PLoS Negl Trop DisplosplosntdsPLoS Neglected Tropical Diseases1935-2735Public Library of ScienceSan Francisco, USA07-PNTD-EC-0176R110.1371/journal.pntd.0000076Expert CommentaryInfectious Diseases/Neglected Tropical DiseasesIvermectin Resistance in Onchocerca volvulus: Toward
a Genetic BasisIvermectin Resistance in
OnchocercaLustigmanSara1*McCarterJames P.23Laboratory of Molecular Parasitology, Lindsley F. Kimball Research
Institute, New York Blood Center, New York, New York, United States of
AmericaDivergence Inc., St. Louis, Missouri, United States of
AmericaDepartment of Genetics, Washington University School of Medicine, St.
Louis, Missouri, United States of AmericaYameyGavinAcademic EditorPLoS, United States of America* E-mail: slustigman@nybloodcenter.org
SL has received research grants for the development of anti-Onchocerca vaccines
and macrofilaricides. JPM is a share-holder and employee of Divergence, Inc., a
company that works on parasite control in plants, animals, and humans.
102007308200711e762007Lustigman, McCarterThis is an open-access article distributed
under the terms of the Creative Commons Attribution License, which permits
unrestricted use, distribution, and reproduction in any medium, provided the
original author and source are credited.Genetic Selection of Low Fertile Onchocerca volvulus
by Ivermectin TreatmentThe authors received no specific funding for this study.Background
Onchocerciasis (river blindness) is a human disease caused by the filarial worm
Onchocerca volvulus. Adult worms can live for over a decade in
skin nodules of affected humans, releasing millions of microfilariae that cause
debilitating itching and blindness [1]. An estimated 37
million people are infected [2], and there are 46,000 new cases of blindness
annually (http://www.apoc.bf/).
International programs supported by the World Health Organization and many other
groups have worked to control the impact of onchocerciasis using vector control with
insecticides beginning in 1974 and mass drug administration (MDA) with ivermectin
(IVM, brand name Mectizan) beginning in 1987 (Figure 1) [3]. IVM is a highly
effective microfilaricide and inhibits female worm microfilarial production for
several months. Annual IVM MDA reduces morbidity [4,5] and lowers transmission [6,7]. From 1974 to
2002, the Onchocerciasis Control Programme (OCP) in West Africa greatly decreased
O. volvulus transmission in the 11 OCP countries and prevented
600,000 cases of blindness [8–10]. IVM without vector control has been the principal tool for
the Onchocerciasis Elimination Program of the Americas (1992–present)
[9]
and the African Programme for Onchocerciasis Control (1995–present). In
the Americas, where O. volvulus is less common, the Onchocerciasis
Elimination Program has substantially reduced transmission and is on track to
eliminate the disease [9].
10.1371/journal.pntd.0000076.g001
Distribution of Onchocerciasis Showing Current Status of Global
Onchocerciasis Control.
Red shading represents areas receiving ivermectin treatment. Yellow shading
represents areas requiring further epidemiological surveys. Green shading
indicates the area covered by the OCP in West Africa. Pink zones indicate
the special intervention zones, i.e., previous OCP areas receiving
ivermectin and some vector control. Figure from [10].
The African Programme for Onchocerciasis Control has extended treatment to 19
countries beyond those originally included in the OCP through sustainable
community-directed IVM treatment [1,11]. By the end of 2005, 400 million treatments had been supplied
in Africa by Merck’s Mectizan Donation Program, with an estimated 40
million people treated by nearly 300,000 community distributors (http://www.apoc.bf/). Nevertheless, the ecology of the disease in
Africa, including the broad geographic range of O. volvulus and its
blackfly vector, leads to the estimation that IVM treatment of at least
65% of the population for 25 or more years will be necessary to eliminate
infection [9,12]. There are
significant logistical obstacles to achieving such broad-ranging and prolonged
treatment, and there is also concern that O. volvulus resistance to
IVM will emerge. IVM resistance has become widespread in many parasitic nematodes of
livestock [13,14]. At present there
are no alternative drugs for IVM for use in the Onchocerca MDA
programs that reduce microfilariae or kill adult worms, which can live up to 15
years in the human host.
The emergence of drug-resistant O. volvulus has been suggested by
reports of patients failing to respond to IVM treatment [15,16]. A recent report from Ghana has
provided the first proof of IVM resistance in O. volvulus: Mike
Osei-Atweneboana and colleagues showed that the ability of IVM to suppress skin
microfilariae repopulation was reduced in some communities that had received
6–18 years of IVM MDA [17]. The authors
predict that a high rate of repopulation of skin with microfilariae will allow
continued parasite transmission, possibly with IVM-resistant O.
volvulus leading to disease recrudescence. Additionally, studies have
associated IVM resistance with genetic markers [18–25], particularly the
β-tubulin gene in human O. volvulus
and the livestock nematode parasite Haemonchus contortus[22,26]. However, previous
O. volvulus genotyping studies were non-longitudinal, using
worms collected from different IVM-naïve and treated individuals.
A New Study: IVM Causes Genetic Selection on O. volvulus
A new study by Catherine Bourguinat and colleagues published in PLoS
Neglected Tropical Diseases extends these previous reports and
concludes not only that IVM causes genetic selection on O. volvulus
worms, but that this selection is also associated with a lower reproductive rate of
the female parasites [27]. In this study of O. volvulus
treatment in a hyperendemic region of central Cameroon, parasite genotypes
(β-tubulin gene and two controls) and phenotypes
(female fertility) were characterized in worms collected from the same individuals
before and after four or 13 IVM treatments over three years. Parasites were
collected pre- and post-treatment from clinical trial patients in four IVM treatment
groups: 150 µg/kg of body weight annually or three-monthly, and 800
µg/kg annually or three-monthly.
Analyses of the genetic polymorphism in parasites pre- and post- treatment clearly
showed a significant selection for β-tubulin heterozygotes
in female worms. The most marked effect was in the three-monthly treated groups,
where the frequency of the β-tubulin
“aa” homozygotes post-IVM was reduced on average from
68.6% to 25.6%, while the “ab”
heterozygotes increased from 20.9% to 69.2% over three years.
Moreover, β-tubulin “aa” homozygous
females were significantly more fertile than heterozygotes before treatment
(67% versus 37%) and 12 months after the last IVM dose in the
groups treated annually (60% versus 17%). No significant
selection was observed in the control genes.
Strengths and Limitations of the Study
A major strength of this study is that the O. volvulus parasites
were collected from the same individuals before and after IVM treatments. Therefore
the observed changes in genotype frequencies between IVM-naïve and treated
O. volvulus populations are not due to factors such as
geographical or sampling effects.
The main limitation of this study was that some worm samples could not be genotyped,
thus reducing the number that could be analyzed, particularly after treatment. This
limitation might also have impeded the genotyping of DNA, ideally prepared from worm
sections instead of just whole females. What is given as a single genotype is, in
fact, a consensus of multiple genotypes including the adult female body and progeny
(uterine embryos and microfilariae). Furthermore, the samples from unfertile
females, which probably represent true singletons, were treated the same as those
from females classified as being of low or high fertility. Consequently, the study
leaves unanswered questions including whether the selection of the
β-tubulin heterozygote genotypes is on the females or
their progeny. If it is on the progeny, questions remain regarding the fitness and
susceptibility to IVM treatment of the β-tubulin
heterozygote microfilariae once they develop into the infective stage larvae and
enter a new human host.
The authors do not present a hypothesis to explain why IVM causes selection for
β-tubulin heterozygote genotypes. The glutamate-gated
chloride channels are thought to be involved in the mode of action of IVM and
resistance to the drug [28]. Treatment with IVM is known to cause a loss of
polymorphism not only at certain β-tubulin gene loci, but
also at certain loci of the genes encoding the gamma-aminobutyric acid receptor,
glutamate-gated chloride channel, and ATP-binding cassette transporter of
IVM-resistant H. contortus[23]. It
will therefore be important to examine for polymorphisms in these genes in the
uniquely collected O. volvulus female worms described in this study
and in future studies.
Despite these caveats, the study indicates that IVM causes genetic selection on
O. volvulus worms and points to the daunting possibility of the
spread of IVM-resistant parasites in endemic regions that have been treated with
IVM.
Implications of the Study for River Blindness Control
The finding that IVM treatment selected for β-tubulin
heterozygotes and that this selection was dependent on dosage raises important
concerns for the current river blindness control programs. These concerns are
heightened by the fact that this gene has been linked with IVM resistance in another
parasitic nematode [26], and by the recent evidence that IVM resistance
is occurring in O. volvulus[17]. Semiannual or more frequent treatments are ongoing
in some endemic areas and are under consideration in other areas. Such treatment
might increase the selection pressure. Therefore, Bourguinat and
colleagues’ study is a wake-up call for control programs to select their
treatment regimens carefully and to develop plans for detecting IVM resistance and
the associated genetic markers (control programs will require additional funding for
these plans). This study presents a possible structure of study design that will
incorporate the detection and validation of the genetic markers associated with IVM
resistance.
Simultaneously, we need to greatly increase our current level of effort and support
to develop and test a new generation of control tools for onchocerciasis. These
tools should include both vaccines and macrofilaricides (drugs which kill adult
worms) that have new classes of chemistry with novel modes of action. Recent
breakthroughs now make macrofilaricide development more feasible, and accordingly
such development is now a high-priority goal with the World Health
Organization’s Special Programme for Research and Training in Tropical
Diseases and the Bill and Melinda Gates Foundation [29,30]. The development of an
anti-Onchocerca vaccine has been the focus of research
supported by the Edna McConnell Clark Foundation [31]. It may be possible to
link such a vaccine with drug treatments in a program of vaccine-linked chemotherapy
[32,33]. These new
generations of control tools would complement the present control
measure—the establishment of sustainable community-directed treatment with
IVM—and ultimately support the long-term goal of eliminating
onchocerciasis as a public health problem in Sub-Saharan Africa.
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