Single Nucleotide Polymorphism Analysis of Pvmdr-1 in Plasmodium Vivax Isolated from Military Personnel of Republic of Korea in 2016 and 2017

Background: Malaria chemoprophylaxis using chloroquine (CQ) and primaquine (PQ) has been administered to resident soldiers in the 3rd Army of Republic of Korea (ROK) to prevent malaria infection since the year 1997. Due to mass chemoprophylaxis against malaria, concern exists about occurrence of chloroquine resistance (CQR). Herein, we investigated the single nucleotide polymorphisms (SNPs) of the Plasmodium vivax multi-drug resistance protein-1 (pvmdr-1) gene to monitor the risk of CQR. Methods: SNPs of the pvmdr-1 gene were analyzed in 73 soldiers of the 3rd Army of ROK diagnosed with infection by Plasmodium vivax (P. vivax). Results: Quintuple mutations (G698S, L845F, M908L, T958M, and F1076L) were detected in 73 soldiers. Mutation in the Y541C position was firstly detected in soldiers at a frequency of 1.3% (1/73). In addition, synonymous mutations were detected at positions K44, L493, T529, and E1233. Based on these SNPs, pvmdr-1 sequences of ROK were classified into 6 haplotypes. The phylogenetic analysis closed to Type of North Korean showed that P. vivax malaria of ROK could be a reason of influx from North Korea. In this study, there was no therapeutic resistance (CQ-mediated parasite clearance within 72 hours) for clinical samples that possessed various SNPs of pvmdr-1. Various SNPs including a newly identified non-synonymous mutation (Y541C) had been introduced into P. vivax malaria-endemic areas in ROK. Conclusions: Our study showed that synonymous and non-synonymous mutations of pvmdr-1 were introduced to the malaria chemoprophylaxis-executed regions of ROK from 2016 to 2017. Thus, to prevent the emergence of CQR, continuous surveillance for SNPs of pvmdr-1 related with CQR in the malaria-endemic regions of ROK is essential.

735. Malaria Chemoprophylaxis Adherence Among U.S. Active Duty Service Members during Deployment to Endemic Regions

Background Military members frequently deploy to malaria-endemic regions. Most cases of travel-related malaria occur due to prophylaxis non-adherence, impacting mission readiness. Factors assessing adherence are described in outbreak settings; we prospectively assess adherence in military travelers. Methods TravMil is a prospective, observational cohort study of US military beneficiaries traveling outside the US (2010-2019). Our analysis includes only active-duty service members traveling with a military purpose to malaria-endemic regions, who were prescribed malaria prophylaxis, and who completed a pre- and post-deployment survey; they could also enroll after return from deployment. All travelers received pre-travel counseling. Survey responses were assessed using descriptive statistics and multivariate regression to determine risk factors for adherence. Results 1504 travelers were included (85% male; median age 28 years; 73% white). Median duration of travel was 77 days (12% traveled ≤ 14 days). Africa was the most common destination (33%). Primary prophylaxis included doxycycline (54%) and atovaquone/proguanil (43%). 969 (64%) were fully adherent to their regimen. The frequency of prophylaxis did not match expected values, as 3.6% of subjects reported taking prophylaxis weekly, and 2.9% did not know how often they took it. 103 (6.9%) did not take any of the prescribed regimen. On multivariate analysis, deployers were more likely to adhere if they traveled for ≤ 14 days or to Africa or practiced other mosquito-avoidance behaviors. Study enrollment post-deployment was associated with decreased odds of adherence, as was use of a tent. The use of daily versus weekly prophylaxis was not associated with a difference in adherence, though we had limited subjects prescribed weekly regimens. Figure 1. Reasons for not taking any of the prescribed chemoprophylaxis (n = 103) Table 1. Odds of full adherence to malaria chemoprophylaxis on multivariate logistic analysis Conclusion Short-duration travel, travel to highly endemic regions, and mosquito-avoidance behaviors were associated with increased adherence to prophylaxis. The lower rate of adherence in post-deployment enrollees may be a surrogate for inadequate counseling or recall bias. Our study highlights potential holes in counseling regarding malaria prophylaxis and the importance of ongoing provider and patient education on malaria. Disclosures Heather Yun, MD, American Board of Internal Medicine (Individual(s) Involved: Self): Board Member

Study Of Malaria Cases Among Seafarers In Croatia And The Causes Of Ineffective Chemoprophylaxis Among Them

Introduction: Malaria in Croatia was endemic until 1954, when the last autochthonous cases were reported. Now we have imported cases and the disease still most commonly affects seafarers and workers temporarily employed in malaria endemic countries. The aim of this study was to investigate ineffective malaria chemoprophylaxis among Croatian seafarers. Materials and methods: This study used the data collected by questionnaires that followed every imported case of malaria in Croatia. In addition, a survey was conducted among the seafarers attending the Special education program at the Faculty of Maritime Studies in Split, regarding their attitudes / reasons for not using chemoprophylaxis using a short questionnaire. Results: From the 1st January 2007 to 31st December 2019 there was a total of 108 imported malaria cases in Croatia, of which 22 (20.37%) referred to seafarers. In situations reporting the information whether the infected seafarers used chemoprophylaxis, twenty seafarers (91% of the overall infected) responded that they had not used chemoprophylaxis, while two of them (9.0%) responded that they had used it incorrectly. Conclusion: It appears that the main reason for avoiding chemoprophylaxis is the fear of side-effects. Some of the respondents, particularly the young ones, stated that they were not vaccinated – which is a proof of ignorance, as the vaccine does not exist. How to familiarise and motivate maritime students and seafarers to participate actively in protection against malaria – this is an issue that is important not only in Croatia but, presumably, in other countries as well.

Single Nucleotide Polymorphism Analysis of Pvmdr-1 in Plasmodium Vivax Isolated From Military Personnel of Republic of Korea in 2016 and 2017

Background: Malaria chemoprophylaxis using chloroquine and primaquine has been administered to resident soldiers in the 3rd Army of Republic of Korea (ROK) to prevent malaria infection since the year 1997. Due to mass chemoprophylaxis against malaria, concern exists about occurrence of chloroquine resistance. Herein, we investigated the single nucleotide polymorphisms (SNPs) of the Plasmodium vivax multi-drug resistance protein-1 (Pvmdr-1) gene to monitor the risk of chloroquine resistance.Methods: To evaluate the risk of malaria chemoprophylaxis, SNPs of the Pvmdr-1 gene were analysed in 73 soldiers of the 3rd Army of ROK diagnosed with infection by Plasmodium vivax. Results: Quintuple mutations (G698S, L845F, M908L, T958M, and F1076L) were detected in 73 soldiers. Mutation in the Y541C position was detected in soldiers at a frequency of 1.3% (1/73). In addition, silent mutations were detected at positions 44K, 493L, 529T, and 1233E. Based on these SNPs, Pvmdr-1 sequences of ROK were classified into 6 haplotypes. Phylogenetic analysis showed that the neighbourhood of the 6 haplotypes were Chaina_NB-16 and Papua New Guinea-PNG58 (Figure 1).Conclusions: Genetic- or phenotypic-based chloroquine resistance was not observed. However, various SNPs including a newly identified non-synonymous mutation (Y541C) have been introduced into Plasmodium vivax malaria endemic areas in ROK. Thus, to prevent the emergence of chloroquine resistance, continuous surveillance for SNPs of Pvmdr-1 related with chloroquine resistance is essential in the malaria chemoprophylaxis-executed regions of ROK.

Study protocol of a cluster randomized controlled trial of strategies to increase antenatal iron and folic acid supplementation and malaria prophylaxis in rural south-central Côte d’Ivoire

Background Coverage of antenatal iron and folic acid supplementation (IFAS) and intermittent preventive treatment of malaria in pregnancy (IPTp) remains low in many countries. Evidence on the most effective ways to increase both IFASIPTp is mixed overall, with only few studies directly identifying cost-effective ways to increase coverage of both interventions. The proposed study aims to assess the cost, impact and relative cost-effectiveness of two complementary strategies of increasing IFAS and malaria chemoprophylaxis coverage among pregnant women relative to the current default system in a rural low-income setting of sub-Saharan Africa. Methods/design This study will be carried out in the Taabo health and demographic surveillance system (HDSS) in south-central Côte d’Ivoire. This is a cluster-randomized trial targeting 720 consenting pregnant women aged ≥15 years. The 118 clusters constituting the Taabo HDSS monitoring area will be randomly allocated to one of the following three groups with equal probability: a control group, an information only group, and an information plus home delivery group. To assess the relative effectiveness of each strategy, we will conduct an endline survey within the first 2 weeks after delivery. The primary outcomes of the trial will be maternal post-partum anaemia and malaria infection. Anaemia will be assessed using HEMOCUE devices; malaria infections will be assessed using standard rapid diagnostic tests named CareStart™ Malaria Pf (HRP2) Ag RDT (Multi Kit with capped lancet and inverted cup specimen transfer device). Other outcomes will include self-reported adherence to supplementation and malaria chemoprophylaxis, as well as miscarriages, stillbirths and low birth weight deliveries. Discussion This study will assess the cost-effectiveness of two alternative strategies to increase antenatal IFAS and malaria chemoprophylaxis coverage among pregnant women in rural Côte d’Ivoire and similar settings. Trial registration ClinicalTrials.govNCT04250428; Registered 31 January 2020.

An open label study of the safety and efficacy of a single dose of weekly chloroquine and azithromycin administered for malaria prophylaxis in healthy adults challenged with 7G8 chloroquine-resistant Plasmodium falciparum in a controlled human malaria infection model

Background Malaria remains the top infectious disease threat facing the U.S. military in many forward operating environments. Compliance with malaria chemoprophylaxis remains a critical component in preventing malaria in the deployed Service Member. Studies of previous military operations show that compliance is consistently higher with weekly versus daily dosing regimens. Current FDA approved weekly chemoprophylaxis options have contraindications that can limit prescribing. The combination of chloroquine (CQ) with azithromycin (AZ) has previously been shown to be an efficacious treatment option for malaria, has pharmacokinetics compatible with weekly dosing, and has shown synergy when combined in vitro. Methods In this open label study, 18 healthy volunteers, aged 18–50 years (inclusive), were randomly assigned to receive either 300 mg CQ or 300 mg CQ and 2 gm azithromycin (CQAZ) of directly observed therapy, weekly for 3 weeks prior to undergoing mosquito bite challenge with chloroquine-resistant Plasmodium falciparum. Volunteers that remained asymptomatic and had no evidence of parasitaemia continued to receive weekly post-exposure chemoprophylaxis for 3 weeks following malaria challenge. The primary endpoint was the number of volunteers that remained asymptomatic and had no evidence of parasitaemia 28 days after the malaria challenge. Results All 6 (100%) volunteers randomized to the CQ control group became symptomatic with parasitaemia during the 28-day post-challenge period. Only 1/12 (8.3%) of volunteers in the CQAZ group developed symptoms and parasitaemia during the 28-day post-challenge period. However, after chemoprophylaxis was discontinued an additional 6 volunteers developed parasitaemia between days 28–41 after challenge, with 4 of 6 experiencing symptoms. 80% of subjects in the CQAZ group experienced treatment related gastrointestinal adverse events (including 13% that experienced severe nausea) compared to 38% in the CQ group. A comparison of the pharmacokinetics in the CQAZ group demonstrated higher azithromycin Cmax (p = 0.03) and AUC (p = 0.044) levels in those volunteers who never became parasitaemic compared to those who did. Conclusion Given the high rate of side effects and poor efficacy when administered for 3 weeks before and after challenge, the combination of weekly chloroquine and azithromycin is a suboptimal regimen combination for weekly malaria chemoprophylaxis. Trial registration ClinicalTrials.gov NCT03278808

An open label study of the safety and efficacy of a single dose of weekly chloroquine and azithromycin administered for malaria prophylaxis in healthy adults challenged with 7G8 chloroquine-resistant Plasmodium falciparum in a controlled human malaria infection model

BackgroundMalaria remains the top infectious disease threat facing the U.S. military in many forward operating environments. Compliance with malaria chemoprophylaxis remains a critical component in preventing malaria in the deployed Service Member. Studies of previous military operations show that compliance is consistently higher with weekly versus daily dosing regimens. Current FDA approved weekly chemoprophylaxis options have contraindications that can limit prescribing. The combination of chloroquine (CQ) with azithromycin (AZ) has previously been shown to be an efficacious treatment option for malaria, has pharmacokinetics compatible with weekly dosing, and has shown synergy when combined in vitro. MethodsIn this open label study 18 healthy volunteers, aged 18-50 years (inclusive), were randomly assigned to receive either 300 mg CQ or 300 mg CQ and 2 gm azithromycin (CQAZ) of directly observed therapy, weekly for 3 weeks prior to undergoing mosquito bite challenge with chloroquine-resistant Plasmodium falciparum. Volunteers that remained asymptomatic and had no evidence of parasitaemia continued to receive weekly post-exposure chemoprophylaxis for 3 weeks following malaria challenge. The primary endpoint was the number of volunteers that remained asymptomatic and had no evidence of parasitaemia 28 days after the malaria challenge.ResultsAll 6 (100%) volunteers randomized to the CQ control group became symptomatic with parasitaemia during the 28-day post-challenge period. Only 1/12 (8.3%) of volunteers in the CQAZ group developed symptoms and parasitaemia during the 28-day post-challenge period. However, after chemoprophylaxis was discontinued an additional 6 volunteers developed parasitaemia between days 28-41 after challenge, with 4 of 6 experiencing symptoms. 80% of subjects in the CQAZ group experienced treatment related gastrointestinal adverse events (including 13 % that experienced severe nausea) compared to 38% in the CQ group. A comparison of the pharmacokinetics in the CQAZ group demonstrated higher azithromycin Cmax (p0.03) and AUC (p0.044) levels in those volunteers who never became parasitaemic compared to those who did. ConclusionGiven the high rate of side effects and poor efficacy when administered for 3 weeks before and after challenge, the combination of weekly chloroquine and azithromycin is a suboptimal regimen combination for weekly malaria chemoprophylaxis.Trial registration: ClinicalTrials.gov NCT03278808

Investigation of glucose-6-phosphate dehydrogenase (G6PD) deficiency prevalence in a Plasmodium vivax-endemic area in the Republic of Korea (ROK)

Background Glucose-6-phosphate dehydrogenase (G6PD) deficiency is the most prevalent inborn disorder. This X-chromosome-linked recessive disease affects more than 400 million people globally, and is associated with haemolytic anaemia after medication with the anti-latent malaria drug, primaquine. To prevent malaria, the Republic of Korea (ROK) Army administers malaria chemoprophylaxis. Due to the previously low G6PD deficiency prevalence in the ROK, prior to primaquine administration, testing for G6PD deficiency was not mandatory. In this study, to evaluate the risk from malaria chemoprophylaxis in the ROK, G6PD deficiency prevalence was investigated. Methods Blood specimens from 1,632 soldiers entering training camp for the 3 rd Infantry of the ROK Army were collected. CareStart™ Biosensor for G6PD and haemoglobin (Hb) was used to detect G6PD levels. G6PD variants using the DiaPlexC G6PD Genotyping kit (Asian type) and full-length sequencing were examined. Results Of 1,632 blood specimens tested, none was observed to be G6PD deficient. The median value of all tested samples was 7.582 U/g Hb. An investigation of 170 G6PD DNA variants was analysed and categorized as partially low normal [n = 131, 30–80% (2.27–6.05 U/g Hb)of the median value], high [n=3, >150% (>11.373 U/g Hb) of the median value], or normal [n = 36, 80–150% (6.05–11.373 U/g Hb) of the median value], and none was amplified by the DiaPlexC kit. Five silent mutations (C→T) in 38 partially abnormal specimens were found at the 1,311th nucleotide position by sequence analysis. Another 8 silent mutations (T93C) were also detected in 131 partially low normal specimens. Thus, it is inferred that these silent mutations could be related to G6PD activity. Conclusions This G6PD deficiency prevalence study, conducted among participants from the 3 rd Infantry of the ROK Army, provided crucial evidence for the safety of malaria chemoprophylaxis. This study showed found that the prevalence of G6PD deficiency among 1,632 young soldiers was wholly absent. Although G6PD phenotypic mutations were not detected, many silent mutations (C1311T and T93C) were observed. Thus, it is inferred that malaria chemoprophylaxis is relatively safe against G6PD deficiency-mediated haemolytic anaemia. However, given the number of individuals whose G6PD were at the low end of the normal range and the frequent detection of G6PD deficiency-related mutations, consistent monitoring of G6PD deficiency is needed. Keywords : Glucose-6-phosphate dehydrogenase deficiency, Prevalence, Single nucleotide polymorphism, Primaquine