Evaluation of the therapeutic efficacy of praziquantel against schistosomes in seven countries with ongoing large-scale deworming programs.

The World Health Organization (WHO) recommends periodic assessment of the therapeutic efficacy of praziquantel (PZQ) to detect reduced efficacy that may arise from drug resistance in schistosomes. In this multi-country study (2014), we assessed the therapeutic efficacy of a single oral dose of PZQ (40 mg/kg) against Schistosoma mansoni (Brazil, Cameroon, Ethiopia, Mali, Madagascar and Tanzania), S. haematobium (Cameroon, Ethiopia, Mali, Tanzania and Zanzibar) and S. japonicum (the Philippines) infections in school-aged children, across a total of 12 different trials. Each trial was performed according to the standardized methodology for evaluating PZQ efficacy as described by the WHO. Overall, therapeutic efficacy, measured as the reduction in arithmetic mean of schistosome egg counts following drug administration (egg reduction rate; ERR), was high for all three schistosome species (S. mansoni: 93.4% (95%CI: 88.8-96.8); S. haematobium: 97.7% (95%CI: 96.5-98.7) and S. japonicum: 90.0% (95%CI: 68.4-99.3). At the trial level, therapeutic efficacy was satisfactory (point estimate ERR ≥90%) for all three Schistosoma species with the exception of S. mansoni in Cameroon where the ERR was 88.5% (95%CI: 79.0-95.1). Furthermore, we observed that in some trials individual drug response could vary significantly (wide 95%CI) and that few non-responsive individuals could significantly impact ERR point estimates. In conclusion, these results do not suggest any established reduced efficacy of the standard PZQ treatment to any of the three schistosome species within these countries. Nevertheless, the substantial degree of variation in individual responses to treatment in some countries underpins the need for future monitoring. The reported ERR values serve as reference values to compare with outcomes of future PZQ efficacy studies to ensure early detection of reduced efficacies that could occur as drug pressure continues increase. Finally, this study highlights that 95%CI should be considered in WHO guidelines to classify the therapeutic efficacy of PZQ.

Piloting an integrated approach for estimation of environmental risk of Schistosoma haematobium infections in pre-school-aged children and their mothers at Barombi Kotto, Cameroon.

Within schistosomiasis control, assessing environmental risk of currently non-treated demographic groups e.g. pre-school-aged children (PSAC) and their mothers is important. We conducted a pilot micro-epidemiological assessment at the crater lake of Barombi Kotto, Cameroon with GPS tracking and infection data from 12 PSAC-mother pairs (n = 24) overlaid against environmental sampling inclusive of snail, parasite and water-use information. Several high-risk locations or 'hotspots' with elevated water contact, increased intermediate snail host densities and detectable schistosome environmental DNA (eDNA) were identified. Exposure between PSAC and mother pairs was temporally and spatially associated, suggesting interventions which can benefit both groups simultaneously might be feasible. When attempting to interrupt parasite transmission in future, overlaid maps of snail, parasite and water contact data can guide fine-scale spatial targeting of environmental interventions.

Comparison of two diagnostic techniques to determine the prevalence of Schistosoma mansoni infections in Cameroonian school children

Background: The Kato-Katz technique is recommended for diagnosis of Schistosoma mansoni infection by the World Health Organization. However, egg counts are subject to variability. The aim of this study is to determine the prevalence of S. mansoni infection in school children using two different techniques and to recommend the technique that should be routinely used in the diagnosis of this infection. Methodology: Field investigations on faecal samples from 299 Cameroonian school children were carried out in 2016 to compare the effectiveness of the Kato-Katz and Formalin-ether techniques in diagnosis of S. mansoni infections. Results: Schistosome eggs were detected in 37 (12.3%) samples with the Kato-Katz technique and 61 (20.4%) samples with the Formalin-ether technique. The difference between the prevalence observed for the two techniques was significant in males and age group 10 - 12 years (p < 0.5). Conclusion: The Formalin-ether technique was more sensitive than the Kato-Katz method for detecting S. mansoni eggs in faecal matter. Despite its cost, the Formalin-ether technique can be routinely used in the laboratory for epidemiological studies of intestinal schistosomiasis

Prospects for the elimination of schistosomiasis and soil-transmitted helminthiasis: exploring disease trends through time at the Barombi crater lakes, South-West Cameroon.

In Cameroon, there is a national programme engaged in the control of schistosomiasis and soil-transmitted helminthiasis. In certain locations, the programme is transitioning from morbidity control towards local interruption of parasite transmission. The volcanic crater lake villages of Barombi Mbo and Barombi Kotto are well-known transmission foci and are excellent context-specific locations to assess appropriate disease control interventions. Most recently they have served as exemplars of expanded access to deworming medications and increased environmental surveillance. In this paper, we review infection dynamics through time, beginning with data from 1953, and comment on the short- and long-term success of disease control. We show how intensification of local control is needed to push towards elimination and that further environmental surveillance, with targeted snail control, is needed to consolidate gains in preventive chemotherapy as well as empower local communities to take ownership of interventions.

Control of soil-transmitted helminths in sub-Saharan Africa: diagnosis, drug efficacy concerns and challenges.

Soil-transmitted helminthiasis (STH) occur throughout the developing world and remain a major public health problem in the poorest communities with enormous consequences on health and development of school-aged and preschool children. The extent of the problem has long been neglected because these diseases rarely kill at a young age, due to their insidious and chronic nature. Today there exists a momentum and an unprecedented opportunity for a cost-effective control of these neglected tropical diseases. The control of these diseases has become a priority on the agenda of many governments, donors and international agencies, which provides important funding opportunities for STH control through integration and co-implementation of control interventions. In the era of preventive chemotherapy, the intensification of mass administration of anthelminthic drugs will result in epidemiological changes of STH transmission levels, with a rapid increase of light infections as far as control programmes are successful. As consequence, the current diagnostic tools may not be always appropriate and may therefore require adaptation or development of alternative assays adapted to the stage of control. This paper reviews aspects relevant to diagnosis and appropriate tools for accurate surveillance and monitoring of STH control programmes and drug efficacy. Prospects, opportunities and challenges for the control are discussed.

Praziquantel: its use in control of schistosomiasis in sub-Saharan Africa and current research needs.

Treatment with praziquantel (PZQ) has become virtually the sole basis of schistosomiasis control in sub-Saharan Africa and elsewhere, and the drug is reviewed here in the context of the increasing rate that it is being used for this purpose. Attention is drawn to our relative lack of knowledge about the mechanisms of action of PZQ at the molecular level, the need for more work to be done on schistosome isolates that have been collected recently from endemic areas rather than those maintained in laboratory conditions for long periods, and our reliance for experimental work mainly on Schistosoma mansoni, little work having been done on S. haematobium. There is no evidence that resistance to PZQ has been induced in African schistosomes as a result of its large-scale use on that continent to date, but there is also no assurance that PZQ and/or schistosomes are in any way unique and that resistant organisms will not be selected as a result of widespread drug usage. The failure of PZQ to produce complete cures in populations given a routine treatment should therefore solicit considerable concern. With few alternatives to PZQ currently available and/or on the horizon, methods to monitor drug-susceptibility in African schistosomes need to be devised and used to help ensure that this drug remains effective for as long a time as possible.

Evaluation of the in vivo activity of different concentrations of Clerodendrum umbellatum Poir against Schistosoma mansoni infection in mice.

Clerodendrum umbellatum Poir (Verbenaceae) is traditionally used in Cameroon for the treatment of many diseases including intestinal helminthiasis. This study was undertaken to assess the in vivo antischistosomal activity of its leaves aqueous extract on a Schistosoma mansoni mice model and to determine the most effective dose of this extract. Mice showing a patent infection of S. mansoni were daily treated with C. umbellatum leaves aqueous extract at the doses of 40, 80 or 160 mg/kg body weight for 14 days. Seven days after administration of the extract, schistosomicidal activity was evaluated on the liver and spleen weights, faecal eggs releasing, liver egg count and worm burden. Treatment using C. umbellatum leaves aqueous extract resulted in an important reduction in faecal egg output by 75.49% and 85.14% for 80 mg/kg and 160 mg/kg of the extract respectively. These reduction rates did not differ significantly from the 100% obtained in the group of infected mice treated with 100 mg/kg of praziquantel. C. umbellatum leaves aqueous extract was lethal to S. mansoni worm. A 100% reduction rate was recorded in the group of infected mice treated with 160 mg/kg of the extract, as well as in praziquantel-treated mice. An amelioration of the hepatosplenomegaly was noticed in both the extract-treated mice and the praziquantel-treated mice. From these results, we can conclude that C. umbellatum leaves aqueous extract demonstrated schistosomicidal properties in S. mansoni model at doses of at least 80 mg/kg body weight.

A single-strand conformation polymorphism (SSCP) approach for investigating genetic interactions of Schistosoma haematobium and Schistosoma guineensis in Loum, Cameroon.

Single-strand conformation polymorphism (SSCP) analysis of the second internal transcribed spacer (ITS2) of nuclear ribosomal DNA provides a molecular tool for the identification of Schistosoma haematobium, Schistosoma guineensis and the hybrids of these two species. This molecular tool was utilized to provide a detailed analysis of the interactions between S. haematobium and S. guineensis in hybrid zones of Loum, Littoral Province, Cameroon. Individual hybrid schistosomes were identified within the natural populations collected from Loum in 1990, 1999 and 2000, which would have been misidentified as S. haematobium using solely morphological and sequence criteria. This study indicates the complexities of the hybridization between S. haematobium and S. guineensis and emphasizes the importance of assessing morphological, biological and molecular data to gain insights into the interaction of these two species over time.

Towards control of schistosomiasis in sub-Saharan Africa.

Approximately 80% of the 200 million people infected with schistosomiasis inhabit sub-Saharan Africa, and the annual mortality is estimated to be 280,000. Praziquantel is the drug of choice in the treatment of schistosomiasis and pregnant women may now be treated. It was agreed at the World Health Assembly in 2001 that at least 75% of school-aged children in high burden areas should be treated for schistosomiasis and soil-transmitted helminth infections by 2010 to reduce morbidity. A grant from the Bill and Melinda Gates Foundation to the Schistosomiasis Control Initiative, Imperial College of Science, Technology and Medicine, London has enabled control programmes to be initiated in Uganda, Tanzania, Zambia, Burkina Faso, Niger and Mali. Additional programmes have recently commenced in Zanzibar with a grant from the Health Foundation to The Natural History Museum, London and in Cameroon. Combination treatment for schistosomiasis, gastrointestinal helminths and filariasis reduces costs of control programmes. The EC Concerted Action Group on 'Praziquantel: its central role in the chemotherapy of schistosome infection' met in Yaoundé Cameroon in 2004 to discuss recent developments in laboratory and field studies. The use of standard operating procedures will enable data on drug action on schistosomes produced in different laboratories to be compared. With the ever increasing use of praziquantel there is a possibility of the development of resistance by schistosomes to the drug, hence the necessity to explore the activities of other compounds. Artemether, unlike praziquantel, is effective against immature schistosomes. The effectiveness of mirazid, an extract of myrrh, is controversial as data from different laboratories are equivocal. It is suggested that an independent body such as the World Health Organization should determine whether mirazid should be used in the treatment of schistosomiasis.

The interaction of Schistosoma haematobium and S. guineensis in Cameroon.

Interactions between schistosomes are complex with some different species being able to mate and hybridize. The epidemiology of schistosomiasis in specific areas of South West Cameroon has evolved remarkably over 30 years as a result of hybridization between Schistosoma guineensis and S. haematobium. Morphological and biological data suggest that S. haematobium replaced S. guineensis in areas of Cameroon through introgressive hybridization. Data are reported on the use of single stranded conformational polymorphism (SSCP) analysis of the nuclear ribosomal second internal transcribed spacer (ITS2) of individual schistosomes from hybrid zones of Cameroon. The data show that since 1990 S. haematobium has completely replaced S. guineensis in Loum, with S. haematobium and the recombinants still present in 2000. This study illustrates the complexities of the dynamics between S. haematobium and S. guineensis in South West Cameroon.

A neotropical snail host of Schistosoma mansoni introduced into Africa and consequences for the schistosomiasis transmission Biomphalaria tenagophila in Kinshasa (Democratic Republic of Congo).

Malacological surveys carried out in the early 1970s in water bodies of the Kinshasa area, Lower Zaire (Democratic Republic of Congo), showed the appearance of a Biomphalaria species which was identified as Biomphalaria camerunensis. In 1976, other surveys confirmed the presence of the species in several sites and showed numerous infected snails with Schistosoma mansoni, demonstrating for the first time an active transmission of the parasite responsible of the intestinal schistosomiasis in this area. The most recent malacological sampling was carried out by one of us in 1994 in Mangungu River and revealed the presence of apparently the same snail species. However, conchological, anatomical and molecular studies showed that this snail may be considered as an introduced neotropical species, B. tenagophila. To our knowledge, this is the second example of the introduction of a neotropical snail host of schistosomes into Africa.

[Urban schistosomiasis in Cameroon: a longitudinal study of its transmission in a new site of an extension of the intestinal schistosomiasis focus in Mélen, Yaoundé]. / Schistosomose urbaine au Cameroun: étude longitudinale de la transmission dans un nouveau site d'extension du foyer de schistosomose intestinale de Mélen, Yaoundé.

In order to to set up the present situation on schistosomiasis in the neighbourhoods around the University of Yaounde I campus, a malacological survey (collection of freshwater snails, their identification and tests on cercarial emergence) was carried out monthly over 12 months in 2 water sources, followed by a parasitological diagnosis of intestinal schistosomiasis in subjects who acknowledge having come into contact with the infested water course. The malacological survey revealed 4 freshwater gastropod species, two (Bulinus globosus and Biomphalaria pfeifferi) of which are intermediate hosts of schistosomes. Biomphalaria pfeifferi specimens from the former quarry pond of Ngoa-Ekellé neighbourhood showed an average infestation rate of 9.7%, and emitted schistosome cercariae throughout the whole period of study The parasitological diagnosis included 112 (81.2%) out of the 138 individuals identified, including 55 males and 57 females; 27 out of the 112 subjects had S. mansoni eggs in stools, making a prevalence rate of 24.1%. These patients were composed of 21 males and 6 females. The prevalence of S. mansoni is therefore significantly higher in males. The average parasitic load for our sample was 248 eggs/g of stools. It was much higher (384 eggs/g stools) for males (280 eggs/g stools) than for females (136 eggs/g stools). The differences of prevalences and parasitic loads between sex can be explained in part by the varying activities of interest according to the different groups in the infested watering places.

Interactions between Schistosoma haematobium and Schistosoma mansoni in humans in north Cameroon.

OBJECTIVES: To analyse the relationships between the frequency of ectopic localizations of Schistosoma haematobium and S. mansoni eggs. METHODS: Studies were conducted in 11 villages in north Cameroon, around Bessoum, a village where an epidemic of bloody diarrhoea caused by S. mansoni occurred in 1997. RESULTS: The results revealed infection prevalence rates of 70.5% for S. haematobium and 30.8% for S. mansoni. Interestingly, S. mansoni eggs were found in 14.5% of the urine samples and S. haematobium eggs in 3% of the stool samples. These ectopic eliminations of schistosome eggs resulted from sexual interactions between the two species of schistosomes, and from a spill-over of high infection loads. The clinical study showed that the morbidity was lower in individuals with mixed infections and high loads of S. haematobium than in those with S. mansoni infections only, suggesting a possible lowering effect of S. haematobium infection on S. mansoni morbidity. DISCUSSION: The results obtained in human populations are discussed in relation to the known schistosome interspecific interactions in animal models.

Polyparasitism with Schistosoma haematobium and soil-transmitted helminth infections among school children in Loum, Cameroon.

Prevalence and abundance of Schistosoma haematobium and soil-transmitted helminths (STH) were assessed among a total of 1600 pupils (urine, n = 1190; faecal samples, n = 1454) attending five schools in Loum, Littoral Province, Cameroon, with the specific aim of assessing the extent of polyparasitism and the extent to which infections were focused in particular subsets of the study group. Prevalence of S. haematobium was 62.8% with an abundance (arithmetic mean of egg counts) of 54 eggs/10 ml urine. For the STH these were 47.7% and 619 eggs per gram of faeces (EPG) for Trichuris trichiura, 65.5% and 3636 EPG for Ascaris lumbricoides, and 1.4% and <0.1 EPG for hookworms. Most children (90.3%) were infected with at least one of these four species, the largest proportion (34.3%) carrying two species; 27.4% carried three and 1.1% carried concurrently all four species of parasites. The average number of species harboured increased with age, as did the prevalences of S. haematobium and T. trichiura but not that of A. lumbricoides. All STH showed marked differences in prevalence between the five schools but only T. trichiura varied significantly between sexes. Mean abundance of infection varied significantly between age classes, among schools and between the sexes, with females showing heavier mean EPGs for A. lumbricoides and T. trichiura and males higher mean eggs/10 ml urine for S. haematobium infections. A highly significant association was detected between A. lumbricoides and T. trichiura, that was not context-dependent. This was confirmed in quantitative analyses after controlling for differences in abundance between schools, sexes and age classes. A weaker context-dependent association (prevalence data) was detected between S. haematobium and A. lumbricoides (sex- and age-dependent) but quantitative associations between these two species, as well as between S. haematobium and T. trichiura, were not convincing.

A phylogeny based on three mitochondrial genes supports the division of Schistosoma intercalatum into two separate species.

Two recognized strains of Schistosoma intercalatum, one from the Democratic Republic of Congo (DRC), formerly Zaire, and the other from Cameroon, have been investigated using DNA sequences from 3 mitochondrial genes, cytochrome oxidase subunit 1 (cox1), NADH dehydrogenase subunit 6 (nad6) and the small ribosomal RNA gene (rrnS). In addition, partial DNA sequences from the nuclear large subunit ribosomal RNA gene (lsrDNA) were included within the study. Although partial lsrDNA alone reveals little taxonomic information, phylogenetic analysis of the mitochondrial data demonstrates a clear dichotomy between the 2 purported strains and it is proposed that they should be treated as distinct taxa. The 'original' S. intercalatum now falls relatively basal in the S. haematobium group, while the proposed new species is more derived and sister taxon to S. bovis and S. curassoni.

Isoenzyme analysis of Schistosoma haematobium, S. intercalatum and their hybrids and occurrences of natural hybridization in Cameroon.

Isoelectric focusing of glucose-6-phosphate dehydrogenase (G6PD) produced clearly identifiable profiles for S. haematobium and S. intercalatum and their hybrids. To provide a more detailed analysis of the interactions of S. haematobium and S. intercalatum in South West Cameroon over the last 12 years, G6PD analyses were carried out on individual schistosomes collected in Kumba in 1990, Loum in 1990, 1999 and 2000 and Barombi Mbo and Barombi Kotto in 1999. Studies were also carried out on the two parental species S. haematobium Barombi Mbo, S. intercalatum Edea and subsequent generations of hybrids resulting from laboratory crosses of the two parental species. The isoenzyme analysis demonstrated that the 1990 isolate from Kumba, was a recombinant of S. intercalatum x S. haematobium, and that 30% of individual schistosomes collected in 1990 in Loum were also recombinants. The remainder gave data indicative of S. haematobium. In 1999, 12.5% of individuals from Loum showed recombination and 10% in 2000. Results from the most recent parasitological survey in October 2000 showed the persistence of the recombinant population in addition to that of S. haematobium. There was also evidence of recombination having taken place in Barombi Kotto but not Barombi Mbo. This study demonstrates how the situation has changed over the last 12 years, and emphasizes the importance of assessing morphological, biological and molecular data together to gain a true picture of the rapidly evolving situation.

Experimental evidence of hybrid breakdown between the two geographical strains of Schistosoma intercalatum.

Reciprocal crosses (Schistosoma intercalatum male Zaire x S. intercalatum female Cameroon: S. intercalatum female Zaire x S. intercalatum male Cameroon) were produced in 10 mice by exposing each mouse to 60 male cercariae of one isolate and 60 female cercariae of the other isolate, and vice versa. Hybrid generations originating from the two crosses were established. The infectivity of the F1, F2, F3 and F4 hybrid generations were evaluated after exposing snails individually to 5 miracidia. A comparative histological study of snails infected with F2 and F4 hybrid sporocysts from both crosses was made to assess abnormalities in the intramolluscan development of the hybrids. The worm recovery rate and fecundity were measured by comparing the fitness of the mid-parents with that of the hybrids. S. intercalatum Cameroon was compatible with Bulinus forskalii and incompatible with B. globosus whereas S. intercalatum Zaire was compatible with B. globosus and incompatible with B. forskalii. In the case of S. intercalatum male Cameroon x S. intercalatum female Zaire, hybrid miracidia develops in both B: forskalii and B. globosus in F1, F2 and F3 generations. The infection rate was much lower for B. globosus and F2 and F3 generations produced few cercariae (less than 30 cercariae in overall per snail) and F4 miracidia were only infective to B. forskalii again producing few cercariae. At 40 days post-infection the sporocyst contains masses of acidophilic granules originating from the breakup of pycnotic nuclei. Similarly the F1, F2 and F3 miracidia of the reciprocal cross (S. intercalatum male Zaire x S. intercalatum female Cameroon) exhibited a dual infectivity for B. forskalii and B. globosus, but cercarial productivity was low (less than 30 cercariae overall per snail for F2 and F3 generations). Histological studies demonstrated sporocyst degeneration in snails infected with F4 generation. In the definitive host, the F, generation (both crosses) exhibited hybrid vigour in that the worm return was greater than that of the mid-parent, F2 and F3 generations. No significant difference in fecundity was demonstrated between the parental strains and the F1 and F2 generations, yet egg production of the F3 generation was significantly lower. It is apparent that there is a post-zygotic barrier in the crosses of S. intercalatum Zaire and S. intercalatum Cameroon from the F2 generations onwards, and sterility of the F4 hybrid sporocyst supports the concept of the existence of 2 distinct taxa.

Lack of prezygotic isolation by assortative mating between the two cryptic species of the polytypic Schistosoma intercalatum taxon.

Ten mice were individually exposed to 30 male and 30 female cercariae of Schistosoma intercalatum from Cameroon and to 30 male and 30 female cercariae of S. intercalatum from Zaire. Ten weeks later, the nine surviving mice were killed, and a total of 263 adult schistosomes (228 pairs, 7 unpaired males, 28 unpaired females) were perfused. All worms were identified utilising the random amplified polymorphic DNA technique using primer OPG06 which enables the differentiation of the Cameroon and Zaire genotypes. The data demonstrate that there were no significant differences between the number of homospecific pairs observed compared with the expected number under the null hypothesis. Matings occurred in a random manner indicating that mate choice did not exist for the two genotypes of S. intercalatum. Hence there is no prezygotic isolation mechanism which would prevent the interaction of the two genotypes if they were sympatric. However, as they are allopatric, the possibility of the production of unfit hybrids does not arise. Consequently, the reinforcement of a prezygotic isolating mechanism does not play a significant role in the speciation process.

Are poor responses to praziquantel for the treatment of Schistosoma mansoni infections in Senegal due to resistance? An overview of the evidence.

This paper summarizes and concludes in-depth field investigations on suspected resistance of Schistosoma mansoni to praziquantel in northern Senegal. Praziquantel at 40 mg/kg usually cures 70-90% of S. mansoni infections. In an initial trial in an epidemic S. mansoni focus in northern Senegal, only 18% of the cases became parasitologically negative 12 weeks after treatment, although the reduction in mean egg counts was within normal ranges (86%). Among other hypotheses to explain the observed low cure rate in this focus, the possibility of drug resistance or tolerance had to be considered. Subsequent field trials with a shorter follow-up period (6-8 weeks) yielded cure rates of 31-36%. Increasing the dose to 2 x 30 mg/kg did not significantly improve cure rates, whereas treatment with oxamniquine at 20 mg/kg resulted in a normal cure rate of 79%. The efficacy of praziquantel in this focus could be related to age and pre-treatment intensity but not to other host factors, including immune profiles and water contact patterns. Treatment with praziquantel of individuals from the area residing temporarily in an urban region with no transmission, and re-treatment after 3 weeks of non-cured individuals within the area resulted in normal cure rates (78-88%). The application of an epidemiological model taking into account the relation between egg counts and actual worm numbers indicated that the low cure rates in this Senegalese focus could be explained by assuming a 90% worm reduction after treatment with praziquantel; in average endemic situations, such a drug efficacy would result in normal cure rates. Laboratory studies by others on the presence or absence of praziquantel resistance in Senegalese schistosome strains have so far been inconclusive. We conclude that there is no convincing evidence for praziquantel-resistant S. mansoni in Senegal, and that the low cure rates can be attributed to high initial worm loads and intense transmission in this area.