Medicinal Plants as Natural Anti-Parasitic Agents Against Blastocystis Species.

BACKGROUND: Blastocystis species (sp.) are enteric parasites that live in both humans' and animals' gastrointestinal tracts. Blastocystis hominis (B. hominis) is the recognizable human isolates in clinical and diagnostic specimens. Human infection occurs via the oro-fecal route, particularly in developing areas due to the lack of sanitation and hygienic facilities. B. hominis can exist in the large intestine for weeks to years until treated appropriately. Metronidazole is the drug of choice for the treatment of Blastocystis infection. However, it induces intolerable side effects and has been shown to have teratogenic and carcinogenic potential. Several medicinal plant extracts have been experimentally tested against Blastocystis infection in comparison to currently available treatments. OBJECTIVE: Based on in vitro and in vivo studies, this article reviewed anti-Blastocystis activity of some medicinal plants. METHODS: To conduct the research for this review, Google Scholar and PubMed were the primary search engines used to find relevant literature. A total of 19 published in vitro and in vivo studies were evaluated to identify the anti-Blastocystis effects of various medicinal plants. RESULTS: Multiplication of Blastocystis parasites as well as nucleic acids and protein synthesis, all be inhibited by extracts from different medicinal plants. These natural agents have been shown to be both safe and effective when compared to the existing treatment options. CONCLUSION: Different medicinal plants can combat Blastocystis infection and could be a good substitute for metronidazole and other synthetic treatments.

IN VIVO AND IN VITRO EFFICACY OF NIGELLA SATIVA AQUEOUS EXTRACT ON BLASTOCYSTIS HOMINIS.

Metronidazole (MTZ) was the most widely accepted treatment for Blastocystis hominis (B. hominis) with high treatment failure rate, resistance and potential mutagenic and carcinogenic effects so there is urgent need to find out new, effective and safe treatment against B. hominis. The present research aimed to evaluate the therapeutic effect of the aqueous extract of Nigella sativa (NS) at different doses on B. hominis in vitro and in vivo in comparison to MTZ as a control drug. Isolates of B. hominis were obtained from patients complaining of diarrhea and abdominal pain. Isolates were cultured in egg diphasic medium (LE) for in vitro study and to adjust proper inoculating dose for in vivo study. The aqueous extract of NS at concentrations of 100 & 500 µg/ml showed a potent lethal effect on B. hominis isolates in vitro. Caecal tissue of experimentally infected and treated mice with two different doses of NS (250 & 500 mg/kg/d) were examined histopathologically and compared with that of mice infected and treated by two doses of MTZ (62 & 125 mg/kg/d) as control drug and Infected untreated mice as negative control group. Histopathological examination of infected untreated group showed all pathological degrees in the caecal tissue while infected treated one showed remission of pathological changes especially with higher dose (500 mg/kg). Present study proved that N. sativa had inhibitory effect on B. hominis in vitro and prevented cytopathic effect in infected mice inoculated orally with B. hominis.

In vivo antiprotozoan effects of garlic (Allium sativum) and ginger (Zingiber officinale) extracts on experimentally infected mice with Blastocystis spp.

Controversy surrounding the pathogenic role of Blastocystis spp. in humans and lack of well-established diagnostic criteria led to debates concerning the treatment for that organism. Furthermore, some strains develop resistance against the recommended drugs. Thus, using natural medicine has many positive aspects to address these points. In an earlier study, we addressed in vitro effect of garlic and ginger on Blastocystis spp. isolates as an alternative treatment. Accordingly, this study was conducted to evaluate in vivo activities of these two herbs on mice infected with Blastocystis spp. Antiprotozoan activities were determined by monitoring Blastocystis shedding in stools and histopathological changes of the intestine of infected mice. Additionally, assessment of the antioxidant effect (via measuring the level of malondialdehyde (MDA) production) of these herbs on the treated groups of mice was done. Also, their effects on nitric oxide (NO) production were assessed. In this work, treatment of infected mice with garlic, ginger, and nitazoxanide (NTZ) reduced the shedding of cysts significantly compared to the infected untreated group, P value ≤0.001, 0.0001, and 0.0003, respectively. As well, histopathological examination revealed that Blastocystis was frequently observed within the lumen, at the tip of the epithelium, and/ or infiltrated in an enterocyte in the infected group without treatment compared to that of the infected treated ones. Furthermore, mice infected with Blastocystis exhibited increased levels of NO (440.09 ± 3.7 vs. 276.66 ± 0.8, P ≤ 0.001) and MDA production (106.19 ± 0.43 vs. 63.06 ± 0.45, P ≤ 0.0004) compared to that of the uninfected controls. Treatment of infected mice with garlic, ginger, and NTZ reduced NO levels to 54.41 ± 1.2, 47.70 ± 1.2, and 37.43 ± 0.98 and MDA levels to 22.38 ± 0.17, 63.34 ± 3.89, and 66.76 ± 9.1, respectively. We conclude that using ginger and garlic for treatment of blastocystosis is beneficial.

Antiprotozoal activities determined in vitro and in vivo of certain plant extracts against Histomonas meleagridis, Tetratrichomonas gallinarum and Blastocystis sp.

A total of 43 plant substances provided as raw material and different kinds of extracts (aqueous, ethanol, and heptane) from 18 different organic wastes obtained from the food/feed industry were investigated for their in vitro activities against clonal cultures of Histomonas meleagridis, Tetratrichomonas gallinarum, and Blastocystis sp. Ethanolic extracts of thyme, saw palmetto, grape seed, and pumpkin fruit proved to be most efficacious. Thus, these extracts were further tested in vivo in turkeys experimentally infected with H. meleagridis by administrating the substances to the birds through the drinking water. Even though a delayed mortality was noticed in some birds medicated with the extracts of thyme, grape seed, and pumpkin fruit, all birds died or had to be euthanized the latest within 5 weeks post infection--with the exception of one bird which was probably never infected with histomonads--due to a severe typhlohepatitis indicative for histomonosis. In addition, none of the substances were able to prevent the spreading of H. meleagridis from infected to in-contact birds. Thus, these studies clearly demonstrate that in vitro studies are of limited value to assess the efficacy of plant substances against histomonosis.

Evaluation of the in vitro effect of Nigella sativa aqueous extract on Blastocystis hominis isolates.

The effect of Nigella sativa aqueous extract was evaluated against the in vitro growth of 2 different isolates of the intestinal protozoan parasite Blastocystis hominis. Different concentrations (10, 100, 500 microg/ml) of Nigella aqueous extract and metronidazole, an active standard drug for B. hominis, were incubated with B. hominis isolates in culture media at 37 degrees C. Their possible effect on B. hominnis living cell count (LCC) was assessed on Day 1, 3 & 6. The aqueous extract of N. sativa at concentrations of 100 and 500 microg/ml showed a potent lethal effect on both B. homninis isolates, but with different extent. There is no significant difference between the inhibitory effect of N. sativa and metronidazole on the LCC on the 6th day. On assessment of living cell rate (LCR) which calculate percentage rate of living cell, N. sativa at 500 microg/ml concentration has a significant inhibitory effect on both isolates. So, it is considered as the most active concentration of Nigella aqueous extract. These results prove that N. sativa aqueous extract could be useful in the treatment of B. hominis.

The effects of extracts from anti-diarrheic Thai medicinal plants on the in vitro growth of the intestinal protozoa parasite: Blastocystis hominis.

The activities of n-hexane, dichloromethane and methanol extracts from five anti-diarrheic Thai medicinal plants, Acacia catechu (Fabaceae) resin, Amaranthus spinosus (Amaranthaceae) whole plant, Brucea javanica (Simaroubaceae) seed, Piper longum (Piperaceae) fruit and Quercus infectoria (Fagaceae) nut gall were tested against the in vitro growth of fresh isolates of the intestinal protozoan parasite, Blastocystis hominis. The extracts at concentrations that ranged from 62.5 to 2000 microg/mL, were incubated with several isolates of Blastocystis hominis for 48 h. The activities were classified as killed, inhibited, moderately inhibited and not-inhibited. Dichloromethane and methanol extracts from the Brucea javanica seed and a methanol extract from Quercus infectoria nut gall showed the highest activity. At a concentration of 2000 microg/mL, the three extracts killed 82, 75 and 67% of the Blastocystis hominis samples tested and inhibited 94, 100 and 76% of them, respectively. Metronidazole, used as a reference antiprotozoan drug, at a concentration of 40 microg/mL, killed 97% of the Blastocystis hominis isolates and inhibited all samples tested at concentrations that ranged from 1.25 to 20 microg/mL.

Polypeptides associated with in vitro cyst formation of Blastocystis hominis.

The objective of this study was to characterize the polypeptides associated with cysts of Blastocystis hominis. This form is believed to be infective and plays a role in parasite resistance to anti-B. hominis drugs currently used for treatment of Blastocystis associated diarrhea. Cysts were induced through in vitro culture of the parasite in complete medium supplemented with bacterial extract with trypticase, metronidazole or doxycycline. SDS-PAGE analysis showed almost similar polypeptide patterns of parasite extracts obtained from in vitro cultured parasites before and after exposure with the three supplements. Polypeptide bands at 76, 58.5, 48, 45, 40, 38, 32, 25 and 22 kDa were constantly seen in all antigenic preparations and no specific cyst-associated polypeptide was present. However, on immunoblot analysis, 3 out of 16 blastocystosis human sera identified a cyst-associated polypeptide at 60 kDa in all parasite extracts prepared from cultures with the three supplements. In addition, there were associated morphological changes detected in these parasites stained with acridine orange and observed under fluorescence microscopy. Metronidazole induced cyst forms (reddish cells) as early as 12 hours post-exposure; more cyst production (with stronger immunoblot bands) occurred after 24 hours exposure. However, cysts rupture with release and destruction of B. hominis daughters cells occurred after 48 hours exposure. Doxycycline induced less cyst-like forms at 24 hours (weaker 60 kDa band) and less destruction of the cysts (60 kDa band still present at 72 hours post exposure). Bacterial extract and trypticase also induced cysts at 12 hours with increasing numbers up to 72 hours exposure (corresponding increase in intensity of 60 kDa band from samples harvested at 12 to 72 hours post exposure) without any sign of deleterious effect on the parasite.

Inhibition of enteric parasites by emulsified oil of oregano in vivo.

Oil of Mediterranean oregano Oreganum vulgare was orally administered to 14 adult patients whose stools tested positive for enteric parasites, Blastocystis hominis, Entamoeba hartmanni and Endolimax nana. After 6 weeks of supplementation with 600 mg emulsified oil of oregano daily, there was complete disappearance of Entamoeba hartmanni (four cases), Endolimax nana (one case), and Blastocystis hominis in eight cases. Also, Blastocystis hominis scores declined in three additional cases. Gastrointestinal symptoms improved in seven of the 11 patients who had tested positive for Blastocystis hominis.

Isolate resistance of Blastocystis hominis to metronidazole.

Isolates of Blastocystis hominis from infected immigrant workers from Indonesia, Bangladesh and infected individuals from Singapore and Malaysia were assessed for growth pattern and degree of resistance to different concentrations of metronidazole. Viability of the cells was assessed using eosin-brillian cresyl blue which stained viable cells green and nonviable cells red. The Bangladeshi and Singaporean isolates were nonviable even at the lowest concentration of 0.01 mg/ml, whereas 40% of the initial inoculum of parasites from the Indonesian isolate at day one were still viable in cultures with 1.0 mg/ml metronidazole. The study shows that isolates of B. hominis of different geographical origin have different levels of resistance to metronidazole. The search for more effective drugs to eliminate th parasite appears inevitable, especially since surviving parasites from metronidazole cultures show greater ability to multiply in subcultures than controls.