Decreasing incidence of Plasmodium vivax in the Republic of Korea during 2010–2012

Background After the re-emergence of Plasmodium vivax in 1993, a total of 31,254 cases of vivax malaria were reported between 1993–2012 in the Republic of Korea (ROK). The purpose of this study was to review Korea Centers for Disease Control and Prevention records to investigate the transmission of malaria from 2010–2012. Methods Reporting of microscopy-diagnosed cases of malaria is mandatory in the ROK. In this study, all available records of malaria cases and malaria vectors collected from 2010 – 2012 in Cheorwon County, Gangwon Province and Ganghwa County, Incheon Metropolitan City, were reviewed. Results Although the number of cases of malaria peaked a third time in 2010 (1,772 cases) since the re-emergence of P. vivax, the incidence decreased two-fold to 838 in 2011 and three-fold to 555 in 2012. The number of cases decreased 52.7% in 2011 compared with that in 2010 and 33.8% in 2012 compared with that in 2011. However, the number of cases increased in Incheon Metropolitan City (15.3%) and Gyeongnam Province (23.1%) in 2012 compared with 2011. Of the 3,165 cases of vivax malaria in 2010–2012, 798 (25.2%) were in ROK military personnel, 519 (16.4%) in veterans, and 1,848 (58.4%) in civilians. In total, there were 2,666 male patients and 499 female patients, and the ratio of female to male patients increased from 1:7.9 in 2011 to 1:4.1 in 2012. Conclusions A rapid decrease in the incidence of malaria was observed in most areas from 2010 to 2012, but the incidence increased again in the western part of the demilitarized zone. Therefore, more intensive surveillance is needed throughout high risk areas to identify factors responsible for increase/decrease in the incidence of malaria in the ROK.

Conclusions: A rapid decrease in the incidence of malaria was observed in most areas from 2010 to 2012, but the incidence increased again in the western part of the demilitarized zone. Therefore, more intensive surveillance is needed throughout high risk areas to identify factors responsible for increase/decrease in the incidence of malaria in the ROK.

Background
Plasmodium vivax is the causative agent of relapsing benign tertian human malaria, the second most common type of malaria in humans, that afflicts several hundred million people annually in the world. Vivax malaria is a major public health problem in many tropical and semitropical regions and temperate countries, including the Democratic People's Republic of Korea (DPRK) and the Republic of Korea (ROK) [1].
The first scientific documentation of malaria was published in 1913, although it had been prevalent throughout the Korean peninsula for several centuries [2]. As a result of a national malaria eradication programme conducted in cooperation with the World Health Organization (WHO), the incidence of vivax malaria in the ROK rapidly decreased until in 1979 when WHO declared the ROK to be malaria free [3][4][5]. In the 1980's two sporadic cases were detected [6] and in 1993, a ROK soldier serving in northern Gyeonggi Province [7], and two civilians were diagnosed with vivax malaria [8]. Thereafter, a number of malaria cases were reported near the demilitarized zone (DMZ), which centers on Paju-si, Yeoncheon-gun, Cheorwon-gun, Gimpo-si, Ganghwa-gun, Goyang-si, and Dongducheon-si.
During the first few years of re-emergence of vivax malaria in the ROK, most cases occurred in ROK military personnel deployed near the DMZ. But some civilians who live in Daeseongdong where located inside of DMZ and Tongilchon surrounded by ROK military installations near the DMZ showed high infection rate as much as ROK military personnel. In addition, as the number of malaria cases increased, the ratio of civilian cases were increased [9].
Thus, it was of considerable concern that vivax malaria might become re-established through out the ROK as veteran soldiers returned to their hometowns throughout all of Korea [10][11][12].
The aim of this study was to determine the annual number of patients with malaria among military personnel, veterans, and civilians in the ROK; the mean daily incidence for weekly intervals; the geographic distribution of cases among military personnel and civilians; and the number of Anopheles mosquitoes captured by black light trap in the malaria risk from 2010-2012 to analyse the current status of malaria in the ROK.

Ethics statement
All participants were informed about the study methodology and provided written informed consent according to ethical standards. The study procedures, potential risks, and benefits were explained to all participants. All adult participants and parents/guardians in the household of participants under 18 years of age provided informed consent. Parents who were unwilling to have their children participate in the study were identified and their children subsequently excluded without prejudice from study surveys. All patient data were anonymous. The Human Ethics Committee of Inha University provided ethical approval, and this study was conducted according to the principles expressed in the Declaration of Helsinki.

Data collection from patients with malaria
Malaria is classified as one of the Group III communicable diseases that should be controlled by the Korean government. For the convenience of data analysis, patients with malaria were categorized as civilians, veterans, and military personnel. For civilians and veterans (who were diagnosed with vivax malaria within 24 months of discharge from military service in malaria epidemic areas), cases of malaria detected in private hospitals or clinics are reported to the local Public Health Center (PHC). The data collected by the PHCs is provided periodically to the Division of Infectious Disease Surveillance, Korea Centers for Disease Control and Prevention. For military personnel, cases of malaria are reported to the Office of the Surgeon General, Army Headquarters, and the Ministry of Health and Welfare [13]. The report usually contains the patient's name, age, sex, address, date of onset of illness, date of diagnosis of malaria, and estimated areas where possibly infected. Most cases of malaria diagnosed in civilian hospital or health clinics include veterans, but might also include ROK soldiers who were diagnosed while leave. However, these cases among ROK soldiers are excluded from the civilian/veteran populations by the Division of Infectious Disease Surveillance, since the patients are contacted directly as part of the verification process [14,15].

Definition of malaria
Thick and thin blood films obtained from suspected malaria patients were fixed with methanol and stained with Giemsa diluted with buffered water at pH 7.2 in accordance with standard procedure [16]. To estimate the densities of blood-stage parasites, the numbers of parasites were counted relative to 200 white blood cells (WBCs) and then the parasite/WBC ratio was multiplied by 8,000 (estimated number of WBCs per microlitre of blood) [17].

Geographic distribution of patients with malaria
The annual geographic distribution of P. vivax malaria in civilians, veterans, and ROK military personnel was determined by the patients residence and travel history during diagnosis. The seasonal incidence was analysed by grouping cases at weekly or monthly intervals.

Calculation of the annual parasite incidence
The annual parasite incidence (API) was calculated as the number of malaria-positive patients per 1,000 inhabitants for each of the study sites: API = (number of microscopically proven malaria cases/1,000 population/year).

Density of Anopheles mosquitoes
The number of Anopheles mosquitoes was monitored using a black light traps in Gyodong-myeon ( Figure 1A, I) and Samsan-myeon ( Figure 1A, II), Ganghwa County of Incheon Metropolitan City and Cheorwon-eup ( Figure 1B, III) and Gimhwa-eup ( Figure 1B, IV), Cheorwon County of Gangwon Province in the ROK during the malaria transmission season from 2010-2012. Mosquito collections were conducted twice a week between 8 PM and 7 AM from April -October, at the same collection sites for comparative analysis. Female mosquitoes were identified using morphological differences and counted regardless of the blood-fed status or malaria parasite infectivity. The average of the number of collected mosquitoes over two days was considered a representative value for the week [14].     Figure 5). The most common occupation was soldier (18.74-31.50%), followed by students (9.73-13.25%), and farmers and fishermen (4.23-6.85%) ( Figure 6).   A total of 110,795 An. sinensis s.l. were collected in Ganghwa County but only 3,963 were collected in Cheorwon County (Figure 7). The high vector density in Ganghwa County may have resulted in the high and increased incidence of vivax malaria in that area.

Discussion
In North Korea, the high-risk areas of vivax malaria are Gaeseong City, the South and North Hwanghae Provinces, and Gangwon Province, which border the DMZ. The DMZ is a 4-km-wide, 250-km-long corridor that extends across the middle part of the Korean peninsula. In general, no civilians have been allowed to enter the DMZ for more than 60 years, even though one exceptional village called "Daesungdong" is located inside of DMZ; therefore, natural ecosystems and biodiversity are highly conserved in the DMZ [18]. To control malaria, North Korea has provided presumptive anti-relapse chemoprophylaxis with primaquine (15 mg for 14 days) before the malaria transmission season for civilians living in these areas since 2002 [19]. As a result, it was believed that transmission of malaria occurred mainly in these areas in North Korea [20]. Therefore, the adjacent to malaria risk areas in North Korea, high risk areas were observed in the ROK, e.g., Incheon Metropolitan City and northern Gyeonggi and Gangwon provinces. There is no objective opinion that mass chemoprophylaxis might largely decrease the number of malaria cases among military personnel, which may alternatively decrease numbers of cases among civilian populations. Chemoprophylaxis started with approximately 16,000 military personnel in 1997 and expanded to more than 200,000 military personnel who are currently serving in malaria high risk areas [13]. Unfortunately, it was found that prophylactic failure occurred even in those who had sufficient plasma concentrations of hydroxychloroquine. It means that a chloroquineresistant P. vivax strain has been reported in the ROK [21]. The appearance of this drug-resistant strain is troublesome for the treatment of patients in the future. Therefore, another standard regimen should be considered to replace the current regimen of chloroquine-primaquine. However, it is not easy to find the next defensive line of anti-malaria drugs because it has been reported that Korean P. vivax isolates are already resistant to pyrimethamine, which is recommended for use in patients with malaria who fail to respond to chloroquine chemotheraphy [22]. It was shown that the existence of antifolate-resistant P. vivax in the ROK. Further detailed geographic mapping of current and changing patterns of vivax malaria drug resistance on a national or regional scale would prove a valuable aid for developing and updating national anti-malarial policy guidelines in the ROK. Control measures and inter-governmental co-operation are also needed to block the spread of drug-resistant malaria in the ROK. It was reported that six Anopheles species in the ROK: Anopheles sinensis sensu stricto, Anopheles lesteri, Anopheles pullus, Anopheles sineroides, Anopheles kleini, and Anopheles belenrae [23][24][25][26]. These six species comprise a species complex called An. sinensis s.l. Because they occur in sympatry and it is hard to distinguish by morphology. Thus, molecular methods are needed to identify the species. But it is hard to apply the newly developed methods for classifications in the surveillance system until developing more precise methods for identification of Anopheles complex in real time manners. Therefore, An. sinensis s.l. was presented in this study. Analysis of the mosquito density of An. sinensis s.l. in study areas over three years showed an extremely higher density  in Ganghwa County compared to Cheorwon County ( Figure 7). Therefore, the potential unknown threatening factors including mosquito density still may exist on the west side of malaria foci compared with the east side; this explains why the API of Gangwon Province, which is the representative location of the east side, was reduced dramatically from 0.123 in 2010 to 0.063 in 2011 and to 0.009 in 2012 and the API of Incheon Metropolitan City changed from 0.093 in 2010 to 0.045 in 2011 but increased to 0.052 in 2012 (Table 3). PHCs are responsible for controlling the transmission of malaria in each of the vivax malaria risk area. As a result of the anti-malaria efforts of the PHC in Cheorwon County, which is a high-risk area in Gangwon Province, the number of cases of malaria decreased dramatically to low level, 27 cases in 2010, 14 cases in 2011, and two cases in 2012. In Cheorwon County, nine cases were first reported in 1997, while there was a peak of 166 cases in 2000 [27]. The interesting finding is that the last peak of each year was shown the similar patterns in three years, that is, increasing the malaria cases in three weeks after Chuseok (Figure 4, ↓). This duration is matched with the incubation period of vivax malaria in the liver. It was probably due to the massive interregional population movement throughout the ROK during Chuseok from high risk areas to malaria free areas, in addition increasing the activities during the night without self protection from malaria vectors. It is suggested that soldiers who leave for vacation to their hometowns should be either checked or treated for malaria just like retiring exsoldiers from their active military service (primaquine/ 15 mg/14 days) [15,28,29].
To evaluate transmission of malaria in a given geographic region, many factors, including temperature, mosquito density, vector capacity, climate, rainfall, and humidity, should be considered [30]. Patient incidence alone cannot provide a complete understanding of the prevalence of malaria in the ROK because there are many factors involved, including the changes in weekly population density of mosquitoes due to environmental factors, vectorial capacity, long-and short-incubation patient ratios, symptomatic and asymptomatic patient ratios, and differences in weekly rainfall and temperatures. This study analysed only two factors: incidence and An. sinensis s.l. However, all factors indicated that transmission of malaria would apparently be reduced except in Incheon Metropolitan City and Gyeongnam Province. There are opportunities for the eradication of malaria in the ROK due to several efforts of the Korea CDC, Government Public Institute of Health and Environment, PHCs, and related institutes.

Conclusions
The number of patients with vivax malaria in the ROK peaked in 2010 for a third time over two decades and then decreased in the subsequent two years. This was the third time the number of cases of malaria peaked since a trend of decreasing cases was observed in 2001. It is expected that the number of cases of malaria will continue to decrease in the future, although there may be some fluctuations. However, more intensive malaria surveillance is needed in the western part of the country where foci have re-emerged to reduce the potential for future increases in the number of malaria cases in the ROK.