T. H. Chua

Seasonal and Spatial Dynamics of the Primary Vector of Plasmodium knowlesi within a Major Transmission Focus in Sabah, Malaysia

Background The simian malaria parasite Plasmodium knowlesi is emerging as a public health problem in Southeast Asia, particularly in Malaysian Borneo where it now accounts for the greatest burden of malaria cases and deaths. Control is hindered by limited understanding of the ecology of potential vector species. Methodology/Principal Findings We conducted a one year longitudinal study of P. knowlesi vectors in three sites within an endemic area of Sabah, Malaysia. All mosquitoes were captured using human landing catch. Anopheles mosquitoes were dissected to determine, oocyst, sporozoites and parous rate. Anopheles balabacensis is confirmed as the primary vector of. P. knowlesi (using nested PCR) in Sabah for the first time. Vector densities were significantly higher and more seasonally variable in the village than forest or small scale farming site. However An. balabacensis survival and P. knowlesi infection rates were highest in forest and small scale farm sites. Anopheles balabacensis mostly bites humans outdoors in the early evening between 1800 to 2000hrs. Conclusions/Significance This study indicates transmission is unlikely to be prevented by bednets. This combined with its high vectorial capacity poses a threat to malaria elimination programmes within the region.

POPULATION STUDIES OF BREVICORYNE BRASSICAE (L.), ITS PARASITES AND HYPERPARASITES IN ENGLAND

Studies on populations ofBrevicoryne brassicae (L.), its parasites and hyperparasites were carried out by actual counting in the sprouts field and by sticky and water traps.B. brassicae was found to be attacked by one primary parasite,Diaretiella rapae (McIntosh), which in turn is parasitized byAlloxysta brassicae (Ashm.),Asaphes vulgaris Walker,A. suspensus (Nees),Pachyneuron minutissimum (Förster) andDendrocerus carpenterii (Curtis). The aphid population in the field was started by immigrant alates which were found flying too early to be synchronized with the sprouts plants. SimilarlyD. rapae was not synchronized with the aphids although many individuals could have been carried into a plot through parasitized immigrant alates, of which less than 30% were found parasitized. Because of high hyperparasitism (especially byA. brassicae)D. rapae was not able to maintain a high rate of parasitism to curb the aphid population growth. The maximum percentage mummies being 27.8%, while the maximum, percentage parasitism being 56.6% recorded only during the early 1974 season (mean=12.9%). The decline of aphid population from September onwards was largely due to the cold weather, Syrphid predation and occasionally fungal attack. The high rate of hyperparasitism byA. brassicae is attributed to its better synchronization withD. rapae. The mean percentage of parasite that emerged from mummies collected during 1973–74 wereD. rapae 31.3%,A. brassicae 64.3%,A. vulgaris andA. suspensus 4.3%,D. carpenterii 0.2% andP. minutissimum 0.1%.

A comparative study of the searching efficiencies of a parasite and a hyperparasite

The searching efficiencies of a primary parasite (Diaeretiella rapae (McIntosh)) and a hyperparasite (Alloxysta brassicae (Ash.)) were investigated and compared. In both species, at all parasite densities, there was a curvilinear relationship (P<0.001) between the number of hosts parasitised and the host density. A linear regression (loga=logQ−m logP) was fitted for log area of discovery against log parasite density (P<0.001). The area of discovery for its immediate (i.e. primary) host (viz.Diaeretiella for the hyperparasite and aphid forDiaeretiella) is lower in the hyperparasite than in the primary parasite. InDiaeretialla both the searching efficiency and the mutual interference constant increased (but not significantly,P>0.05) in the presence of its males.

Development of Allele-Specific Single-Nucleotide Polymorphism-Based Polymerase Chain Reaction Markers in Cytochrome Oxidase I for the Differentiation of Bactrocera papayae and Bactrocera carambolae (Diptera: Tephritidae)

ABSTRACT Differentiation of Bactrocera papayae Drew & Hancock and Bactrocera carambolae Drew & Hancock (Diptera: Tephritidae) based on morphological characters has often been problematical. We describe here a single-nucleotide polymorphism (SNP)-based polymerase chain reaction (PCR) assay to differentiate between these two species. For detection of SNPs, fragments derived from each species were amplified using two primer pairs, COIF/COIR and UEA7/UEA10, sequenced, and aligned to obtain a contiguous 1,517-bp segment. Two new sets of primers were designed based on the 11 SNPs identified in the region. Results of the SNP-PCR test using any one of these species-specific primer sets indicate that these two species could be differentiated on basis of presence or absence of a band in the gel profile. We also tested the SNP-PCR primers on Bactrocera umbrosa F., Bactrocera cucurbitae Coquillett, Bactrocera latifrons Hendel, and Bactrocera tau (Walker) but did not detect any band in the gel, indicating the likelihood of a false positive for B. papayae is nil. This SNP-PCR method is efficient and useful, especially for immature life stages or when only adult body parts of the two species are available for identification, as encountered often in quarantine work.

Species determination of Malaysian Bactrocera pests using PCR-RFLP analyses (Diptera: Tephritidae).

BACKGROUND Identification of Bactrocera carambolae Drew and Hancock, B. papayae Drew and Hancock, B. tau Walker, B. latifrons Hendel, B. cucurbitae Coquillett, B. umbrosa Fabricius and B. caudata Fabricius would pose a problem if only a body part or an immature stage were available. Analysis of polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) of cytochrome oxidase I (COI) gene using primers COIR, COIF, UEA7 and UEA10 and restriction enzymes (MseI, RsaI and Alu1) was carried out. The banding profiles in the electrophoresis gel were analysed. RESULTS The COI gene in six Bactrocera spp. was successfully amplified by COIR and COIF, as well as UEA7 and UEA10, while B. caudata was amplified successfully only by UEA primers. Using COI amplified PCR products and restriction enzymes, distinct banding profiles for B. tau, B. latifrons, B. cucurbitae and B. umbrosa were observed, but not for B. carambolae and B. papayae. However, using UEA7, UEA10 and RsaI, B. caudata could be identified, while B. carambolae and B. papayae might possibly be separated from one another. It was also shown that adult body parts or immature life stages of B. carambolae, B. papayae, B. latifrons and B. cucurbitae produced the same banding profiles as the adults. CONCLUSION PCR-RFLP analyses are able to identify positively five Bactrocera species, while B. papayae and B. carambolae might possibly be separated from one another, even if immature life stages or adult body parts are used.

Searching efficiency and multiparasitism in Aphidius smithi and A. ervi (Hym., Aphidiidae), parasites of pea aphid, Acyrthosiphon pisum (Hom., Aphididae)

Aphidius smithi and A. ervi were introduced into the United States to control aphid pests of alfalfa. Laboratory experiments showed that A. smithi was more efficient in searching for hosts (i.e., more hosts parasitized) than A. ervi, when measured separately. However, when the two species searched together for the same hosts, a lesser percentage of A. smithi adults than A. ervi emerged from the mummies. Studies on multiple parasitism which involved hosts parasitized by one species exposed to the other species indicated that larvae of A. ervi were more competitive and survived better to adulthood than larvae of A. smithi. These results might help explain for the decline of A. smithi in California.

Investigating the Contribution of Peri-domestic Transmission to Risk of Zoonotic Malaria Infection in Humans

Background In recent years, the primate malaria Plasmodium knowlesi has emerged in human populations throughout South East Asia, with the largest hotspot being in Sabah, Malaysian Borneo. Control efforts are hindered by limited knowledge of where and when people get exposed to mosquito vectors. It is assumed that exposure occurs primarily when people are working in forest areas, but the role of other potential exposure routes (including domestic or peri-domestic transmission) has not been thoroughly investigated. Methodology/Principal Findings We integrated entomological surveillance within a comprehensive case-control study occurring within a large hotspot of transmission in Sabah, Malaysia. Mosquitoes were collected at 28 pairs households composed of one where an occupant had a confirmed P. knowlesi infection within the preceding 3 weeks (“case”) and an associated “control” where no infection was reported. Human landing catches were conducted to measure the number and diversity of mosquitoes host seeking inside houses and in the surrounding peri-domestic (outdoors but around the household) areas. The predominant malaria vector species was Anopheles balabacensis, most of which were caught outdoors in the early evening (6pm - 9pm). It was significantly more abundant in the peri-domestic area than inside houses (5.5-fold), and also higher at case than control households (0.28±0.194 vs 0.17±0.127, p<0.001). Ten out of 641 An. balabacensis tested were positive for simian malaria parasites, but none for P. knowlesi. Conclusions/Significance This study shows there is a possibility that humans can be exposed to P. knowlesi infection around their homes. The vector is highly exophagic and few were caught indoors indicating interventions using bednets inside households may have relatively little impact.

Variations in carambola infestation rates byBactrocera carambolae drew and hancock (Diptera: Tephritidae) with fruit availability in a carambola orchard

The relationship between the infestation rate of carambola fruits byBactrocera carambolae Drew and Hancock was investigated in a carambola orchard. Phenology of the fruits was found to influence percentage infestation of fruits byB. carambolae. The proportion of unbagged or susceptible fruits infested varied with time of year and tended to decrease with the increase of unbagged fruits available on the tree. The number of ovipunctures per fruit varied with fruit size and was also found to be indicative of the number of adult insects (B. carambolae and parasitoids) that will emerge from each fruit. Higher number of susceptible fruits available on each tree also decreased both the number of ovipunctures per fruit and the number of eggs laid per fruit, which could possibly be due to the strategy of spreading the risk adopted by the femaleB. carambolae or a result of random selection with more hosts available. The main parasitoids recorded wereBiosteres vandenboschi (Fullaway) andB. arisanus (Sonan). The mean percentage of parasitism was 38.3% and it followed roughly that of infestation of fruits.

Nesting Frequency and Clutch Size in Dermochelys coriacea in Malaysia

Analysis of 37,654 leatherback visit records shows that a mean of 81% (range 51-95%) of visits resulted in successful nests every year at Rantau Abang beach, peninsular Malaysia. The internesting interval within a season (intraseasonal interval) varied from 0-49 days, but the highest mode (polymodal histogram) was 9-10 days. The percentage of tagged turtles returning to breed in consecutive years and once in 2 years was 48.9 and 24.6, respectively. Few turtles returned after a long absence (e.g., 4 after 8 years). On average, each turtle returns to breed after an interval of 1.95 years (interseasonal interval). Most turtles (38% of the daily total) arrived at the beach at 0100-0300 hours, and laid at 0200-0400 hours. Mean clutch size for those turtles with only one clutch per season was 83 normal and 20 yolkless eggs. For those with more than one clutch, the mean total clutch of normal and yolkless eggs varied little (range 100-105) although yolkless eggs may be fewer in the later clutches (e.g., 11th and 12th). The major difference was in variation in the maximum and minimum number of normal and yolkless eggs in later clutches. For 3 clutches, the minimum-maximum normal and yolkless egg numbers were 84-85 and 18-22, respectively, while for 12 clutches the corresponding figures were 75-96 and 8-29. The number of yolkless eggs depended on the number of normal eggs laid (y = 47.13 - 0.34x (P < 0.001). Clutch size was also found to vary with the beach sector where the eggs were laid.

Nesting Population and Frequency of Visits in Dermochelys coriacea in Malaysia

A total of 37,654 leatherback turtle visits were recorded over a period of 10 yrs (1967- 1976) on the east coast of peninsular Malaysia. The mean number of visits/yr was 3765 (range 1962-6721), made by 1924 individual turtles (1067-3103), with a mean number of visits/turtle/yr of 1.96 (overall general mean). The mean number of visits/yr made by a turtle tagged in any one yr was 2.89 (overall tagged mean). The number of visits declined somewhat during the last 4 yrs of the program. The turtles visited mostly between May and September, with a peak of 626 individuals and 1278 visits (34% of the mean annual total) recorded in July. The visit (nesting) frequency/mo/turtle increased from April (1.84 visits per month) to June (2.20) and tapered off in September (1.24). The breeding population was estimated to be 15,525 females, the yearly survival rate 0.89, and the proportion of return of tagged turtles 0.31. About 2039 newly matured females were calculated to join the breeding population every year, 65% of which (=1328 individuals) would return to this beach. Maximum use was recorded in the middle part of the nesting beach, and the highest number of nests/km was 886, recorded in 1972. 33.4% of the total tagged turtles (12,997) visited the beach only once (when they were tagged). Those that returned 2, 3, and 4 times constituted 19.9%, 15.6%, and 11.0% of all turtles that returned at least once in subsequent years. The maximum number of visits made by a turtle during the ten yrs was 14, recorded for 6 turtles. The number of visits was found to differ significantly among years, among months, and among sectors of the beach (P < 0.001).