoplasma gondii activity of Trametes versicolor (Turkey tail) mushroom extract.

Toxoplasma gondii (T. gondii) infection continues to rise globally in humans and animals with high socioeconomic and public health challenges. Current medications used against T. gondii infection are limited in efficacy, safety, and affordability. This research was conducted to assess the higher fungi extract effect on T. gondii tachyzoites growth in vitro and possibly decipher its mechanism of action. Furthermore, we evaluated the extract's effect on human foreskin fibroblast viability. The methanol extracts of Turkey tail (TT) mushroom was tested against T. gondii tachyzoites growth using an RH-RFP type I strain that expresses red fluorescent protein throughout culture in a dose-dependent manner using a fluorescent plate reader. Similarly, we tested the effect of the extract on host cell viability. We observed that TT extract inhibited tachyzoites growth with a 50% minimum inhibitory concentration (IC50s), IC50 = 5.98 ± 1.22 µg/mL, and 50% cytotoxic concentration (CC50s), CC50 ≥ 100 µg/mL. It was discovered that TT extract induced strong mitochondria superoxide and  reactive oxygen species production and disrupted mitochondria membrane potential in T. gondii tachyzoites. Additionally, scanning electron microscopy depicted that TT extract and pyrimethamine (PY) caused a morphological deformation of tachyzoites in vitro. In conclusion, TT methanol extract made up of phytosterols, bioactive sphingolipids, peptides, phenolic acids, and lactones could be a promising source of new compounds for the future development of anti-Toxoplasma gondii drugs. Extracts were non-cytotoxic, even at higher concentrations.

Targeting the Plasmodium falciparum proteome and organelles for potential antimalarial drug candidates.

There is an unmet need to unearth alternative treatment options for malaria, wherein this quest is more pressing in recent times due to high morbidity and mortality data arising mostly from the endemic countries coupled with partial diversion of attention from the disease in view of the SARS-Cov-2 pandemic. Available therapeutic options for malaria have been severely threatened with the emergence of resistance to almost all the antimalarial drugs by the Plasmodium falciparum parasite in humans, which is a worrying situation. Artemisinin combination therapies (ACT) that have so far been the mainstay of malaria have encountered resistance by malaria parasite in South East Asia, which is regarded as a notorious ground zero for the emergence of resistance to antimalarial drugs. This review analyzes a few key druggable targets for the parasite and the potential of specific inhibitors to mitigate the emerging antimalarial drug resistance problem by providing a concise assessment of the essential proteins of the malaria parasite that could serve as targets. Moreover, this work provides a summary of the advances made in malaria parasite biology and the potential to leverage these findings for antimalarial drug production.

Inhibition of Toxoplasma gondii Growth by Dihydroquinine and Its Mechanisms of Action.

Toxoplasma gondii is a zoonotic parasite that infects the brain of humans and causes cerebral toxoplasmosis. The recommended drugs for the treatment or prophylaxis of toxoplasmosis are pyrimethamine (PY) and sulfadiazine (SZ), which have serious side effects. Other drugs available for toxoplasmosis are poorly tolerated. Dihydroquinine (DHQ) is a compound closely related to quinine-based drugs that have been shown to inhibit Plasmodium falciparum and Plasmodium berghei in addition to its anti-arrhythmia properties. However, little is known about the effect of DHQ in T. gondii growth and its mechanism of action in vitro. In this study, we report the anti-Toxoplasma and anti-invasion properties of DHQ. DHQ significantly inhibited T. gondii tachyzoite growth with IC50s values of 0.63, 0.67, and 0.00137 µM at 24, 48, and 72 h, respectively. Under similar conditions, SZ and PY, considered as the gold standard drugs for the treatment of toxoplasmosis, had IC50s values of 1.29, 1.55, and 0.95 and 3.19, 3.52, and 2.42 µM, respectively. The rapid dose-dependent inhibition of T. gondii tachyzoites by DHQ compared to the standard drugs (SZ and PY) indicates that DHQ has high selective parasiticidal effects against tachyzoite proliferation. Remarkably, DHQ had an excellent selectivity index (SI) of 149- and 357-fold compared to 24- and 143-fold for PY and SZ, respectively, using fibroblast cells. In addition, DHQ disrupted T. gondii tachyzoite mitochondrial membrane potential and adenosine triphosphate (ATP) production and elicited high reactive oxygen species (ROS) generation. Taking all these findings together, DHQ promises to be an effective and safe lead for the treatment of toxoplasmosis.

Capsaicin Potently Blocks Salmonella typhimurium Invasion of Vero Cells.

Salmonella typhimurium (S. typhimurium) is one of the major food and waterborne bacteria that causes several health outbreaks in the world. Although there are few antibiotics against this bacterium, some of these drugs are challenged with resistance and toxicity. To mitigate this challenge, our group explored the ethnomedicinal/herbalism knowledge about a certain spice used in Northern Ghana in West Africa against bacterial and viral infection. This plant is Capsicum chinense (C. chinense). The plant is one of the commonest food spices consumed across the world. The seed of the plant contains both capsaicin and dihydrocapsaicin. Apart from capsaicin and dihydrocapsaicin, other major capsaicinoids in C. chinense include nordihydrocapsaicin, homodihydrocapsaicin, and homocapsaicin. In this pilot work, we investigated the antibacterial activity of pure capsaicin and capsaicin extract obtained from C. chinense against S. typhimurium in vitro. Capsaicin extract showed potent inhibition of S. typhimurium growth at concentrations as low as 100 ng/mL, whereas pure capsaicin comparatively showed poorer inhibition of bacteria growth at such a concentration. Interestingly, both capsaicin extract and pure capsaicin were found to potently block a S. typhimurium invasion of the Vero cell in vitro. Taken together, we believed that capsaicin might work synergistically with dihydrocapsaicin or the other capsaicinoids to inhibit S. typhimurium growth, whereas individually, capsaicin or dihydrocapsaicin could potently block the bacteria entry and invasion of Vero cells.

Interaction of apigenin-7-O-glucoside with pyrimethamine against Toxoplasma gondii growth.

Apigenin-7-O-glucoside, a flavonoid glucoside known to inhibit cancer cell growth, fungi growth, both intra and extracellular reactive oxygen species generation, causing cell arrest and damage to the plasma membrane, was tested alone or in combination with a dihydrofolate inhibitor (pyrimethamine) against Toxoplasma gondii (T. gondii) growth. The anti-T. gondii activity was carried out using a high throughput antiparasitic drug screening cell-based assay known as 2-(2-methoxy-4-nitrophenyl)-3-(4-nitrophenyl)-5-(2,4-disulfophenyl)-2H tetrazolium, monosodium salt (WST-8) and fluorescence plate reader. The 50% effective concentration inhibition and 95% confidence interval values for individual and combination treatments against T. gondii were 0.80 (0.38-1.29) µg/mL, 1.05 (0.275-2.029) µg/mL, and 0.40 (0-1.06) µg/mL for apigenin-7-O-glucoside, pyrimethamine, and apigenin-7-O-glucoside plus pyrimethamine, respectively. Interestingly, the apigenin-7-O-glucoside plus pyrimethamine combination showed an additive inhibition effect against T. gondii growth in vitro using the fractional inhibitory concentration index method. It was discovered that the apigenin-7-O-glucoside combination with pyrimethamine had a high selectivity index 62.5, which implies 62-fold inhibition activity against the parasite versus human foreskin fibroblast cell cytotoxicity. This new combination hit is novel and will have the potential for future effective, safe, and less costly anti-Toxoplasma drug development, if its in vivo activity shows similar findings.

Anti-Toxoplasma activity of Sorghum bicolor-derived lipophilic fractions.

OBJECTIVE: Toxoplasma gondii, an intracellular zoonotic parasite, infects approximately a third of the world population. Current drugs for treatment of T. gondii infection have been challenged with ineffectiveness and adverse side effects. This necessitates development of new anti-Toxoplasma drugs. Sorghum bicolor [Moench] leaf extract has been used in African traditional medicine for the management of anemia and treatment of infectious diseases. We tested the in vitro anti-Toxoplasma inhibitory activity of S. bicolor's oil-like crude extracts and fractions against T. gondii and determined their cytotoxic effects on human host cells. RESULTS: Significant inhibitory activities against the growth of T. gondii tachyzoites were observed for the crude extract (IC50 = 3.65 µg/mL), the hexane-methanol fraction (IC50 = 2.74 µg/mL), and the hexane fraction (IC50 = 3.55 µg/mL) after 48 h of culture. The minimum cytotoxicity concentrations against HFF were 34.41, 16.92 and 7.23 µg/mL for crude extract, hexane-methanol and hexane fractions, respectively. The crude extract and fractions showed high antiparasitic effects with low cytotoxic effects. Further studies to determine synergistic activities and modes of action would provide impetus for the development of new toxoplasmosis drugs or nutraceuticals.

Racial Disparities and Preventive Measures to Renal Cell Carcinoma.

Kidney cancer ranks among the top 10 cancers in the United States. Although it affects both male and female populations, it is more common in males. The prevalence rate of renal cell carcinoma (RCC), which represents about 85% of kidney cancers, has been increasing gradually in many developed countries. Family history has been considered as one of the most relevant risk factors for kidney cancer, although most forms of an inherited predisposition for RCC only account for less than four percent. Lifestyle and other factors such as occupational exposure, high blood pressure, poor diet, and heavy cigarette smoking are highly associated with its incidence and mortality rates. In the United States, White populations have the lowest prevalence of RCC compared to other ethnic groups, while Black Americans suffer disproportionally from the adverse effects of RCC. Hence, this review article aims at identifying the major risk factors associated with RCC and highlighting the new therapeutic approaches for its control/prevention. To achieve this specific aim, articles in peer-reviewed journals with a primary focus on risk factors related to kidney cancer and on strategies to reduce RCC were identified. The review was systematically conducted by searching the databases of MEDLINE, PUBMED Central, and Google Scholar libraries for original articles. From the search, we found that the incidence and mortality rates of RCC are strongly associated with four main risk factors, including family history (genetics), lifestyle (poor diet, cigarette smoking, excess alcohol drinking), environment (community where people live), and occupation (place where people work). In addition, unequal access to improvement in RCC cancer treatment, limited access to screening and diagnosis, and limited access to kidney transplant significantly contribute to the difference observed in survival rate between African Americans and Caucasians. There is also scientific evidence suggesting that some physicians contribute to racial disparities when performing kidney transplant among minority populations. New therapeutic measures should be taken to prevent or reduce RCC, especially among African Americans, the most vulnerable population group.

In vitro activity of the interaction between taxifolin (dihydroquercetin) and pyrimethamine against Toxoplasma gondii.

Toxoplasmosis is one of the most neglected zoonotic foodborne parasitic diseases that cause public health and socioeconomic concern worldwide. The current drugs used for the treatment of toxoplasmosis have been identified to have clinical limitations. Hence, new drugs are urgently needed to eradicate T.gondii infections globally. Here, an in vitro anti-Toxoplasma gondii activity of taxifolin (dihydroquercetin) and dihydrofolate inhibitor (pyrimethamine) alone and in combination with a fixed concentration of pyrimethamine were investigated against the rapidly proliferating T.gondii RH strain at 48 hr using colorimetric assay. Pyrimethamine showed the highest anti-T. gondii activity with IC50P of 0.84 µg/ml (p > .05), respectively. The combination of pyrimethamine with dihydroquercetin gave a significant inhibitory activity against tachyzoites in in vitro with IC50p of 1.39 µg/ml (p < .05). The IC50p ranges obtained for the individual and the combination of taxifolin with pyrimethamine inhibition of parasite growth were not cytotoxic to the infected HFF and Hek-293 cell lines used. These compounds combination should be investigated further using in vivo model of toxoplasmosis.

In vitro antagonistic and indifferent activity of combination of 3-deoxyanthocyanidins against Toxoplasma gondii.

Toxoplasma gondii is a ubiquitous intracellular zoonotic parasite estimated to affect about 30-90% of the world's human population. The most affected are immunocompromised individuals such as HIV-AIDS and cancer patients, organ and tissue transplant recipients, and congenitally infected children. No effective and safe drugs and vaccines are available against all forms of the parasite. We report here the antagonistic and indifferent activity of the combination of five different formulations of pure synthetic 3-deoxyanthocyaninidin (3-DA) chloride compounds against T. gondii tachyzoites and the synergistic and additive interaction against a human foreskin fibroblast (HFF) cell line in vitro using fluorescence microscopy, trypan blue assay, and fractional inhibitory concentration index. The individual and the combined pure 3-DA compounds were observed to have effective inhibition against T. gondii parasites with less cytotoxic effect in a ratio of 1:1. The IC50 values for parasite inhibition ranged from 1.88 µg/mL (1.51-2.32 µg/mL) for luteolinindin plus 7-methoxyapigeninindin (LU/7-MAP) and 2.23 µg/mL (1.66-2.97 µg/mL) for apigeninindin plus 7-methoxyapigeninindin (AP/7-MAP) combinations at 95% confidence interval (CI) after 48 h of culture. We found LU/7-MAP to be antagonistic and AP/7-MAP to be indifferent in interaction against T. gondii growth. Both individual and combination 3-DA compounds not only depicted very strong inhibitory activity against T. gondii, but also had synergistic and additive cytotoxic effects against HFF cells. These synthetic 3-DAs have potential as antiparasitic agents for the treatment of human toxoplasmosis.

Knockdown of phosphoethanolamine transmethylation enzymes decreases viability of Haemonchus contortus.

The phosphobase methylation pathway, in which phosphoethanolamine N-methyltransferases (PMTs) successively catalyze the methylation of phosphoethanolamine to phosphocholine, is essential in the free-living nematode Caenorhabditis elegans. Two PMT-encoding genes (HcPMT1 and HcPMT2) cloned from Haemonchus contortus have been shown, by in vitro assays, to possess enzymatic characteristics similar to those of C. elegans PMTs, but their physiological significance in H. contortus is yet to be elucidated. Therefore, in this study, we endeavored to determine the importance of HcPMT1 and HcPMT2 in the survival of H. contortus by adapting the use of phosphorodiamidate morpholino oligomers (PPMO) antisense approach to block the translation of HcPMT1 and HcPMT2 in the worms. We found that PPMOs targeting HcPMT1 and HcPMT2 down-regulated the expression of HcPMT1 and HcPMT2 proteins in adult H. contortus. Analysis of the effect of HcPMT1 and HcPMT2 knockdown showed that it significantly decreased worm motility and viability, thus validating HcPMT1 and HcPMT2 as essential enzymes for survival of H. contortus. Studies of gene function in H. contortus have been constrained by limited forward and reverse genetic technologies for use in H. contortus. Thus, our success in adaptation of use of PPMO antisense approach in H. contortus provides an important reverse genetic technological advance for studying this parasitic nematode of veterinary significance.

In vitro activity of Sorghum bicolor extracts, 3-deoxyanthocyanidins, against Toxoplasma gondii.

We investigated dried red leaf extracts of Sorghum bicolor for activity against Toxoplasma gondii tachyzoites. S. bicolor red leaf extracts were obtained by bioassay-guided fractionation using ethanol and ethyl acetate as solvents. Analysis of the crude and fractionated extracts from S. bicolor using electrospray ionization mass spectrometry (ESI-MS) showed that they contained significant amounts of apigeninidin, luteolinidin, 7-methoxyapigeninidin, 5-methoxyapigeninidin, 5-methoxyluteolinidin, 7-methoxyluteolinidin 5,7-dimethoxyapigeninidin or 5,7-dimethoxyluteolinidin, based on mass per charge (m/z). When tested in vitro, the IC50s for inhibitory activity against T. gondii tachyzoites' growth of the ethanol and ethyl acetate extracts were 2.3- and 4-fold, respectively, lower than their cytotoxic IC50s in mammalian cells. Ethyl acetate extracts fractionated in chloroform-methanol and chloroform had IC50s against T. gondii that were 56.1- and 3-fold lower than their respective cytotoxic IC50s in mammalian cells. These antiparasitic activities were found to be consistent with those of the respective pure 3-deoxyanthocyanidin compounds identified to be contained in the fractions in significant amounts. Further, we observed that, the position and number of methoxy groups possessed by the 3-deoyanthocyanidins influenced their antiparasitic activity. Together, our findings indicate that S. bicolor red-leaf 3-deoxyanthocyanidins-rich extracts have potent in vitro inhibitory activity against the proliferative stage of T. gondii parasites.