Molecular Tools for Helminth Control and Elimination: Time to Get them Out of the Laboratory and itno Programmes and Policies?

By Corrado Minetti

On my way back from Ghana, where we have been testing the molecular protocols for the detection of filarial parasites in mosquitoes, in the laboratory of Mike Osei-Atweneboana at the Council for Scientific and Industrial Research (CSIR) in Accra; I had some thoughts about how far molecular diagnostics has come but also questioned how can we make it a sustainable reality to assist effectively in disease control and elimination.

corrado-molecular-diagnostics-blog-post-08-11-2016

DNA extraction from pooled mosquitoes for the detection of filarial worms (on the left) and an example of amplification of parasite DNA (+) with the LAMP method (on the right) (Photo: Corrado Minetti)

In order to achieve the goals of the London Declaration on Neglected Tropical Diseases for the effective and sustainable control and long term elimination of Lymphatic Filariasis, onchocerciasis, soil-transmitted helminthiasis and schistosomiasis; the deployment of appropriate diagnostic tools is crucial at every stage of these disease control and elimination programmes from initial mapping to post-elimination surveillance. With the rapidly changing epidemiological scenario of these diseases due to the scaling up of mass drug administration, and the push towards more sustainable and cost-effective multi-disease interventions, the implementation of more sensitive and cost-effective diagnostic tools is a priority well recognized and advocated by the World Health organization.

Molecular diagnostics tools, including (multiplex) real-time polymerase chain reaction and more recent isothermal amplification assays such as loop-mediated isothermal amplification and recombinase polymerase amplification do offer increased sensitivity compared to traditional approaches but they are yet to be used in control and elimination programmes due to their cost and technical requirements. There are various gaps that need to be highlighted and solved in order to allow these approaches to become potentially embedded into disease control programmes & policies, and to inform decision-making.

In order to identify these much-needed gaps, we have recently published a review paper where we compared the features of published real-time PCR and isothermal amplification assays for the detection of Lymphatic Filariasis, onchocerciasis, soil-transmitted helminthiasis and schistosomiasis in clinical and vector/intermediate host samples. Despite the availability of a wide range of assays for both patient diagnosis and xenomonitoring (parasite detection in insect vectors or snails), little or no research has been devoted to estimate the real costs and logistics of implementing these approaches on a wider scale for control and elimination. We highlight the need for a major focus on the implementation aspects of these tools in developing countries, and how barriers for their full adoption in resource-poor settings could be overcome. Key issues are the technical requirements and the related need for capacity building, the abatement of costs and the economic sustainability of molecular screening over time. For example, diagnosing multiple parasites from the same clinical sample can heavily reduce the number of samples that a community may need to provide, resulting in a far less invasive procedure for the communities, as well as reducing significantly the cost of processing. A multi-disease approach to diagnostics will certainly benefit the health system as well, both logistically and economically.

Writing this review paper has been extremely valuable to get a clearer picture of the progress in the field so far and to identify the best and most cost-effective diagnostic approaches for our project. In a broader sense and within the COUNTDOWN research consortium, we hope this review could serve as a starting point of discussion in the NTDs control and elimination community, leading to a more comprehensive analysis of what molecular diagnostics can offer and how we can make sure these tools can finally get out from the laboratory becoming embedded into policy, to strengthen disease control and elimination programmes and the health system itself.

Find more information on COUNTDOWN’s activities visit us here.

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Onchocerciasis in Three Decades Part II: Building the Next Generation of Parasitologists

By Prof Samuel Wanji & Pamela Bongkiyung

This post continues the discussion with Prof. Wanji on the strides undertaken towards the control of Onchocerciasis.

In this segment, he discusses his hopes for a drug that could not only work against Onchocerciasis but also Loa Loa without causing harm to the human host. He highlights the foundation he has laid down to help the next generation of parasitologists, empowering them to seek solutions to problems endemic to their respective environments.

Question: Understanding that we have just the one tool, does it complicate matters or make it worse that there is Loa Loa in all of this to deal with?

Prof. Wanji: Of course because that tool presents a problem with Loa Loa. Just because it is one tool remains a problem but the fact that in some areas you cannot use it, makes it more difficult. It presents a double problem and sometimes people often talk of double penalty with regards to Loa Loa. That is something which was discovered recently that the area where Loa Loa exists, the tool which was being used to map Lymphatic Filariasis can have false positivity. It can be positive not because of Lymphatic Filariasis but because of Loa Loa. Loa Loa has been inflicting a double penalty for the control of lymphatic filariasis and onchocerciasis in the central African region.

Q: Is there a drug to mitigate Loa Loa as well as Onchocerciasis without causing any adverse reactions.  It appears there isn’t the one stone that can kill two birds but is there one that can kill each bird at a time?

Prof. Wanji: Unfortunately we do not have a drug that can kill Loa Loa safely for now. That is a major problem. In our laboratory, we are trying to develop experimental models but at the in vitro and nvivo that can help screening and developing drugs for Loa loa. If we can have drugs that can kill Loa Loa alone, that already will be a great achievement. If we could have a good drug that could kill Loa Loa in a safe manner for the host (the human) and also kill onchocerciasis, that will be a tremendous achievement. Actually, we were handicapped, we don’t have a solution for Loa Loa as we stand today.

Q: This problem is very endemic in the Central Africa region. At the level of education especially within the universities, are there any incentives to get students more interested in research work for diseases that affect their environment? And is there available training to enable them conduct research that could possibly bring a much needed solution in the future?

Prof. Wanji: I think the teaching of parasitology (because all of this belongs to microbiology and parasitology), is difficult because most of our African students are likely to learn computer science, business administration and Economics.

We, at the University of Buea for the past five years in the Department of Microbiology and Parasitology have created postgraduate programmes that are specifically oriented to Neglected Tropical Diseases (NTDs). We have an MSc in Molecular Parasitology and Vector Biology, MSc in Microbiology, MSc in Epidemiology and Control of Infectious Diseases, a PhD in Cellular and Molecular Parasitology and a PhD in Microbiology. And if you look at our PhD and Master degree programmes, we have two arms: we have the arm of research and development which includes the MSc in Molecular Parasitology and vector biology, PhD in Cellular and Molecular Parasitology.

They give the basis for students to be involved in developing new tools for diagnosis of parasites, new tools for understanding the immune responses and developing vaccines, new tools for developing new drugs. The thing is that our students have to understand and be part of the momentum. But we also work with epidemiology and control orientation in the programme. We want them to be ready to uptake the product of the research to the end users because the epidemiology conceives control programmes, monitors and evaluates them. We want them to be able to say, this is what we have achieved, this is where we are having difficulties or encountering bottlenecks and we can do this to change.

We have conceived a programme to address the problem of NTDs and infectious diseases at large maintaining a parasitology focus.

Q: What legacy would you want to leave behind?

Prof. Wanji: Difficult question. Legacy is difficult because it is not easy to praise yourself. I would like to be known as somebody who conscious of the dimension and width of NTDs devoted part of his life to contribute, to understand and fight those tropical diseases; by teaching students about NTDs, creating and developing research capacity that has contributed to the training of those students; by participating in research work that has contributed to change people’s lives, contributing to fight in the field of those diseases and by anticipating what the future will be in my discipline; re-orienting the teaching programme at the university to get African students to be more proactive in the solution to those problems created by  NTDs.

Thank you for those last words Prof. Wanji.

 

Onchocerciasis in Three Decades: Through the Lens of Prof. Samuel Wanji

By Prof Samuel Wanji & Pamela Bongkiyung

Prof. Samuel Wanji is Head of Department for Microbiology and Parasitology at the University of Buea, Cameroon. He is also Executive Director of the Research Foundation in Tropical Diseases and Environment, Buea. He heads the COUNTDOWN programme’s partnership with the University of Buea. He has worked extensively on Onchocerciasis control and been instrumental in its control. We caught up with him while he was on a visit to the Liverpool School of Tropical Medicine.

Pamela: Hello Prof. Wanji. Thank you for taking the time to talk to us. How long have you worked on Onchocerciasis?

Prof. Wanji:  My journey with onchocerciasis started as far back as 1988. We could be talking of almost 30 years of research in onchocerciasis. Everything started with an experimental model when I was doing my postgraduate studies in Paris, France. And later on, I remained within Filariasis/Onchocerciasis for my research and university career. So actually, it has been a very faithful relationship with research and onchocerciasis throughout my life as a student and a worker both for teaching and research.

P: In the almost three decades that you have worked on various projects in relation to onchocerciasis, what changes have you observed?

PW: Yes, there have been changes. I started almost when Ivermectin was introduced as a new tool for the control of Onchocerciasis. And we witnessed the huge impact Ivermectin has made in the control of Onchocerciasis through the African Programme for Onchocerciasis Control – APOC. We were among the researchers involved in mapping the disease across Africa for the implementation of the control. We also witnessed the decline of the disease in many of foci where the endemicity was really high.

But what has been my major contribution for the control of onchocerciasis came from another filarial parasite Loa Loa. Originally, nobody knew that Loa Loa could cause a problem for the onchocerciasis control. So, when it was established that some people taking Ivermectin could develop severe adverse events and could even die, the link was established with Loa Loa and it became a priority to know those places where the endemicity of Loa Loa that is dangerous for the intake of Ivermectin was.

We participated in those early days around 2000 to develop the rapid assessment procedure for Loa Loa which was later on extended to 15 African countries, where we coordinated the mapping exercises in those countries, to generate the first map for Loa Loa in Africa. This map was operationally very useful in saying this place is very dangerous, this place is safe. And that is how we contributed heavily to the control of onchocerciasis in those areas where there was co-endemicity with Loa Loa.

Besides that, at the level of the laboratory we developed an experimental model in baboons to understand the mechanisms of encephalopathy due to Loa Loa with the treatment we have with Ivermectin. Today we know that the microfilaria are massively killed and they block the micro capillary of the brain and that is how people almost get killed. People become withdrawn and they go into coma because of such. We know that mechanism and we even have an idea of how such things could be prevented with ivermectin and aspirin. We have worked a lot through Loa Loa to see how the control of onchocerciasis could be safe in the forested areas of Africa.

P: What are the major impediments to controlling Onchocerciasis not just in Cameroon as you seem to have worked extensively across Africa, but also how does Loa Loa add to these impediments that you might have encountered in your work?

PW: As we know Onchocerciasis is a Neglected Tropical Disease. So just by that fact, it is already an impediment in controlling such disease. Less attention was paid to it. Fortunately for the past ten years there has been a lot of momentum around Neglected Tropical Diseases and Onchocerciasis has also benefitted from that.

As you can imagine that for almost 30years, the only drug that has been used for the control of Onchocerciasis has been Ivermectin. Ivermectin does not kill adult Onchocerciasis besides of the fact that it creates problems with Loa Loa in areas where the two diseases co-exist, Ivermectin kills only Microfilaria, the children of filarial of Onchocerca Volvulus (O. Volvulus).

So you need to take the drug for longer than 15years, to expect getting rid of the disease. And that is a very long time because people can easily go into fatigue. If the resources are not properly mobilised people may not have resources to sustain such long term control.

One probable consequence of such impediment has been the suspicion (and I will call it suspicion because there has been a lot of controversy around it) of sub-optimal response of the worms to Ivermectin. That means when you have a long term pressure of the same one medication on a parasite, the parasite may develop a strategy to not be sensitive to that drug anymore. And that has been a very shortcoming of onchocerciasis control to know that instead of having two or three drugs to play with, only one drug exists.

It is only of recent that there is a lot of work going on to develop microfilaria drug that will kill the adult worm. That is why in many areas we are really doubting how the elimination will be feasible. Those areas where the transmission was very high or is very high, we are almost sure that Ivermectin alone cannot do the job. We need alternative strategies. That is why you have been hearing about Doxycycline which was developed recently. Doxycycline can kill adult Onchocerca Volvulus but it needs at least 4weeks to do that job. The four weeks treatment is far better than 15years yearly treatments because with 15yrs yearly treatment you may have fatigue effect more than 4wks continuous treatment. At the level of the public health people think it is not suitable to have a regimen of 4wks, they insist on shorter regimens.

Here at the Liverpool School of Tropical Medicine, we have the AWOL consortium which is developing a shorter regimen of Anti-Wolbachia. Doxycycline is an anti-wolbachia. It is a drug that kills the bacteria that lives in a worm. Because that bacteria is starkly associated to the worm, they exchange some functions. So if you kill the bacteria, the worm also dies. It is an indirect effect. So the AWOL consortium is developing a shorter regimen of antibiotics that can do the same job like doxycycline. We hope that doxycycline will play its own role in the elimination of onchocerciasis or anti-wolbachia drugs globally. But we are expecting contributions from other drugs like Flubendazole which is in the pipeline.

Globally to answer your question, the impediment has come from the fact that we are dealing with only one tool, for the control of onchocerciasis.

To be continued…

Old dog, New Tricks? Assessing the Potential of Integrating Focal Vector Suppression with Drug Cure to Control and Eliminate River Blindness

By Louise Hamill

Onchocerciasis, also known as river blindness, is one of the vector-borne neglected tropical diseases (NTDs); in this case, transmitted by many different species of black fly. The majority of infections (99%) occur in sub-Saharan Africa. The disease was previously also found in South America but is now thankfully close to total elimination; with only a few isolated, extremely remote areas still to be verified disease-free. The aim for Africa is to achieve continent-wide interruption of transmission by 2025.  Current control of river blindness in Africa, which, as well as blindness, leads to debilitating, disfiguring skin pathology, is based upon the mass delivery of ivermectin to entire populations in endemic areas. Ivermectin kills microfilariae in the skin, but has no significant effects on adult worms. This necessitates repeated rounds of ivermectin mass delivery for a period of 12 – 15 years, with sustained high coverage of the at-risk population essential for successful disease control and eventual elimination.

This approach has had notable successes in several areas, led by the efforts of the African Programme for Onchocerciasis Control (APOC). However in other areas, the impact of sustained delivery of ivermectin for fifteen years, and in some areas more than two decades, has yet to result in the predicted interruption of transmission. Furthermore, where the eye worm Loa loa and onchocerciasis occur together, mass delivery of ivermectin cannot be easily rolled out. Ivermectin causes unwanted side effects, and in rare cases death, in individuals infected with L. loa as the drug rapidly kills this parasite. L. loa is common in large swathes of West and Central Africa, allowing onchocerciasis to endure in these areas, where many people are still infected and transmission of both pathogens is actively taking place, despite ongoing control efforts. Clearly there is no one-size-fits-all approach to curtail river blindness, and there is a need to seek alternative strategies to ivermectin-based control in areas where river blindness and L. loa overlap.

In the nineteen seventies and eighties the WHO onchocerciasis control programme, OCP, ran an extremely successful vector control strategy against onchocerciasis in savannah areas of West Africa. This programme relied exclusively on aerial application of larvicide to kill black fly larval as they resided in rivers and streams. It is estimated this past use of vector control prevented 600,000 cases of blindness and prevented 40 million people being infected. The scale of this undertaking, including the huge financial cost and human resource needed, means that the use of mass vector elimination as a tool for onchocerciasis control is very much consigned to the history books. Before turning the page completely on this chapter of onchocerciasis control, are there any lessons to be learnt from this “old dog”?

The COUNTDOWN meeting on Focal black fly Control in Cameroon

This is exactly the question we set out to debate when COUNTDOWN convened a technical advisory panel at the Liverpool School of Tropical Medicine on 22nd of July 2016. Although mass vector control is out of the question, is there any way in which short-term, localised approaches can be used to augment and complement existing strategies?

Vector Meeting

Attendees at the COUNTDOWN meeting on Focal Black fly Control, from Left to Right: Professor Graham Matthews, Professor Rory Post, Dr Frank Walsh, Didier Bakajika, Dr John B. Davies, Dr Louise Hamill, Dr Hans Dobson, Dr Joseph Turner, Professor María-Gloria Basáñez, Professor Mark Taylor, Isobel Routledge, Professor Russell Stothard, Professor Robert Cheke. Not pictured; Professor Janet Hemingway, Dr Lisa Reimer.

Previous work in South West Cameroon by members of the COUNTDOWN consortium indicates that ten years of ivermectin delivery in our study area has not had the expected impact on disease prevalence. The average community-level of skin microfilaria prevalence stands at 52.7 percent, with the infection intense even in children under ten years of age. Additional work in South West Cameroon found current adherence to ivermectin mass delivery by local residents is not adequate to achieve onchocerciasis control. This is an area where alternative and complementary strategies are urgently needed.

The COUNTDOWN consortium has proposed that larvicidal treatment of vector breeding sites at the same time as testing and treating the human population with doxycycline could offer a complementary onchocerciasis control strategy. This two-pronged approach, it is hoped, will have a greater impact on disease transmission than using either technique in isolation. Doxycycline targets “friendly” bacteria living within the adult onchocerciasis worms, resulting in a significant shortening of their lifespans and giving doxycycline a very different mode of action to ivermectin. Since L. loa does not harbour the same bacteria, individuals co-infected with L. loa who take doxycycline will not suffer the same side effects as can happen with ivermectin. From the evidence above, it is clear to see ivermectin mass delivery has not had the desired impact on disease prevalence over the past ten years in this area of South West Cameroon; could targeted vector suppression jump start the path to onchocerciasis control?

At the meeting, debate revolved around the factors influencing choice of larvicide; when, how often & for how long the larvicide should be applied; the most suitable sampling methods to monitor impact of the larviciding on adult and larval black fly; and how best to undertake monitoring of the impact of insecticide application on non-target organisms. The optimal timing of any vector suppression to best amplify the impact of the community test-and-treat strategy is crucial. The way ahead is far from straight forward, highlighting the importance of rigorously assessing the evidence and our proposed strategy in this way. Although the use of localised vector control against black flies is not a new proposal, there is little information on how this could be implemented against free-living black fly larvae.

Where next?

The control and elimination of NTDs in Africa has repeatedly been in the post-millennium development goals policy spotlight, with (among others) the WHO roadmap to elimination, the London Declaration on NTDs and recently the launch of the Expanded Special Programme for Elimination of NTDS (ESPEN). Similar to the situation for lymphatic filariasis, scale-up of mass delivery of ivermectin will not be enough to achieve the London Declaration 2020 targets for onchocerciasis control and elimination. The use of both doxycycline and focal vector suppression are separately recommended by WHO and APOC as alternative onchocerciasis control strategies, to accelerate progress towards onchocerciasis control, however as relatively new control strategies evidence on their implementation is scarce and evidence on integrated, dual-strategy implementation is wholly absent. The specific contexts in which these tools could be successfully implemented together are unclear.  Going forward with our onchocerciasis work in Cameroon, COUNTDOWN’s focus is consolidation of the evidence gathered at the vector control meeting to assess the possibility of implementing localised vector suppression as an adjunct to existing and alternative control and elimination strategies. This will bridge vital evidence gaps and provide clarity on if and where these techniques can be used, and the optimal conditions in which to implement them.