Adaptation of Plasmodium falciparum to its transmission environment.

Success in eliminating malaria will depend on whether parasite evolution outpaces control efforts. Here, we show that Plasmodium falciparum parasites (the deadliest of the species causing human malaria) found in low-transmission-intensity areas have evolved to invest more in transmission to new hosts (reproduction) and less in within-host replication (growth) than parasites found in high-transmission areas. At the cellular level, this adaptation manifests as increased production of reproductive forms (gametocytes) early in the infection at the expense of processes associated with multiplication inside red blood cells, especially membrane transport and protein trafficking. At the molecular level, this manifests as changes in the expression levels of genes encoding epigenetic and translational machinery. Specifically, expression levels of the gene encoding AP2-G-the transcription factor that initiates reproduction-increase as transmission intensity decreases. This is accompanied by downregulation and upregulation of genes encoding HDAC1 and HDA1-two histone deacetylases that epigenetically regulate the parasite's replicative and reproductive life-stage programmes, respectively. Parasites in reproductive mode show increased reliance on the prokaryotic translation machinery found inside the plastid-derived organelles. Thus, our dissection of the parasite's adaptive regulatory architecture has identified new potential molecular targets for malaria control.

The changing malaria landscape in Aseer region, Kingdom of Saudi Arabia: 2000-2015.

BACKGROUND: In 2004, a revised action plan was developed, supported by the World Health Organization, to eliminate malaria from Saudi Arabia by preventing re-introduction of malaria into regions since declared malaria free, eliminating foci of transmission in the Mecca and Medina areas and a concerted effort of foci surveillance and control, to eliminate malaria from the regions of Jazan and Aseer. This paper provides the context, activities, progress, and possible contributions toward malaria elimination in the Aseer region since 2000, with a more detailed analysis of the spatial location of locally acquired case incidence since 2012. METHODS: This is a descriptive study of all available Ministry of Health surveillance data and process reports since 2000, with higher spatial resolution analysis of data between 2012 and 2015. RESULTS: In 2000, there were 511 cases of Plasmodium falciparum locally acquired infection. The following 4 years witnessed a dramatic decline in cases to only 18 locally acquired infections reported in 2005. A resurgence in local infections was reported in 2006 (93) and 2007 (165), thereafter (2008-2014) local cases continued to decline to fewer than 40 per year across the region. However, in 2015, a small rise was noted (51). All locally acquired infections were P. falciparum. There has been a constant flow of imported infections into Aseer since 2000, mostly among immigrant labour from Pakistan, India, Sudan, and Yemen. Imported infections have included both Plasmodium vivax and P. falciparum. The spatial extent of malaria appears to be changing, but there remain two intractable areas Sarat Abeda and Dhran Aljanub, where risks per reporting centre have changed little since 2001, remaining above 0.5 per 10,000 population. Only seven villages contributed 55% of all locally acquired infection since 2012. DISCUSSION: Aseer has reached a state of very low incidence of locally acquired infections, despite a constant source of imported infections from outside the country. How many of the local infections are F2 generations from imported infections or how many are a result of residual active transmission between asymptomatic carriers of infections transmitted by pockets of existing Anopheles arabiensis populations remains unknown. A more detailed investigation of the spatial and temporal patterns of infected hosts, parasites and vectors would help define whether this region has managed to effectively prevent local transmission of new infections.

Progress toward malaria elimination in Jazan Province, Kingdom of Saudi Arabia: 2000-2014.

BACKGROUND: The draft Global Technical Strategy for malaria aims to eliminate malaria from at least 10 countries by 2020. Yemen and Saudi Arabia remain the last two countries on the Arabian Peninsula yet to achieve elimination. Over the last 50 years, systematic efforts to control malaria in the Kingdom of Saudi Arabia has successfully reduced malaria cases to a point where malaria is now constrained largely to Jazan Province, the most south-western area along the Red Sea. The progress toward elimination in this province is reviewed between 2000 and 2014. METHODS: Data were obtained from the Ministry of Health case-reporting systems, activity reports, unpublished consultants reports, and relevant scientific published papers. Sub-provincial population data were obtained the national household censuses undertaken in 2004 and 2010. Rainfall data were obtained from the Meteorological Department in Jazan. RESULTS: Between 2000 and 2014 there were 5522 locally acquired cases of malaria and 9936 cases of imported malaria. A significant reduction in locally acquired malaria cases was observed from 2000 to 2014, resulting in an average annual incidence (2010-2014) of 0.3 cases per 10,000 population. Conversely imported cases, since 2000, remain consistent and higher than locally acquired cases, averaging between 250 and 830 cases per year. The incidence of locally acquired cases is heterogeneous across the Province, with only a few health districts contributing the majority of the cases. The overall decline in malaria case incidence can be attributed to coincidental expansion of control efforts and periods of exceptionally low rainfall. CONCLUSIONS: Jazan province is poised to achieve malaria elimination. There is a need to change from a policy of passive case detection to reactively and proactively detecting infectious reservoirs that require new approaches to surveillance. These should be combined with advanced epidemiological tools to improve the definitions of epidemiological receptive and hotspot malaria risk mapping. The single largest threat currently remains the risks posed by imported infections from Yemen.

Genetic Diversity of Plasmodium falciparum Field Isolates in Central Sudan Inferred by PCR Genotyping of Merozoite Surface Protein 1 and 2.

BACKGROUND: Characterization of Plasmodium falciparum diversity is commonly achieved by amplification of the polymorphic regions of the merozoite surface proteins 1 (MSP1) and 2 (MSP2) genes. AIMS: The present study aimed to determine the allelic variants distribution of MSP1 and MSP2 and multiplicity of infection in P. falciparum field isolates from Kosti, central Sudan, an area characterized by seasonal malaria transmission. MATERIALS AND METHODS: Total 121 samples (N = 121) were collected during a cross-sectional survey between March and April 2003. DNA was extracted and MSP1 and MSP2 polymorphic loci were genotyped. RESULTS: The total number of alleles identified in MSP1 block 2 was 11, while 16 alleles were observed in MSP2 block 3. In MSP1, RO33 was found to be the predominant allelic type, carried alone or in combination with MAD20 and K1 types, whereas FC27 family was the most prevalent in MSP2. Sixty two percent of isolates had multiple genotypes and the overall mean multiplicity of infection was 1.93 (CI 95% 1.66-2.20). Age correlated with parasite density (P = 0.017). In addition, a positive correlation was observed between parasite densities and the number of alleles (P = 0.022). CONCLUSION: Genetic diversity in P. falciparum field isolates in central Sudan was high and consisted of multiple clones.

Genotyping of human papillomavirus in paraffin embedded cervical tissue samples from women in Ethiopia and the Sudan.

Cervical cancer is the most frequent female malignancy in most developing countries. Previous studies have demonstrated a strong association of human papillomavirus (HPV) infection with dysplasia and carcinoma of the uterine cervix. The objective of this study was to identify the prevailing HPV genotypes responsible for the development of cervical cancer among women in Ethiopia and the Sudan. A molecular characterization of HPV was done on 245 paraffin embedded cervical biopsy samples collected from the two countries. Amplification of HPV and subsequent genotyping was done using SPF10 primers and Line probe assay. Of samples collected from Ethiopian patients, 93% (149/160) and 13% (21/160) had high risk and low risk HPV genotypes, respectively. Among samples collected from the Sudan, 94% (80/85) harbored high risk and 11.7% (10/85) low risk HPV genotypes. Human papillomavirus 16 was the most frequent genotype identified in samples from Ethiopia (91%, 136/149) and the Sudan (82.5%, 66/80). HPV 52, 58, and 18 were the second, third and fourth common genotypes identified in Ethiopia, whereas HPV 18, 45, and 52 were the second, third, and fourth genotypes identified in samples collected from the Sudan. Thus, individuals living in different geographical localities should receive vaccines based on the specific genotypes circulating in the area and a vaccine targeting HPV 16, 18, 45, 52, and 58 may be optimal for the control of cervical cancer in the two countries.

Identifying residual foci of Plasmodium falciparum infections for malaria elimination: the urban context of Khartoum, Sudan.

BACKGROUND: Identifying the location and size of residual foci of infections is critical where malaria elimination is the primary goal. Here the spatial heterogeneity of Plasmodium falciparum infections within the urban extent of Khartoum state in Sudan is investigated using data from cross-sectional surveys undertaken from 1999 to 2008 to inform the Khartoum Malaria Free Initiative (KMFI). METHODS: From 1999-2008 the KMFI undertook cross-sectional surveys of 256 clusters across 203 random samples of residential blocks in the urban Khartoum state in September of each year. Within sampled blocks, at least five persons, including at least one child under the age of five years, were selected from each household. Blood smears were collected from the sampled individuals to examine the presence of P. falciparum parasites. Residential blocks were mapped. Data were analysed for spatial clustering using the Bernoulli model and the significance of clusters were tested using the Kulldorff scan statistic. RESULTS: A total of 128,510 malaria slide examinations were undertaken during the study period. In 1999, overall prevalence was 2.5%, rising to 3.2% in 2000 and consistently staying below 1% in subsequent years. From 2006, over 90% of all surveyed clusters reported no infections. Spatial clustering of infections was present in each year but not statistically significant in the years 2001, 2002, 2004 and 2008. Spatial clusters of high infection were often located at the junction of the Blue and White Niles. CONCLUSION: Persisting foci of malaria infection in Khartoum are likely to distort wide area assessments and disproportionately affect future transmission within the city limits. Improved investments in surveillance that combines both passive and active case detection linked to a geographic information system and a more detailed analysis of the location and stability of foci should be undertaken to facilitate and track malaria elimination in the state of Khartoum.

Malaria infection by sporozoite challenge induces high functional antibody titres against blood stage antigens after a DNA prime, poxvirus boost vaccination strategy in Rhesus macaques.

BACKGROUND: A DNA prime, poxvirus (COPAK) boost vaccination regime with four antigens, i.e. a combination of two Plasmodium knowlesi sporozoite (csp/ssp2) and two blood stage (ama1/msp142) genes, leads to self-limited parasitaemia in 60% of rhesus monkeys and survival from an otherwise lethal infection with P. knowlesi. In the present study, the role of the blood stage antigens in protection was studied in depth, focusing on antibody formation against the blood stage antigens and the functionality thereof. METHODS: Rhesus macaques were immunized with the four-component vaccine and subsequently challenged i.v. with 100 P. knowlesi sporozoites. During immunization and challenge, antibody titres against the two blood stage antigens were determined, as well as the in vitro growth inhibition capacity of those antibodies. Antigen reversal experiments were performed to determine the relative contribution of antibodies against each of the two blood stage antigens to the inhibition. RESULTS: After vaccination, PkAMA1 and PkMSP119 antibody titres in vaccinated animals were low, which was reflected in low levels of inhibition by these antibodies as determined by in vitro inhibition assays. Interestingly, after sporozoite challenge antibody titres against blood stage antigens were boosted over 30-fold in both protected and not protected animals. The in vitro inhibition levels increased to high levels (median inhibitions of 59% and 56% at 6 mg/mL total IgG, respectively). As growth inhibition levels were not significantly different between protected and not protected animals, the ability to control infection appeared cannot be explained by GIA levels. Judged by in vitro antigen reversal growth inhibition assays, over 85% of the inhibitory activity of these antibodies was directed against PkAMA1. CONCLUSIONS: This is the first report that demonstrates that a DNA prime/poxvirus boost vaccination regimen induces low levels of malaria parasite growth inhibitory antibodies, which are boosted to high levels upon challenge. No association could, however, be established between the levels of inhibitory capacity in vitro and protection, either after vaccination or after challenge.

Improved outcome for children with acute lymphoblastic leukemia after risk-adjusted intensive therapy: a single-institution experience.

BACKGROUND AND OBJECTIVE: Because of the need for more comprehensive information on the least toxic and most effective forms of therapy for children with acute lymphoblastic leukemia (ALL), we reviewed our experience in the treatment of children with ALL at King Faisal Specialist Hospital and Research Centre (KFSH&RC) and King Fahad National Center for Children's Cancer and Research (KFNCCC&R) over a period of 18 years with a focus on patient characteristics and outcome. METHODS: During the period of 1981 to 1998, records of children with ALL were retrospectively reviewed with respect to clinical presentation, laboratory findings, risk factors, stratification, therapy and outcome. The protocols used in treatment included 4 local protocols (KFSH 81, 84, 87 and 90), and subsequently, Children's Cancer Group (CCG) protocols, and these were grouped as Era 1 (1981-1992) and Era 2 (1993-1998). RESULTS: Of 509 children with ALL treated during this period, 316 were treated using local protocols and 193 using CCG protocols. Drugs used in Era 1 included a 4-drug induction using etoposid (VP-16) instead of L-asparaginase. Consolidation was based on high dose methotrexate (MTX) 1 g/m(2) and maintenance was based on oral mercaptopurine (6-MP) and MTX with periodic pulses using intravenous teniposide (VM-26), Ara-C, L-asparaginase, adriamycin, prednisone, VP-16 and cyclophosphamide. International protocols were introduced in Era 2, which was also marked by intensification of early treatment, a wider selection of cytoreductive agents, and the alternating use of non-cross-resistant pairs of drugs during the post-remission period. The end-of-induction remission rate improved from 90% in Era 1 to 95% in Era 2, which was of borderline statistical significance (P=.049). The 5-year event-free survival (EFS) improved from 30.6% in Era 1 to 64.2% in Era 2 (P<.001). Improvement in outcome was achieved without any significant increase in morbidity or mortality, due to improvement in both systemic therapy and supportive care. The most important independent prognostic factors were intensity of therapy, poor risk category assignment and CNS disease at diagnosis. CONCLUSION: Outcome in children with ALL has improved because of intensification of treatment protocols and better supportive care.

VIP (etoposide, ifosfamide, cisplatin) in adult patients with recurrent or refractory Ewing sarcoma family of tumors.

Despite the fact that Ewing sarcoma family of tumors (ET) is chemosensitive, long-term survival is extremely rare for patients with primary refractory or recurrent disease. There is no standard salvage chemotherapy regimen available in this context. In this study the authors reviewed their experience with the combination of etoposide, ifosfamide, and cisplatin in adult patients with recurrent or refractory disease. From February 1997 through December 2001, they evaluated the efficacy of etoposide (75 mg/m2/day for 5 days), ifosfamide (1,200 mg/m2/day for 5 days), and cisplatin (20 mg/m2/day for 5 days) combination chemotherapy (VIP regimen), as second-line salvage therapy in 27 patients with recurrent or refractory ET. All patients were evaluated for response, time to progression, and overall survival. Twenty-one male and 6 female patients with recurrent (n = 14) and refractory (n = 13) disease were treated with the VIP regimen. Median age was 18 years (range, 16-34 years). Twenty-two patients were previously treated with vincristine, Adriamycin, ifosfamide, and actinomycin-D; and 5 patients were treated with cyclophosphamide, Adriamycin, and vincristine. Sites of recurrent or progressive disease included local (n = 3), distant (n = 11), and both local and distant (n = 13). A total of 129 cycles of VIP were given (median, 5 cycles/patient; range, 1-14 cycles/patient). One patient (4%) had a complete response (CR) and 8 patients (30%) had a partial response (PR), for an overall response rate of 34%. The median number of cycles given to patients with CR + PR was 6 (range, 3-14 cycles). Nine patients (33%) had stable disease and 9 (33%) had disease progression. Median time to progression and median overall survival were 6.6 months and 8.1 months respectively for all patients, and 12.8 months and 14.2 months respectively for responders. There were no toxic deaths. Major toxicities included grade IV granulocytopenia in 19 patients and grades III/IV thrombocytopenia in 15 patients. At a median follow-up of 8 months (range, 2-56 months), 24 patients died of disease progression, 2 patients are alive with disease, and 1 patient is alive with no evidence of disease. The authors conclude that the VIP combination is active in patients with recurrent/refractory ET, with acceptable toxicity, and offers good palliation. Cisplatin-based combination chemotherapy merits further investigation, possibly as first-line treatment in this disease.