Genetic Profiling of Plasmodium ovale wallikeri Relapses With Microsatellite Markers and Whole-Genome Sequencing.

Like Plasmodium vivax, both Plasmodium ovale curtisi and Plasmodium ovale wallikeri have the ability to cause relapse in humans, defined as recurring asexual parasitemia originating from liver-dormant forms subsequent to a primary infection. Here, we investigated relapse patterns in P ovale wallikeri infections from a cohort of travelers who were exposed to the parasite in sub-Saharan Africa and then experienced relapses after their return to France. Using a novel set of 8 highly polymorphic microsatellite markers, we genotyped 15 P ovale wallikeri relapses. For most relapses, the paired primary and relapse infections were highly genetically related (with 12 being homologous), an observation that was confirmed by whole-genome sequencing for the 4 relapses we further studied. This is, to our knowledge, the first genetic evidence of relapses in P ovale spp.

The primate malaria parasites Plasmodium malariae, Plasmodium brasilianum and Plasmodium ovale spp.: genomic insights into distribution, dispersal and host transitions.

During the twentieth century, there was an explosion in understanding of the malaria parasites infecting humans and wild primates. This was built on three main data sources: from detailed descriptive morphology, from observational histories of induced infections in captive primates, syphilis patients, prison inmates and volunteers, and from clinical and epidemiological studies in the field. All three were wholly dependent on parasitological information from blood-film microscopy, and The Primate Malarias" by Coatney and colleagues (1971) provides an overview of this knowledge available at that time. Here, 50 years on, a perspective from the third decade of the twenty-first century is presented on two pairs of primate malaria parasite species. Included is a near-exhaustive summary of the recent and current geographical distribution for each of these four species, and of the underlying molecular and genomic evidence for each. The important role of host transitions in the radiation of Plasmodium spp. is discussed, as are any implications for the desired elimination of all malaria species in human populations. Two important questions are posed, requiring further work on these often ignored taxa. Is Plasmodium brasilianum, circulating among wild simian hosts in the Americas, a distinct species from Plasmodium malariae? Can new insights into the genomic differences between Plasmodium ovale curtisi and Plasmodium ovale wallikeri be linked to any important differences in parasite morphology, cell biology or clinical and epidemiological features?

Plasmodium ovale wallikeri and P. ovale curtisi Infections and Diagnostic Approaches to Imported Malaria, France, 2013-2018.

We retrospectively analyzed epidemiologic, clinical, and biologic characteristics of 368 Plasmodium ovale wallikeri and 309 P. ovale curtisi infections treated in France during January 2013­December 2018. P. ovale wallikeri infections displayed deeper thrombocytopenia and shorter latency periods. Despite similar clinical manifestations, P. ovale wallikeri­infected patients were more frequently treated with artemisinin-based combination therapy. Although the difference was not statistically significant, P. ovale wallikeri­infected patients were 5 times more frequently hospitalized in intensive care or intermediate care and had a higher proportion of severe thrombocytopenia than P. ovale curtisi­infected patients. Rapid diagnostic tests that detect aldolase were more efficient than those detecting Plasmodium lactate dehydrogenase. Sequence analysis of the potra gene from 90 P. ovale isolates reveals an insufficient polymorphism for relapse typing.

Epidemiological profile of Plasmodium ovale spp. imported from Africa to Anhui Province, China, 2012-2019.

BACKGROUND: Although autochthonous malaria cases are no longer reported in Anhui Province, China, imported malaria has become a major health concern. The proportion of reported malaria cases caused by Plasmodium ovale spp. increased to levels higher than expected during 2012 to 2019, and showed two peaks, 19.69% in 2015 and 19.35% in 2018. METHODS: A case-based retrospective study was performed using data collected from the China Information System for Disease Control and Prevention (CISDCP) and Information System for Parasitic Disease Control and Prevention (ISPDCP) from 2012 to 2019 to assess the trends and differences between Plasmodium ovale curtisi (P. o. curtisi) and Plasmodium ovale wallikeri (P. o. wallikeri). Epidemiological characteristics were analyzed using descriptive statistics. RESULTS: Plasmodium o. curtisi and P. o. wallikeri were found to simultaneously circulate in 14 African countries. Among 128 patients infected with P. ovale spp., the proportion of co-infection cases was 10.16%. Six cases of co-infection with P. ovale spp. and P. falciparum were noted, each presenting with two clinical attacks (the first attack was due to P. falciparum and the second was due to P. ovale spp.) at different intervals. Accurate identification of the infecting species was achieved among only 20.00% of cases of P. ovale spp. infection. At the reporting units, 32.17% and 6.96% of cases of P. ovale spp. infection were misdiagnosed as P. vivax and P. falciparum infections, respectively. CONCLUSION: The present results indicate that the potential of P. ovale spp. to co-infect with other Plasmodium species has been previously underestimated, as is the incidence of P. ovale spp. in countries where malaria is endemic. P. o. curtisi may have a long latency period of > 3 years and potentially cause residual foci, thus posing challenges to the elimination of malaria in P. ovale spp.-endemic areas. Considering the low rate of species identification, more sensitive point-of-care detection methods need to be developed for P. ovale spp. and introduced in non-endemic areas.

A powerful qPCR-high resolution melting assay with taqman probe in plasmodium species differentiation.

BACKGROUND: The use of highly sensitive molecular tools in malaria diagnosis is currently largely restricted to research and epidemiological settings, but will ultimately be essential during elimination and potentially eradication. Accurate diagnosis and differentiation down to species levels, including the two Plasmodium ovale species and zoonotic variants of the disease, will be important for the understanding of changing epidemiological patterns of the disease. METHODS: A qPCR-high resolution melting (HRM) method was to detect and differentiate all human Plasmodium species with one forward and one reverse primer set. The HRM detection method was further refined using a hydrolysis probe to specifically discriminate Plasmodium falciparum. RESULTS: Out of the 113 samples tested with the developed HRM-qPCR- P. falciparum probe assay, 96 (85.0 %) single infections, 12 (10.6 %) mixed infections, and 5 (4.4 %) were Plasmodium negative. The results were concordant with those of the nested PCR at 98.2 %. The assay limit of detection was varied from 21.47 to 46.43 copies /µl, equivalent to 1-2.11 parasites/µl. All P. falciparum infections were confirmed with the associated Taqman probe. CONCLUSIONS: Although the dependence on qPCR currently limits its deployment in resource-limited environments, this assay is highly sensitive and specific, easy to perform and convenient for Plasmodium mono-infection and may provide a novel tool for rapid and accurate malaria diagnosis also in epidemiological studies.

High prevalence of asymptomatic malaria infections in adults, Ashanti Region, Ghana, 2018.

BACKGROUND: Ghana is among the high-burden countries for malaria infections and recently reported a notable increase in malaria cases. While asymptomatic parasitaemia is increasingly recognized as a hurdle for malaria elimination, studies on asymptomatic malaria are scarce, and usually focus on children and on non-falciparum species. The present study aims to assess the prevalence of asymptomatic Plasmodium falciparum and non-falciparum infections in Ghanaian adults in the Ashanti region during the high transmission season. METHODS: Asymptomatic adult residents from five villages in the Ashanti Region, Ghana, were screened for Plasmodium species by rapid diagnostic test (RDT) and polymerase chain reaction (PCR) during the rainy season. Samples tested positive were subtyped using species-specific real-time PCR. For all Plasmodium ovale infections additional sub-species identification was performed. RESULTS: Molecular prevalence of asymptomatic Plasmodium infection was 284/391 (73%); only 126 (32%) infections were detected by RDT. While 266 (68%) participants were infected with Plasmodium falciparum, 33 (8%) were infected with Plasmodium malariae and 34 (9%) with P. ovale. The sub-species P. ovale curtisi and P. ovale wallikeri were identified to similar proportions. Non-falciparum infections usually presented as mixed infections with P. falciparum. CONCLUSIONS: Most adult residents in the Ghanaian forest zone are asymptomatic Plasmodium carriers. The high Plasmodium prevalence not detected by RDT in adults highlights that malaria eradication efforts must target all members of the population. Beneath Plasmodium falciparum, screening and treatment must also include infections with P. malariae, P. o. curtisi and P. o. wallikeri.

Prospective Clinical and Molecular Evaluation of Potential Plasmodium ovale curtisi and wallikeri Relapses in a High-transmission Setting.

BACKGROUND: Plasmodium ovale curtisi and wallikeri are perceived as relapsing malarial parasites. Contrary to Plasmodium vivax, direct evidence for this hypothesis is scarce. The aim of this prospective study was to characterize the reappearance patterns of ovale parasites. METHODS: P. ovale spp. infected patients were treated with artemether-lumefantrine and followed biweekly for up to 1 year for the detection of reappearing parasitemia. Molecular analysis of reappearing isolates was performed to identify homologous isolates by genotyping and to define cases of relapse following predefined criteria. RESULTS: At inclusion, 26 participants were positive for P. ovale curtisi and/or P. ovale wallikeri. The median duration of follow-up was 35 weeks. Reappearance of the same P. ovale species was observed in 46% of participants; 61% of P. ovale curtisi and 19% of P. ovale wallikeri infection-free intervals were estimated to end with reappearance by week 32. Based on the predefined criteria, 23% of participants were identified with 1 or 2 relapses, all induced by P. ovale curtisi. CONCLUSION: These findings are in line with the currently accepted relapse theory inasmuch as the reappearance of P. ovale curtisi strains following initial blood clearance was conclusively demonstrated. Interestingly, no relapse of P. ovale wallikeri was observed.

A comparison of two PCR protocols for the differentiation of Plasmodium ovale species and implications for clinical management in travellers returning to Germany: a 10-year cross-sectional study.

BACKGROUND: To assess the occurrence of Plasmodium ovale wallikeri and Plasmodium ovale curtisi species in travellers returning to Germany, two real-time PCR protocols for the detection and differentiation of the two P. ovale species were compared. Results of parasite differentiation were correlated with patient data. METHODS: Residual nucleic acid extractions from EDTA blood samples of patients with P. ovale spp. malaria, collected between 2010 and 2019 at the National Reference Centre for Tropical Pathogens in Germany, were subjected to further parasite discrimination in a retrospective assessment. All samples had been analysed by microscopy and by P. ovale spp.-specific real-time PCR without discrimination on species level. Two different real-time PCR protocols for species discrimination of P. o. curtisi and P. o. wallikeri were carried out. Results were correlated with patient data on gender, age, travel destination, thrombocyte count, and duration of parasite latency. RESULTS: Samples from 77 P. ovale spp. malaria patients were assessed, with a male:female ratio of about 2:1 and a median age of 30 years. Parasitaemia was low, ranging from few visible parasites up to 1% infected erythrocytes. Discriminative real-time PCRs revealed 41 cases of P. o. curtisi and 36 cases of P. o. wallikeri infections. Concordance of results by the two PCR approaches was 100%. Assessment of travel destinations confirmed co-existence of P. o. curtisi and P. o. wallikeri over a wide range of countries in sub-Saharan Africa. Latency periods for the two P. ovale species were similar, with median values of 56.0 days for P. o. curtisi and 58.0 days for P. o. wallikeri; likewise, there was no statistically significant difference in thrombocyte count with median values of 138.5/µL for patients with P. o. curtisi and 152.0/µL for P. o. wallikeri-infected patients. CONCLUSIONS: Two different real-time PCR protocols were found to be suitable for the discrimination of P. o. curtisi and P. o. wallikeri with only minor differences in sensitivity. Due to the overall low parasitaemia and the lack of differences in severity-related aspects like parasite latency periods or thrombocyte counts, this study supports the use of P. ovale spp. PCR without discrimination on species level to confirm the diagnosis and to inform clinical management of malaria in these patients.

Plasmodium ovale wallikeri in Western Lowland Gorillas and Humans, Central African Republic.

Human malaria parasites have rarely been reported from free-ranging great apes. Our study confirms the presence of the human malaria parasite Plasmodium ovale wallikeri in western lowland gorillas and humans in Dzanga Sangha Protected Areas, Central African Republic, and discusses implications for malaria epidemiology.

Molecular evidence for relapse of an imported Plasmodium ovale wallikeri infection.

BACKGROUND: Malaria caused by Plasmodium ovale spp. has been neglected by and large from research and has received only little scientific attention during the past decades. Ovale malaria is considered to feature relapses by liver hypnozoites although scientific evidence for this paradigm is scarce. CASE PRESENTATION: Here, the case of a 16-year-old male, who presented with fevers to the outpatient department in Vienna, Austria, after travelling to Uganda and Papua New Guinea is described. Infection with Plasmodium malariae was diagnosed by microscopy and the patient was treated accordingly with a full course of supervised artemether-lumefantrine. He was discharged in good clinical condition with a negative blood smear. One month after initial diagnosis, he returned complaining of fever. Thick blood smear was positive again for malaria parasites, which were confirmed as P. ovale wallikeri by PCR. Retrospective analysis revealed the identical Plasmodium spp. in the initial blood samples. Molecular analysis of various gene loci (nuclear porbp2, 18S rRNA and potra genes) gave identical results providing further evidence for relapse by an identical parasite genotype. Consecutively, the patient was retreated with artemether-lumefantrine and received a regimen of primaquine according to WHO guidelines. CONCLUSION: Conclusive evidence for relapses with P. ovale spp. is rare. The presented case provides convincing confirmation for the relapse paradigm based on re-appearing parasitaemia following supervised treatment in a non-endemic region with a parasite strain of identical genotype.

Prospective comparative multi-centre study on imported Plasmodium ovale wallikeri and Plasmodium ovale curtisi infections.

BACKGROUND: Few previous retrospective studies suggest that Plasmodium ovale wallikeri seems to have a longer latency period and produces deeper thrombocytopaenia than Plasmodium ovale curtisi. Prospective studies were warranted to better assess interspecies differences. METHODS: Patients with imported P. ovale spp. infection diagnosed by thick or thin film, rapid diagnostic test (RDT) or polymerase chain reaction (PCR) were recruited between March 2014 and May 2017. All were confirmed by DNA isolation and classified as P. o. curtisi or P. o. wallikeri using partial sequencing of the ssrRNA gene. Epidemiological, analytical and clinical differences were analysed by statistical methods. RESULTS: A total of 79 samples (35 P. o. curtisi and 44 P. o. wallikeri) were correctly genotyped. Males predominate in wallikeri group (72.7%), whereas were 48.6% in curtisi group. Conversely, 74.3% of curtisi group were from patients of African ethnicity, whilst 52.3% of Caucasians were infected by P. o. wallikeri. After performing a multivariate analysis, more thrombocytopaenic patients (p = 0.022), a lower number of platelets (p = 0.015), a higher INR value (p = 0.041), and shorter latency in Caucasians (p = 0.034) were significantly seen in P. o. wallikeri. RDT sensitivity was 26.1% in P. o. curtisi and 42.4% in P. o. wallikeri. Nearly 20% of both species were diagnosed only by PCR. Total bilirubin over 3 mg/dL was found in three wallikeri cases. Two patients with curtisi infection had haemoglobin under 7 g/dL, one of them also with icterus. A wallikeri patient suffered from haemophagocytosis. Chemoprophylaxis failed in 14.8% and 35% of curtisi and wallikeri patients, respectively. All treated patients with various anti-malarials which included artesunate recovered. Diabetes mellitus was described in 5 patients (6.32%), 4 patients of wallikeri group and 1 curtisi. CONCLUSIONS: Imported P. o. wallikeri infection may be more frequent in males and Caucasians. Malaria caused by P. o. wallikeri produces more thrombocytopaenia, a higher INR and shorter latency in Caucasians and suggests a more pathogenic species. Severe cases can be seen in both species. Chemoprophylaxis seems less effective in P. ovale spp. infection than in P. falciparum, but any anti-malarial drug is effective as initial treatment. Diabetes mellitus could be a risk factor for P. ovale spp. infection.

Polymorphisms analysis of the Plasmodium ovale tryptophan-rich antigen gene (potra) from imported malaria cases in Henan Province.

BACKGROUND: Plasmodium ovale has two different subspecies: P. ovale curtisi and P. ovale wallikeri, which may be distinguished by the gene potra encoding P. ovale tryptophan-rich antigen. The sequence and size of potra gene was variable between the two P. ovale spp., and more fragment sizes were found compared to previous studies. Further information about the diversity of potra genes in these two P. ovale spp. will be needed. METHODS: A total of 110 dried blood samples were collected from the clinical patients infected with P. ovale, who all returned from Africa in Henan Province in 2011-2016. The fragments of potra were amplified by nested PCR. The sizes and species of potra gene were analysed after sequencing, and the difference between the isolates were analysed with the alignment of the amino acid sequences. The phylogenetic tree was constructed by neighbour-joining to determine the genetic relationship among all the isolates. The distribution of the isolates was analysed based on the origin country. RESULTS: Totally 67 samples infected with P. o. wallikeri, which included 8 genotypes of potra, while 43 samples infected with P. o. curtisi including 3 genotypes of potra. Combination with the previous studies, P. o. wallikeri had six sizes, 227, 245, 263, 281, 299 and 335 bp, and P. o. curtisi had four sizes, 299, 317, 335 and 353 bp, the fragment sizes of 299 and 335 bp were the overlaps between the two species. Six amino acid as one unit was firstly used to analyse the amino acid sequence of potra. Amino acid sequence alignment revealed that potra of P. o. wallikeri differed in two amino acid units, MANPIN and AITPIN, while potra of P. o. curtisi differed in amino acid units TINPIN and TITPIS. Combination with the previous studies, there were ten subtypes of potra exiting for P. o. wallikeri and four subtypes for P. o. curtisi. The phylogenetic tree showed that 11 isolates were divided into two clusters, P. o. wallikeri which was then divided into five sub-clusters, and P. o. curtisi which also formed two sub-clusters with their respective reference sequences. The genetic relationship of the P. ovale spp. mainly based on the number of the dominant amino acid repeats, the number of MANPIN, AITPIN, TINPIN or TITPIS. The genotype of the 245 bp size for P. o. wallikeri and that of the 299 and 317 bp size for P. o. curtisi were commonly exiting in Africa. CONCLUSION: This study further proved that more fragment sizes were found, P. o. wallikeri had six sizes, P. o. curtisi had four sizes. There were ten subtypes of potra exiting for P. o. wallikeri and four subtypes for P. o. curtisi. The genetic polymorphisms of potra provided complementary information for the gene tracing of P. ovale spp. in the malaria elimination era.

Evidence of non-Plasmodium falciparum malaria infection in Kédougou, Sénégal.

BACKGROUND: Expanded malaria control efforts in Sénégal have resulted in increased use of rapid diagnostic tests (RDT) to identify the primary disease-causing Plasmodium species, Plasmodium falciparum. However, the type of RDT utilized in Sénégal does not detect other malaria-causing species such as Plasmodium ovale spp., Plasmodium malariae, or Plasmodium vivax. Consequently, there is a lack of information about the frequency and types of malaria infections occurring in Sénégal. This study set out to better determine whether species other than P. falciparum were evident among patients evaluated for possible malaria infection in Kédougou, Sénégal. METHODS: Real-time polymerase chain reaction speciation assays for P. vivax, P. ovale spp., and P. malariae were developed and validated by sequencing and DNA extracted from 475 Plasmodium falciparum-specific HRP2-based RDT collected between 2013 and 2014 from a facility-based sample of symptomatic patients from two health clinics in Kédougou, a hyper-endemic region in southeastern Sénégal, were analysed. RESULTS: Plasmodium malariae (n = 3) and P. ovale wallikeri (n = 2) were observed as co-infections with P. falciparum among patients with positive RDT results (n = 187), including one patient positive for all three species. Among 288 negative RDT samples, samples positive for P. falciparum (n = 24), P. ovale curtisi (n = 3), P. ovale wallikeri (n = 1), and P. malariae (n = 3) were identified, corresponding to a non-falciparum positivity rate of 2.5%. CONCLUSIONS: These findings emphasize the limitations of the RDT used for malaria diagnosis and demonstrate that non-P. falciparum malaria infections occur in Sénégal. Current RDT used for routine clinical diagnosis do not necessarily provide an accurate reflection of malaria transmission in Kédougou, Sénégal, and more sensitive and specific methods are required for diagnosis and patient care, as well as surveillance and elimination activities. These findings have implications for other malaria endemic settings where species besides P. falciparum may be transmitted and overlooked by control or elimination activities.

Species and genotype diversity of Plasmodium in malaria patients from Gabon analysed by next generation sequencing.

BACKGROUND: Six Plasmodium species are known to naturally infect humans. Mixed species infections occur regularly but morphological discrimination by microscopy is difficult and multiplicity of infection (MOI) can only be evaluated by molecular methods. This study investigated the complexity of Plasmodium infections in patients treated for microscopically detected non-falciparum or mixed species malaria in Gabon. METHODS: Ultra-deep sequencing of nucleus (18S rRNA), mitochondrion, and apicoplast encoded genes was used to evaluate Plasmodium species diversity and MOI in 46 symptomatic Gabonese patients with microscopically diagnosed non-falciparum or mixed species malaria. RESULTS: Deep sequencing revealed a large complexity of confections in patients with uncomplicated malaria, both on species and genotype levels. Mixed infections involved up to four parasite species (Plasmodium falciparum, Plasmodium malariae, Plasmodium ovale curtisi, and P. ovale wallikeri). Multiple genotypes from each species were determined from the asexual 18S rRNA gene. 17 of 46 samples (37%) harboured multiple genotypes of at least one Plasmodium species. The number of genotypes per sample (MOI) was highest in P. malariae (n = 4), followed by P. ovale curtisi (n = 3), P. ovale wallikeri (n = 3), and P. falciparum (n = 2). The highest combined genotype complexity in samples that contained mixed-species infections was seven. CONCLUSIONS: Ultra-deep sequencing showed an unexpected breadth of Plasmodium species and within species diversity in clinical samples. MOI of P. ovale curtisi, P. ovale wallikeri and P. malariae infections were higher than anticipated and contribute significantly to the burden of malaria in Gabon.

A novel diagnostic method for malaria using loop-mediated isothermal amplification (LAMP) and MinION™ nanopore sequencer.

BACKGROUND: A simple and accurate molecular diagnostic method for malaria is urgently needed due to the limitations of conventional microscopic examination. In this study, we demonstrate a new diagnostic procedure for human malaria using loop mediated isothermal amplification (LAMP) and the MinION™ nanopore sequencer. METHODS: We generated specific LAMP primers targeting the 18S-rRNA gene of all five human Plasmodium species including two P. ovale subspecies (P. falciparum, P. vivax, P. ovale wallikeri, P. ovale curtisi, P. knowlesi and P. malariae) and examined human blood samples collected from 63 malaria patients in Indonesia. Additionally, we performed amplicon sequencing of our LAMP products using MinION™ nanopore sequencer to identify each Plasmodium species. RESULTS: Our LAMP method allowed amplification of all targeted 18S-rRNA genes of the reference plasmids with detection limits of 10-100 copies per reaction. Among the 63 clinical samples, 54 and 55 samples were positive by nested PCR and our LAMP method, respectively. Identification of the Plasmodium species by LAMP amplicon sequencing analysis using the MinION™ was consistent with the reference plasmid sequences and the results of nested PCR. CONCLUSIONS: Our diagnostic method combined with LAMP and MinION™ could become a simple and accurate tool for the identification of human Plasmodium species, even in resource-limited situations.

Comparison of imported Plasmodium ovale curtisi and P. ovale wallikeri infections among patients in Spain, 2005-2011.

Sequencing data from Plasmodium ovale genotypes co-circulating in multiple countries support the hypothesis that P. ovale curtisi and P. ovale wallikeri are 2 separate species. We conducted a multicenter, retrospective, comparative study in Spain of 21 patients who had imported P. ovale curtisi infections and 14 who had imported P. ovale wallikeri infections confirmed by PCR and gene sequencing during June 2005-December 2011. The only significant finding was more severe thrombocytopenia among patients with P. ovale wallikeri infection than among those with P. ovale curtisi infection (p = 0.031). However, we also found nonsignificant trends showing that patients with P. ovale wallikeri infection had shorter time from arrival in Spain to onset of symptoms, lower level of albumin, higher median maximum core temperature, and more markers of hemolysis than did those with P. ovale curtisi infection. Larger, prospective studies are needed to confirm these findings.

The two parasite species formerly known as Plasmodium ovale.

Plasmodium ovale was the last of the exclusively human malaria parasites to be described, in 1922, and has remained the least well studied. Beginning in 1995, two divergent forms of the parasite, later termed 'classic' and 'variant', were described. By 2010, it was realised that these forms are two closely related, but genetically distinct and non-recombining species; they were given the names Plasmodium ovale curtisi and Plasmodium ovale wallikeri. Since then, substantial additional data have confirmed that the two parasites are indeed separate species, but the trinomial nomenclature has often led to confusion about their status, with many authors describing them as subspecies. We hereby formally name them Plasmodium ovalecurtisi and Plasmodium ovalewallikeri.

A case of Plasmodium ovale wallikeri infection in a Chinese worker returning from West Africa.

In contrast to the gradual reduction in the number of locally transmitted malaria cases in China, the number of imported malaria cases has been increasing since 2008. Here, we report a case of a 39-year-old Chinese man who acquired Plasmodium ovale wallikeri infection while staying in Ghana, West Africa for 6 months in 2012. Microscopic examinations of Giemsa-stained thin and thick blood smears indicated Plasmodium vivax infection. However, the results of rapid diagnostic tests, which were conducted 3 times, were not in agreement with P. vivax. To further check the diagnosis, standard PCR analysis of the small-subunit rRNA gene was conducted, based on which a phylogeny tree was constructed. The results of gene sequencing indicated that this malaria is a variant of P. ovale (P. ovale wallikeri). The infection in this patient was not a new infection, but a relapse of the infection from the one that he had contracted in West Africa.

Genetic Diversity and Phylogenetic Relatedness of Plasmodium ovale curtisi and Plasmodium ovale wallikeri in sub-Saharan Africa.

P. ovale was until recently thought to be a single unique species. However, the deployment of more sensitive tools has led to increased diagnostic sensitivity, including new evidence supporting the presence of two sympatric species: P. ovale curtisi (Poc) and P. ovale wallikeri (Pow). The increased reports and evolution of P. ovale subspecies are concerning for sub-Saharan Africa where the greatest burden of malaria is borne. Employing published sequence data, we set out to decipher the genetic diversity and phylogenetic relatedness of P. ovale curtisi and P. ovale wallikeri using the tryptophan-rich protein and small subunit ribosomal RNA genes from Gabon, Senegal, Ethiopia and Kenya. Higher number of segregating sites were recorded in Poc isolates from Gabon than from Ethiopia, with a similar trend in the number of haplotypes. With regards to Pow, the number of segregating sites and haplotypes from Ethiopia were higher than from those in Gabon. Poc from Kenya, had higher segregating sites (20), and haplotypes (4) than isolates from Senegal (8 and 3 respectively), while nucleotide from Senegal were more diverse (θw = 0.02159; π = 0.02159) than those from Kenya (θw = 0.01452; π = 0.01583). Phylogenetic tree construction reveal two large clades with Poc from Gabon and Ethiopia, and distinct Gabonese and Ethiopian clades on opposite ends. A similar observation was recorded for the phylogeny of Poc isolates from Kenya and Senegal. With such results, there is a high potential that ovale malaria control measures deployed in one country may be effective in the other since parasite from both countries show some degree of relatedness. How this translates to malaria control efforts throughout the continent would be next step deserving more studies.

Epidemiological, Physiological and Diagnostic Comparison of Plasmodium ovale curtisi and Plasmodium ovale wallikeri.

Nowadays, Plasmodium ovale is divided into two non-recombinant sympatric species: Plasmodium ovale wallikeri and Plasmodium ovale curtisi. In this mini review, we summarize the available knowledge on the clinical/biological aspects of P. ovale spp. malaria and current techniques for the diagnosis/characterisation of P. ovale curtisi and P. ovale wallikeri. P. ovale wallikeri infections are characterized by a deeper thrombocytopenia and shorter latency compared to P. ovale curtisi infections, indicating that P. ovale wallikeri is more pathogenic than P. ovale curtisi. Rapid diagnosis for effective management is difficult for P. ovale spp., since specific rapid diagnostic tests are not available and microscopic diagnosis, which is recognized as the gold standard, requires expert microscopists to differentiate P. ovale spp. from other Plasmodium species. Neglect in addressing these issues in the prevalence of P. ovale spp. represents the existing gap in the fight against malaria.