This Month in Blastocystis Research - MAR 2015

"Show me your gut bacteria and I'll tell you if you're infected with Entamoeba"

One of my 'partners in crime', science reporter Jop de Vrieze, made me aware of a study just published now by Elise R Morton and colleagues. The study appeared in bioRxiv—The Preprint Server for Biology, operated by Cold Spring Harbor Laboratory. The study is totally in line with one of the research foci in our lab.

The paper is called 'Variation in rural African gut microbiomes is strongly shaped by parasitism and diet', and can be downloaded here. The backbone in this type of research is the recognition that studies revealing a large contrast between the microbiomes of populations in developing countries and those of populations in urban industrialised areas have shown that geography is an important factor associated with the gut microbiome, but that such studies yet have to disentangle the effects of factors such as climate, diet, host genetics, hygiene and parasitism.

It's very refreshing that for once, 'parasitism' is included in such considerations. As mentioned in one or more of my previous blog posts, we have metagenomics data stongly indicating that Blastocystis colonisation is associated with certain microbial communities. As of yet, we have no idea about cause and effet, but the idea alone is immensely intriguing.

A large and a small cyst of Entamoeba coli. Courtesy of Dr Marianne Lebbad.

Now, Morton et al. have produced data that suggest that the presence of Entamoeba—another gut-associated eukaryotic genus comprising multiple species of varying pathogencitiy—is strongly correlated with microbial composition and diversity. They showed that an individual's liability to being infected by Entamoeba could be predicted with 79% accuracy based on gut microbiome composition.

The authors used 16S PCR and Illumina-based sequencing of 16S amplicons, and I could have wished that molecular assays, e.g., the 18S PCR that we have developed in our lab + associated software, had also been used to test the faecal samples from the 64 individuals enrolled in the study in order to obtain more precise data, not only on Entamoeba but also on other human-associated gut protists, such as Blastocystis.

While alpha (intra-host) diversity of Entamoeba-positive individuals was significantly higher than that of Entamoeba-negative individuals, analysis of the beta (inter-host) diversity revealed that gut communities across Entamoeba-positive individuals were more similar than across Entamoeba-negative individuals, suggesting that, as alpha diversity increases, there are fewer potential stable states for individual gut communities, or that infection by Entamoeba drives changes in the microbiome that are dominant over other factors.

Right—this is Entamoeba, I know, but in principle, the type of analyses that were performed in the present study could be applicable to Blastocystis, Dientamoeba, and other gut parasites, which may help us understand their role in health and disease. Are these parasites able to influence gut microbiota? Can they be used for gut microbiota manipulation? Or do they only infect people with certain microbiota profiles? Time will show... maybe.

For those of you who would like to read more about what is shaping our microbiomes and how the gut microbiota may impact on our gastrointestinal health, I recently did a couple of blog posts for United European Gastroenterology (UEG) Education that might be of some interest:

Are we finally saluting the fungal kingdom as a co-ruler of GI health and disease?

The intestinal microbiome—Rosetta Stone or Tower of Babel?


Reference:

Morton ER, Lynch J, Froment A, Lafosse S, Heyer E, Przeworski M, Blekham R, Segurel L.
Variation in rural African gut microbiomes is strongly shaped by parasitism and diet. bioRxiv doi http://dx.doi.org/10.1101/016949 - accessible here.

This Month in Blastocystis Research (FEB 2015)

Before heading off to visit dear colleagues at the Public Health Agency of Sweden tomorrow morning, I thought I'd do a quick 'This Month...' post.

Tropical Parasitology has published a paper by Elghareeb and colleagues on  'Laboratory Diagnosis of Blastocystis in Diarrheic Patients'. I was asked to do a Guest Commentary on their paper, and if your're interested you can download my comments here for free (html version). The paper by Elghareeb et al. should also be free for download at the website.

I have been very lucky to work together with Dr Prashant K Pandey and his colleauges in Pune, India. Together we just published the first data on Blastocystis subtypes ever to appear in India for what I know. We subtyped Blastocystis in a cohort of healthy Indian individuals, and found ST1 and ST3 in 27/100 adult individuals tested, while other common subtypes, ST2 and ST4, were absent. Remarkably, ST3 was seen in all positive individuals, while ST1 was seen only in mixed infections. The strains (alleles) found in India were no different to those found in for instance Europe.

There is a paper out by Rossen and colleagues from The Netherlands showing that Blastocystis is relatively uncommon in patients with active ulcerative colitis (UC) and significantly less common in UC patients (13.3%) than in healthy individuals (32.5%). This is completely in line with data that we generated in Denmark a couple of years ago. In fact, at two separate occasions we have been able to look into patients with inflammatory bowel disease. In both cases (one study has been submitted for publication), hardly any Blastocystis was found in patients with Crohn's disease, while a few patients with UC were positive; however, mostly patients with inactive disease appeared to have Blastocystis, while those with flare-ups were negative. Therefore, the influence of dysbiosis on Blastocystis colonisation should be subject to further scrutiny.

A lot of action goes on at the official website for the 1st International Blastocystis Symposium in Ankara in May, with exactly three months to go! Why not take a minute to browse the programme for the Pre-Symposium Course and the Scientific Programme for the actual Symposium? Please go here to familiarise yourself with the new content. 
Also, conference abstracts are pouring in, - did you submit yours yet?

References

Elghareeb AS, Younis MS, El Fakahany AF, Nagaty IM, & Nagib MM (2015). Laboratory diagnosis of Blastocystis spp. in diarrheic patients. Tropical Parasitology, 5 (1), 36-41 PMID: 25709951

Stensvold, C. (2015). Laboratory diagnosis of Blastocystis spp Tropical Parasitology, 5 (1) DOI: 10.4103/2229-5070.149885  

Pandey PK, Verma P, Marathe N, Shetty S, Bavdekar A, Patole MS, Stensvold CR, & Shouche YS (2015). Prevalence and subtype analysis of Blastocystis in healthy Indian individuals. Infection, Genetics and Evolution: Journal of Molecular Epidemiology and Evolutionary Genetics in Infectious Diseases PMID: 25701123

Rossen NG, Bart A, Verhaar N, van Nood E, Kootte R, de Groot PF, D'Haens GR, Ponsioen CY, & van Gool T (2015). Low prevalence of Blastocystis sp. in active ulcerative colitis patients. European Journal of Clinical Microbiology & Infectious Diseases: Official Publication of the European Society of Clinical Microbiology PMID: 25680316

Updated programme for the 1st International Blastocystis Symposium + Pre-Symposium Course programme

There has been changes to the programme for the 1st International Blastocystis Symposium (May 28-29), please go here to see the updated programme.

Also, the Pre-Symposium Course programme (May 27) can be viewed here.

We will be absolutely spoilt with speakers!

Please do not forget to visit the official website for updates on the social programme plus abstract/registration deadlines.

Scientific Programme for the 1st International Blastocystis Symposium in Ankara, May 2015, is now available!

The Scientific Programme for the 1st International Blastocystis Symposium in Ankara, May 2015, is now available! Please note that the programme may be subject to change.

Click here to view it!

Go here to view the official website for the symposium.

This Month in Blastcystis Research - JAN 2015

I'm going to dedicate this blog post entirely to the upcoming 1st International Blastocystis Symposium.

I'm not sure how much advertising there is for this congress (our budgets are limited), but the fact that we are already receiving abstracts is a good sign! Abstacts may be submitted until April 1st, 2015. Please note that the 'early bird' registration discount expires at the 15th of February.

You will find the online abstract submission form here.

If you think about going but have not paid a visit to the official conference website, I recommend you to do so, clicking this link. You'll hopefully find most if not all the information that you'd be looking for, and there's a lot to be learned. Please also make sure to browse the social programme in order to be able to make appropriate arrangments.

It's a two-day symposium, running from the 28th to the 29th of May, 2015. Moreover, on the 27th, there will be an all-day workshop on various diagnostic and molecular epidemiological aspects, including a barcoding (subtyping) course. There will be more info on that very soon, - please keep an eye on the website.

We are doing all we can to attract scientists with vast experience in Blastocystis research to cover the floor with exciting and stimulating talks, and I think we're doing more than OK. Some of the speakers will be writing up reviews on their respective topics, and these reviews will appear in a special themed issue in Parasitology International.

There will be a quite a few prizes for best talks and posters, etc., thanks to ELSEVIER among others.

It will be one-track symposium, and the first day will focus mostly on some fundamental topics, such as genomics and biochemistry, while the next day will include talks on clinical and diagnostic data.

It's my clear impression that main organiser Dr Funda Dogruman-Al is working 25 hours a day to make everything come together, and Dr Hisao Yoshikawa has also already invested a lot of energy.

Again: please note that early registration will close at the 15th of February, and abstract submission deadline has been extended to April 1st, 2015.

Looking very much forward to seeing you there!

This Month In Blastocystis Research - Welcome 2015! Edition

2014 is coming to an end, and I'm looking very much forward to 2015, which will be the year of the 1st International Blastocystis Symposium. We are busy making all the preparations, and we hope that many of our colleagues will put priority to the event. For more details, please look up www.blastomeeting2015.com

Just before Christmas, I handed in a funding proposal that will hopefully turn out to be successful; if it is, it will enable me to fund three post docs/experienced scientists who will be working on Blastocystis genomics, animal experimental model work, and Blastocystis in the gut microbiota context, all three projects serving to identify the role of the parasite in health and disease. I've never been crossing my fingers as much as I am now... I will only know in 6 months from now whether I've been successful or not, so it's a long wait!

Two new papers have come to my attention. One deals with identification of a new host for Blastocystis - the brown bear (Ursus arctos); the other one is about the finding of Blastocystis in a case of appendicular peritonitis.

Ursus arctos - new host for Blastocystis (Source).

So far, Blastocystis has been identified in many larger mammals. Mostly synanthropic animals and animals in captivity have been sampled. Meanwhile, we know little about Blastocystis in 'the wild', in sylvatic animals. A team from Slovakia took to collecting 16 stool samples from brown bears in the Poloniny National Park; eight of them were positive for Blastocystis ST3, a subtype found mainly in primates, but also occasionally in a variety of other hosts. Interestingly, the team reported that the ST3 sequence amplified from the samples were identical to a sequence that we have deposited in GenBank (HQ909889) isolated from a human. We have studied ST3 from human and non-human primates, who are both common hosts of ST3. However, we have also come to realise that ST3 strains in humans differ genetically from ST3 of non-human primates, i.e., it appears not to be the same strains circulating in human and non-human primates. I wouldn't have been surprised to find a bear-specific allele of ST3, but apparently, the bears in the study harboured ST3 identical to ST3 in humans.

Blastocystis is a frequent finding in large mammals such as cows, pigs, camels, and, possibly, bears. One of my colleagues recently asked me: How about horses? A parasitologist herself with a vast experience in both human and animal parasitology, she told me that she did not remember coming across Blastocystis in horse manure. And we wondered why... Assuming that exposure does not differ between horses and other larger, synanthropic mammals, the reason why horses might not harbour Blastocystis may be due to factors such as diet, digestion, microbiota, and maybe competing eukaryotic intestinal fauna. However, I believe that horses have been infrequently sampled, and so we need more data to be able to 'ruminate' further on this.

Since parasites such as Dientamoeba and Blastocystis are lodged in the colon, including the coecum, their potential role in e.g. appendicitis and related conditions could warrant scrutiny. Fréalle and colleagues reported a case of appendicitis with suppuration into the recto-uterine pouch and reflex ileus in a 9-year-old girl returning to France after a 1-month stay in Casablanca, Morocco, admitted to Lille University Hospital, France. The authors were unable to detect enteropathogenic bacteria such as Salmonella, Shigella, Campylobacter, Yersinia enterocolitica, adenovirus, and rotavirus, but "multimicrobial flora" was detected in the peritoneal liquid and recto-uterine pouch. It would seem plausible to me that Blastocystis ended up in the peritoneum by chance along with other enteric micro-organisms, although the etiology of the inflammation remains uncertain. Appendicitis usually develops following obstruction of the lumen of the appendix, either due to hyperplasia of lymphoid tissue or faeces becoming incarcerated in the mucosal tissue; however, other causes are have also been observed, e.g. pinworm infection.

It would like to finish off by a ***Happy New Year***, at the same time celebrating page view #300,000!

References:

Fréalle, E., El Safadi, D., Cian, A., Aubry, E., Certad, G., Osman, M., Wacrenier, A., Dutoit, E., Creusy, C., Dubos, F., & Viscogliosi, E. (2015). Acute Blastocystis-Associated Appendicular Peritonitis in a Child, Casablanca, Morocco Emerging Infectious Diseases, 21 (1), 91—94 DOI: 10.3201/eid2101.140544

Valenčáková, A., Kandráčová, P., Kalinová, J., Danišová, O., Hasajová, A. (2014). Detection of Blastocystis hominis subtype 3 in the brown bear in the Slovak Republic. Folia Veterinaria, 58, 3: 175—178. http://www.uvlf.sk/sites/default/files/folia-veterinaria/fv_3_14_web.pdf#page=32

This Month in Blastocystis Research (NOV 2014): Blasting Blastocystis Edition

The 'This Month' post is triggered by a paper emerging in the journal Gut Pathogens describing a clinical pilot study on the efficacy of triple antibiotic therapy in Blastocystis positive IBS patients. The article is free for download here. The triple therapy consisted of fourteen days of diloxanide furoate 500 mg thrice daily, trimethoprim/sulfamethoxazole (cotrimoxazole) 160/80 mg twice daily, and secnidazole 400 mg thrice daily. Six of ten patients achieved eradication. Please have a look at the paper for more information.

Sometimes I get contacted by people who have been trying to get rid of Blastocystis. And on the odd occasion, I receive accounts that I'd like to share - completely anonymously of course - hoping that the information will benefit those interested and that I can stimulate interest in the field a bit. But also because I think that sometimes people expose themselves to MASSIVE antibiotic treatment that might cause more harm than good (microbiota perturbation).

Below you'll find three examples dealing with the eradication of Blastocystis. Kindly note that this is not a post on IF or WHEN one should seek to eradicate Blastocystis, and please also note that this should not be interpreted as 'medical advice'.

I obtained permission from the patients in Examples #1 and #2 to share their stories, which have been eidted slightly for clarity.


Example #1:

"Two years ago, I was declared positive for Blastocystis after traveling to India. My symptoms included abdominal pain, weight loss, rectal itching, constipation or diarrhoea (yes, it's supposed to be 'or') -
I could be constipated for 7-10 days and then have a big diarrhoea "in one go". I took:

• January 2012: Fasygin (tinidazole) twice daily for 3 days => still positive after treatment.

• February 2012: Bactrim Forte (co-trimoxazole) three times daily for 10 days => still positive after treatment.

• March 2012: A combination of Bactrim Forte (cotrimoxazole) three times daily for 10 days and Tiberal (ornidazole) twice a day for 5 days. Then, Intetrix (tiliquinol) twice a day for 10 days => still positive after treatment. 

• May 2012: first-line-treatment from Australia = combination of Bactrim Forte (co-trimoxazole) twice a day for 10 days / Secnidazole 3 times a day for 10 days / Diloxanide Furoate 3 times a day for 10 days  => 3 consecutive Blastocystis-negative stools (tests in July 2012).

Then no symptoms anymore till February 2014, when the same symptoms came back, and I was stool-positive for Blastocystis. I took:

• March 2014: Flagyl (metronidazole) 3 times a day for 10 days, then a combination of Paromomycin 6 times a day for 10 days / Doxycyclin 2 times a day for 10 days / Bactrim Forte (co-trimoxazole) 3 times a day for 10 days / Saccharomyces boulardii 4 times a day for 10 days.

• Test in April: Stool-positive for Blastocystis.

• May 2014: Nitazoxanide 2 times a day for 10 days / Furazolidone 3 times a day for 10 days / Secnidazole 3 times a day for 10 days.

I'm now in a period with phases (after pain during 2/3 weeks, no more pain during 2/3 weeks, then pain again, then no more...). All tests were carried out in the same way at the same lab."

The patient's current Blastocystis carrier status remains unknown. However, the present story demonstrates the ferocious concoctions taken into use to clear Blastocystis.

Example #2:

"Metronidazole for 10 days failed, then, a few months later, I tried metronidazole plus paromomycin for 10 days, flanked with 3 doses of nitazoxanide and one dose of albendazole, and I am now convinced that that heavy chemo-treatment worked, since several tests, including my most recent one, have been negative since that multi-drug treatment. Some lingering mild symptoms, possibly related, or not, kept me wondering, but I am now convinced the bugs are gone." 

Example #3:

The last story is my own, and it describes how I inadvertently lost my Blastocystis strain. Please note that I have no financial interests to disclose. Moreover, I don't believe that I ever suffered symptoms from Blastocystis colonisation.

I spent most of my childhood in the countryside in Denmark. Moving down from Norway, my parents had bought a small farm, although they were not farmers. We did have some animals though, e.g. cats, a dog, chickens, sheep, and at some point even a couple of tortoises. I don't think I ever received antibiotics throughout childhood, except from once when being hospitalised due to surgery back in 1975. In 1990, I sustained a severe bicycle accident and was admitted to hospital; I believe I must have received some antibiotics back then, too. I have travelled extensively, and spent several months in e.g. Laos and Thailand in 2003/2004, three weeks in India in 2007, etc.

I started testing myself for Blastocystis only in 2009, and just like at least 20% of mankind, I was positive. Since then I re-checked myself every now and then, and I was always positive for the same strain (evidenced by DNA analysis), ST1 allele 4. However, in early 2014 I had major dental surgery, and I was prescribed tablets three times daily for six days. These tablets contained amoxicillin (500 mg) + the beta-lactamase inhibitor clavulanic acid (125 mg). A couple of weeks after completing antibiotic treatment, I tested myself a couple of times, and Blastocystis had vanished! Also today there is no sign of it...

I wish that I had been able to map my intestinal bacterial communities both before and after treatment to identify the effect of the drugs on my gut microbiota, thinking that Blastocystis disappeared due to microbiota perturbation rather than a direct effect on the parasite. I don't remember changing anything in my diet around the time of 'conversion'; only thing that I can think of is that - for a reason I no longer remember - I took to ingesting large amounts of freshly chopped ginger and consumed quite a few cups of 'ginger tea' (basically just a ginger infusion) around that time. But since ginger consumption is very common in parts of the world where Blastocystis is common, I don't attribute eradication to ginger consumption. I may be wrong of course.

For now, I just wanted to post the information and let the examples speak for themselves.

Reference: 

Nagel R, Bielefeldt-Ohmann H, & Traub R (2014). Clinical pilot study: efficacy of triple antibiotic therapy in Blastocystis positive irritable bowel syndrome patients. Gut pathogens, 6 PMID: 25349629