Morphological features and differential counts of Plasmodium knowlesi parasites in naturally acquired human infections

Background: Human infections with Plasmodium knowlesi, a simian malaria parasite, are more common than previously thought. They have been detected by molecular detection methods in various countries in Southeast Asia, where they were initially diagnosed by microscopy mainly as Plasmodium malariae and at times, as Plasmodium falciparum. There is a paucity of information on the morphology of P. knowlesi parasites and proportion of each erythrocytic stage in naturally acquired human infections. Therefore, detailed descriptions of the morphological characteristics and differential counts of the erythrocytic stages of P. knowlesi parasites in human infections were made, photographs were taken, and morphological features were compared with those of P. malariae and P. falciparum. Methods: Thick and thin blood films were made prior to administration of anti-malarial treatment in patients who were subsequently confirmed as having single species knowlesi infections by PCR assays. Giemsa-stained blood films, prepared from 10 randomly selected patients with a parasitaemia ranging from 610 to 236,000 parasites per mu l blood, were examined. Results: The P. knowlesi infection was highly synchronous in only one patient, where 97% of the parasites were at the late trophozoite stage. Early, late and mature trophozoites and schizonts were observed in films from all patients except three; where schizonts and early trophozoites were absent in two and one patient, respectively. Gametocytes were observed in four patients, comprising only between 1.2 to 2.8% of infected erythrocytes. The early trophozoites of P. knowlesi morphologically resemble those of P. falciparum. The late and mature trophozoites, schizonts and gametocytes appear very similar to those of P. malariae. Careful examinations revealed that some minor morphological differences existed between P. knowlesi and P. malariae. These include trophozoites of knowlesi with double chromatin dots and at times with two or three parasites per erythrocyte and mature schizonts of P. knowlesi having 16 merozoites, compared with 12 for P. malariae. Conclusion: Plasmodium knowlesi infections in humans are not highly synchronous. The morphological resemblance of early trophozoites of P. knowlesi to P. falciparum and later erythrocytic stages to P. malariae makes it extremely difficult to identify P. knowlesi infections by microscopy alone.Publisher PDFPeer reviewe

Knowlesi malaria in Vietnam

The simian malaria parasite Plasmodium knowlesi is transmitted in the forests of Southeast Asia. Symptomatic zoonotic knowlesi malaria in humans is widespread in the region and is associated with a history of spending time in the jungle. However, there are many settings where knowlesi transmission to humans would be expected but is not found. A recent report on the Ra-glai population of southern central Vietnam is taken as an example to help explain why this may be so

Polymerase chain reaction of dried blood spots to detect parasite DNA in individuals with lymphatic filariasis

Lymphatic filariasis caused by Brugia malayi has traditionally been detected in the blood of infected individuals by microscopy.Screening for blood-stage microfilaria (mf) by microscopy is labour intensive with user fatigue and poor specimen handling responsible for false negative results. Recently a method to detect the DNA from circulating microfilaria using the polymerase chain reaction(PCR) has been described (Lizotte et al., 1994 ). However,the specimen collection method described was unsuitable for routine screening in field situations.The aim of the study reported here was to adapt the PCR method to a simple blood spot sampling and DNA extraction method suitable for remote areas without compromising the sensitivity of PCR. Blood spots were collected from individuals in Kelantan and Terengganu to optimise the technique. A one tube DNA extraction method was developed and coupled to a nested PCR assay that was field tested on an endemic community in Sabah. There was 100% sensitivity when comparing PCR to microscopy but only 70% sensitivity when comparing microscopy to PCR. The increased sensitiyity of PCR coupled with simple sample collection and DNA extraction provides a valuable alternative to microscopy for detecting B. malayi positive individuals in endemic regions of the world

Evaluating force field accuracy with long-time simulations of a tryptophan zipper peptide

We have combined a custom implementation of the fast multiple-time-stepping LN integrator with parallel tempering to explore folding properties of small peptides in implicit solvent on the time scale of microseconds. We applied this algorithm to the synthetic {\beta}-hairpin trpzip2 and one of its sequence variants W2W9. Each simulation consisted of over 12 {\mu}s of aggregated virtual time. Several measures of folding behavior showed convergence, allowing comparison with experimental equilibrium properties. Our simulations suggest that the electrostatic interaction of tryptophan sidechains is responsible for much of the stability of the native fold. We conclude that the ff99 force field combined with ff96 {\phi} and {\psi} dihedral energies and implicit solvent can reproduce plausible folding behavior in both trpzip2 and W2W9.Comment: 10 pages, 11 figures, submitted to the Journal of Chemical Physics on June 28, 201

Cytoadherence and virulence - the case of Plasmodium knowlesi malaria.

BACKGROUND: Cytoadherence of infected red blood cells to brain endothelium is causally implicated in malarial coma, one of the severe manifestations of falciparum malaria. Cytoadherence is mediated by specific binding of variant parasite antigens, expressed on the surface of infected erythrocytes, to endothelial receptors including, ICAM-1, VCAM and CD36. In fatal cases of severe falciparum malaria with coma, blood vessels in the brain are characteristically congested with infected erythrocytes. Brain sections from a fatal case of knowlesi malaria, but without coma, were similarly congested with infected erythrocytes. The objective of this study was to determine the binding phenotype of Plasmodium knowlesi infected human erythrocytes to recombinant human ICAM-1, VCAM and CD36. METHODS: Five patients with PCR-confirmed P. knowlesi malaria were recruited into the study with consent between April and August 2010. Pre-treatment venous blood was washed and cultured ex vivo to increase the proportion of schizont-infected erythrocytes. Cultured blood was seeded into Petri dishes with triplicate areas coated with ICAM-1, VCAM and CD36. Following incubation at 37°C for one hour the dishes were washed and the number of infected erythrocytes bound/mm2 to PBS control areas and to recombinant human ICAM-1 VCAM and CD36 coated areas were recorded. Each assay was performed in duplicate. Assay performance was monitored with the Plasmodium falciparum clone HB3. RESULTS: Blood samples were cultured ex vivo for up to 14.5 h (mean 11.3 ± 1.9 h) to increase the relative proportion of mature trophozoite and schizont-infected red blood cells to at least 50% (mean 65.8 ± 17.51%). Three (60%) isolates bound significantly to ICAM-1 and VCAM, one (20%) isolate bound to VCAM and none of the five bound significantly to CD36. CONCLUSIONS: Plasmodium knowlesi infected erythrocytes from human subjects bind in a specific but variable manner to the inducible endothelial receptors ICAM-1 and VCAM. Binding to the constitutively-expressed endothelial receptor CD36 was not detected. Further work will be required to define the pathological consequences of these interactions

Micro-geographic risk factors for malarial infection.

BACKGROUND: Knowledge of geography is integral to the study of insect-borne infectious disease such as malaria. This study was designed to evaluate whether geographic parameters are associated with malarial infection in the East Sepik province of Papua New Guinea (PNG), a remote area where malaria is a major cause of morbidity and mortality. METHODS: A global positioning system (GPS) unit was used at each village to collect elevation, latitude and longitude data. Concurrently, a sketch map of each village was generated and the villages were sub-divided into regions of roughly equal populations. Blood samples were taken from subjects in each region using filter paper collection. The samples were later processed using nested PCR for qualitative determination of malarial infection. The area was mapped using the GPS-information and overlaid with prevalence data. Data tables were examined using traditional chi square statistical techniques. A logistic regression analysis was then used to determine the significance of geographic risk factors including, elevation, distance from administrative centre and village of residence. RESULTS: Three hundred and thirty-two samples were included (24% of the total estimated population). Ninety-six were positive, yielding a prevalence of 29%. Chi square testing within each village found a non-random distribution of cases across sub-regions (p < 0.05). Multivariate logistic regression techniques suggested malarial infection changed with elevation (OR = 0.64 per 10 m, p < 0.05) and distance from administrative centre (OR = 1.3 per 100 m, p < 0.05). CONCLUSION: These results suggest that malarial infection is significantly and independently associated with lower elevation and greater distance from administrative centre in a rural area in PNG. This type of analysis can provide information that may be used to target specific areas in developing countries for malaria prevention and treatment

Dipole-dipole interaction between a quantum dot and graphene nanodisk

We study theoretically the dipole-dipole interaction and energy transfer in a hybrid system consisting of a quantum dot and graphene nanodisk embedded in a nonlinear photonic crystal. In our model a probe laser field is applied to measure the energy transfer between the quantum dot and graphene nanodisk while a control field manipulates the energy transfer process. These fields create excitons in the quantum dot and surface plasmon polaritons in the graphene nanodisk which interact via the dipole-dipole interaction. Here the nonlinear photonic crystal acts as a tunable photonic reservoir for the quantum dot, and is used to control the energy transfer. We have found that the spectrum of power absorption in the quantum dot has two peaks due to the creation of two dressed excitons in the presence of the dipole-dipole interaction. The energy transfer rate spectrum of the graphene nanodisk also has two peaks due to the absorption of these two dressed excitons. Additionally, energy transfer between the quantum dot and the graphene nanodisk can be switched on and off by applying a pump laser to the photonic crystal or by adjusting the strength of the dipole-dipole interaction. We show that the intensity and frequencies of the peaks in the energy transfer rate spectra can be modified by changing the number of graphene monolayers in the nanodisk or the separation between the quantum dot and graphene. Our results agree with existing experiments on a qualitative basis. The principle of our system can be employed to fabricate nano-biosensors, optical nano-switches, and energy transfer devices