Functional analysis of <Emphasis Type="Italic">Plasmodium falciparum</Emphasis> subpopulations associated with artemisinin resistance in Cambodia

Background

Plasmodium falciparum malaria is one of the most widespread parasitic infections in humans and remains a leading global health concern. Malaria elimination efforts are threatened by the emergence and spread of resistance to artemisinin-based combination therapy, the first-line treatment of malaria. Promising molecular markers and pathways associated with artemisinin drug resistance have been identified, but the underlying molecular mechanisms of resistance remains unknown. The genomic data from early period of emergence of artemisinin resistance (2008–2011) was evaluated, with aim to define k13 associated genetic background in Cambodia, the country identified as epicentre of anti-malarial drug resistance, through characterization of 167 parasite isolates using a panel of 21,257 SNPs.

Results

Eight subpopulations were identified suggesting a process of acquisition of artemisinin resistance consistent with an emergence-selection-diffusion model, supported by the shifting balance theory. Identification of population specific mutations facilitated the characterization of a core set of 57 background genes associated with artemisinin resistance and associated pathways. The analysis indicates that the background of artemisinin resistance was not acquired after drug pressure, rather is the result of fixation followed by selection on the daughter subpopulations derived from the ancestral population.

Conclusions

Functional analysis of artemisinin resistance subpopulations illustrates the strong interplay between ubiquitination and cell division or differentiation in artemisinin resistant parasites. The relationship of these pathways with the P. falciparum resistant subpopulation and presence of drug resistance markers in addition to k13, highlights the major role of admixed parasite population in the diffusion of artemisinin resistant background. The diffusion of resistant genes in the Cambodian admixed population after selection resulted from mating of gametocytes of sensitive and resistant parasite populations.

     

A hybrid of bees algorithm and flux balance analysis with OptKnock as a platform for in silico optimization of microbial strains

Microbial strain optimization focuses on improving technological properties of the strain of microorganisms. However, the complexities of the metabolic networks, which lead to data ambiguity, often cause genetic modification on the desirable phenotypes difficult to predict. Furthermore, vast number of reactions in cellular metabolism lead to the combinatorial problem in obtaining optimal gene deletion strategy. Consequently, the computation time increases exponentially with the increase in the size of the problem. Hence, we propose an extension of a hybrid of Bees Algorithm and Flux Balance Analysis (BAFBA) by integrating OptKnock into BAFBA to validate the result. This paper presents a number of computational experiments to test on the performance and capability of BAFBA. Escherichia coli, Bacillus subtilis and Clostridium thermocellum are the model organisms in this paper. Also included is the identification of potential reactions to improve the production of succinic acid, lactic acid and ethanol, plus the discussion on the changes in the flux distribution of the predicted mutants. BAFBA shows potential in suggesting the non-intuitive gene knockout strategies and a low variability among the several runs. The results show that BAFBA is suitable, reliable and applicable in predicting optimal gene knockout strategy.     

An evaluation of the method used in digestibility estimations of a dietary ingredient and comparisons on external and internal markers, and time of faeces collection in digestibility studies in the fish Oreochromis aureus (Steindachner)

Results of experiments carried out on the tilapiine fish Oreochromis aureus (Steindachner) to evaluate the method used in the determination of the apparent dry matter and nutrient digestibilities of an ingredient, together with validity of estimations based on day and night collection of faeces, and internal and external markers, are presented. The apparent dry matter and protein digestibilities of the test diets were linearly correlated to the percent substitution of the test ingredient, percent fibre and ash in the test diets (P less than 0.05); the correlation coefficient for the individual relationships decreased in that order. Dry matter or protein digestibilities were not correlated to the dietary protein content (P greater than 0.05). Similarly there were no statistical differences between digestibility estimations based on faecal material, voided in the day or the night; and estimated using different markers viz crude fibre and Cr2O3. The apparent dry matter and protein digestibilities of the test ingredients, leaf meal, were curvilinearly related to the percent substitution of the ingredient in the test diets. The ingredient digestibilities estimated, using crude fibre as the marker, were consistently higher than those estimated with Cr2O3 (P less than 0.05). The present study suggests that, for ingredient digestibility estimations, test diets prepared by mixing 15-20% of the ingredient to a reference diet would be more desirable.     

三种裸趾虎核型及银带的研究

裸趾虎属Ｃｙｒｔｏｄａｃｔｙｌｕｓ动物 ,已知有 6 0余种 (Ｗｅｒ ｍｕｔｈ ,196 5 ) ,分布于亚州南部及西南部、欧州及大洋洲。中国记载有 5种 (Ｚｈａｏ等 ,1993) ,分布于西北部的干旱荒漠区及西藏东南部。有关其染色体研究报道甚少 ,仅国外记载 7种 (Ｓｚｃｚｅｒｂａｋ等 ,1996 ) ,且所提供的数据较为粗略。为此 ,本文对长裸趾虎Ｃ ｅｌｏｎｇａｔｕｓ、墨脱裸趾虎Ｃ ｍｅｄｏｇｅｎｓｉｓ和西藏裸趾虎Ｃ ｔｉｂｅｔａｎｕｓ核型进行研究 ,并首次报道其银带。旨在为分类及系统进化学提供细胞遗传学信息。1　材料和方法实验动物见表 1…     

Cambodian patients' and health professionals' views regarding the allocation of antiretroviral drugs

The way Cambodian patients and health professionals judge the priority of HIV-infected patients in relation to the allocation of antiretroviral drugs was examined. Participants were either HIV-infected patients attending the HIV/AIDS Care and Support Centre for People Living with HIV/AIDS in Phnom Penh (29 females and 21 males) or members of the staff (9 physicians, 6 pharmacists and 15 health counsellors and health educators). They were presented with stories of a few lines depicting a patient's situation and were instructed to judge the extent to which the patient should be given priority for HIV drugs. The stories were composed according to a four within-subject factor design: (a) the patient's family responsibilities, (b) the severity of infection, (c) the time elapsed since the first consultation, and (d) the financial difficulties of the family. Most patients expressed the view that the drugs should be used for the patients who are most important from a familial point of view, namely, when the family contains small children and/or is already in a precarious financial condition.     

Theoretical and functional analysis of the SIV fusion peptide.

The fusion domain of simian immunodeficiency virus (SIV) envelope glycoproteins is a hydrophobic region located at the amino-terminal extremity of the transmembrane protein (gp32). Assuming an alpha helical structure for the SIV fusogenic domain of gp32 in a lipid environment, theoretical studies have predicted that the fusion peptide would insert obliquely in the lipid bilayer. This oblique insertion could be an initial step of the fusion process by disorganizing locally the structure of the lipid bilayer. We have tested this hypothesis by selectively mutagenizing the SIV gp160 expressed via a vaccinia virus vector, to alter the theoretical angle of insertion of the fusion peptide. The fusogenic activity of the wild-type and mutant glycoproteins was tested after infection of T4 lymphocytic cell lines by the recombinant vaccinia virus, and measure of syncytia formation. Mutations that modified the oblique orientation reduced the fusogenic activity. In contrast, mutations that conserve the oblique orientation did not alter the fusogenic properties. Our results support the hypothesis that oblique orientation is important for fusogenic activity.     

Prediction of macrozoobenthic species distribution in the Korean Saemangeum tidal flat based on a logistic regression model of environmental parameters

This study aims to contribute to the development of heuristic statistical models, which are able to predict benthic macrofaunal responses to environmental gradients in coastal areas, such as tidal flats. Ecological response surfaces were derived for 15 intertidal macrobenthic species, using logistic regression based on three separate environmental parameters (shore level, mud content, and organic content) measured on the tidal flats of the Saemangeum estuary, Korea. The presence/absence of the 15 intertidal macrofauna was accurately predicted from each separate environmental factor in the environmental models, with a prediction accuracy of ~65–92%. Subsequently, geographical comparison was made between the mapped probability surfaces and maps of observed species occurrence. The results indicated that the models developed for different species exhibited a wide variety of functional forms, highlighting potential variability in species response to changes in habitat conditions, even for closely associated species with a similar trophic type. Our modeling approach was capable of predicting macrobenthic species distributions with a relatively high degree of accuracy, although the ecological processes controlling intertidal macrobenthic distribution could not be fully determined. Overall, good agreement between modeling results and field observations, with relatively high concordance regardless of target species, emphasized that such an approach would be of practical use in terms of ecosystem approach to tidal flat management.     

Lead uptake and potentiometric titration studies with live and dried cells of Rhodotorula glutinis

The ability of live cells (LC), freeze dried cells (FDC) and oven dried cells (ODC) of the yeast Rhodotorula glutinis to remove lead from aqueous solution has been studied. Discernible differences were found between the biosorption properties of LC and the other two types of cell preparation. The LC preparation exhibited an uptake level of about 12 mg g⁻¹ in a batch contactor with a biomass dosage of 2 g l⁻¹ and an initial lead concentration of 100 mg l⁻¹. This compared with, respectively, about 26 and 30 mg g⁻¹ for the FDC and ODC biosorbents under the same experimental conditions. It is seen that the level of lead uptake by the two latter biosorbents was increased to, respectively, 2.2- and 2.5-fold of the level observed for the LC preparation. The superior performance of the FDC and ODC biosorbents in the lead binding process was attributed to the presence of additional binding sites on their cell wall surfaces as indicated by potentiometric titration data. These binding sites were ascribed to carboxylic and phosphoric groups, which are the primary sites of divalent metal complexation. Modeling of the titration data revealed that subjecting R. glutinis biomass to freeze drying or oven drying increased its proton binding site concentration by a factor of 3. It appears that the two simple physical treatments were able to compromise the R. glutinis cell wall structure in such a way as to make sites normally inaccessible to become active in proton and lead binding.     

Differential Bees Flux Balance Analysis with OptKnock for In Silico Microbial Strains Optimization

Microbial strains optimization for the overproduction of desired phenotype has been a popular topic in recent years. The strains can be optimized through several techniques in the field of genetic engineering. Gene knockout is a genetic engineering technique that can engineer the metabolism of microbial cells with the objective to obtain desirable phenotypes. However, the complexities of the metabolic networks have made the process to identify the effects of genetic modification on the desirable phenotypes challenging. Furthermore, a vast number of reactions in cellular metabolism often lead to the combinatorial problem in obtaining optimal gene deletion strategy. Basically, the size of a genome-scale metabolic model is usually large. As the size of the problem increases, the computation time increases exponentially. In this paper, we propose Differential Bees Flux Balance Analysis (DBFBA) with OptKnock to identify optimal gene knockout strategies for maximizing the production yield of desired phenotypes while sustaining the growth rate. This proposed method functions by improving the performance of a hybrid of Bees Algorithm and Flux Balance Analysis (BAFBA) by hybridizing Differential Evolution (DE) algorithm into neighborhood searching strategy of BAFBA. In addition, DBFBA is integrated with OptKnock to validate the results for improving the reliability the work. Through several experiments conducted on Escherichia coli, Bacillus subtilis, and Clostridium thermocellum as the model organisms, DBFBA has shown a better performance in terms of computational time, stability, growth rate, and production yield of desired phenotypes compared to the methods used in previous works.