Impacts of Agricultural Practices on Insecticide Resistance in the Malaria Vector Anopheles arabiensis in Khartoum State,Sudan

Background

Agricultural pesticides may play a profound role in selection of resistance in field populations of mosquito vectors. The objective of this study is to investigate possible links between agricultural pesticide use and development of resistance to insecticides by the major malaria vector Anopheles arabiensis in northern Sudan.

Methodology/Principal Findings

Entomological surveys were conducted during two agricultural seasons in six urban and peri-urban sites in Khartoum state. Agro-sociological data were collected from 240 farmers subjected to semi-structured questionnaires based on knowledge attitude and practice (KAP) surveys. Susceptibility status of An. arabiensis (n=6000) was assessed in all sites and during each season using WHO bioassay tests to DDT, deltamethrin, permethrin, Malathion and bendiocarb. KAP analysis revealed that pesticide application was common practice among both urban and peri-urban farmers, with organophosphates and carbamates most commonly used. Selection for resistance is likely to be greater in peri-urban sites where farmers apply pesticide more frequently and are less likely to dispose of surpluses correctly. Though variable among insecticides and seasons, broad-spectrum mortality was slightly, but significantly higher in urban than peri-urban sites and most marked for bendiocarb, to which susceptibility was lowest. Anopheles arabiensis from all sites showed evidence of resistance or suspected resistance, especially pyrethroids. However, low-moderate frequencies of the L1014F kdr allele in all sites, which was very strongly associated with DDT, permethrin and deltamethrin survivorship (OR=6.14-14.67) suggests that resistance could increase rapidly.

Conclusions

Ubiquitous multiple-resistance coupled with presence of a clear mechanism for DDT and pyrethroids (kdr L1014F) in populations of An. arabiensis from Khartoum-Sudan suggests careful insecticide management is essential to prolong efficacy. Our findings are consistent with agricultural insecticide use as a source of selection for resistance and argue for coordination between the integrated vector control program and the Ministry of Agriculture to permit successful implementation of rational resistance management strategies.     

Pharmaceutical and Pharmacokinetic Evaluation of a Novel Fast Dissolving Film Formulation of Flupentixol Dihydrochloride

The objective of the present study was to develop fast dissolving oral film of the antipsychotic drug, flupentixol dihydrochloride, to enhance its bioavailability, optimize its therapeutic effect when used to treat depression with anxiety, and increase the convenience and compliance by the mentally ill, developmentally disable, elderly, and pediatric patients. Six formulae were prepared with different concentrations of water-soluble polymers vis. hydroxypropyl methylcellulose (HPMC E5) and carboxymethyl cellulose (CMC) by solvent casting technique. The prepared films were subjected to characterization for folding endurance, weight variations, thickness, disintegration time, drug release pattern, and drug content. Physical compatibility between the drug and excipients was guaranteed in the selected formulation (2% HPMC) by means of differential scanning calorimetry analysis and Fourier-transform infrared spectroscopy. This formulation revealed high stability after testing according to the International Conference on Harmonisation guidelines. In vivo studies based on single phase parallel design were carried out for the optimized formulation in healthy human volunteers. The concentration of flupentixol dihydrochloride in plasma samples was analyzed by a developed validated LC-MS/MS assay method and the pharmacokinetic parameters of the established formulation were compared with the commercially available oral tablets. Faster rate of absorption of flupentixol could be obtained from the oral film formulation and the relative bioavailability was found to be 151.06% compared to the marketed product.KEY WORDS: fast dissolving, flupentixol dihydrochloride, LC-MS/MS analysis, oral film, pharmacokinetics     

An interplay between 2 signaling pathways: Melatonin-cAMP and IP3–Ca signaling pathways control intraerythrocytic development of the malaria parasite Plasmodium falciparum

Plasmodium falciparum spends most of its asexual life cycle within human erythrocytes, where proliferation and maturation occur. Development into the mature forms of P. falciparum causes severe symptoms due to its distinctive sequestration capability. However, the physiological roles and the molecular mechanisms of signaling pathways that govern development are poorly understood. Our previous study showed that P. falciparum exhibits stage-specific spontaneous Calcium (Ca²⁺) oscillations in ring and early trophozoites, and the latter was essential for parasite development. In this study, we show that luzindole (LZ), a selective melatonin receptor antagonist, inhibits parasite growth. Analyses of development and morphology of LZ-treated P. falciparum revealed that LZ severely disrupted intraerythrocytic maturation, resulting in parasite death. When LZ was added at ring stage, the parasite could not undergo further development, whereas LZ added at the trophozoite stage inhibited development from early into late schizonts. Live-cell Ca²⁺ imaging showed that LZ treatment completely abolished Ca²⁺ oscillation in the ring forms while having little effect on early trophozoites. Further, the melatonin-induced cAMP increase observed at ring and late trophozoite stage was attenuated by LZ treatment. These suggest that a complex interplay between IP₃–Ca²⁺ and cAMP signaling pathways is involved in intraerythrocytic development of P. falciparum.     

Design,synthesis, anticancer evaluation,and molecular modelling studies of novel tolmetin derivatives as potential VEGFR-2 inhibitors and apoptosis inducers

Novel tolmetin derivatives 5a–f to 8a–c were designed, synthesised, and evaluated for antiproliferative activity by NCI (USA) against a panel of 60 tumour cell lines. The cytotoxic activity of the most active tolmetin derivatives 5b and 5c was examined against HL-60, HCT-15, and UO-31 tumour cell lines. Compound 5b was found to be the most potent derivative against HL-60, HCT-15, and UO-31 cell lines with IC₅₀ values of 10.32 ± 0.55, 6.62 ± 0.35, and 7.69 ± 0.41 µM, respectively. Molecular modelling studies of derivative 5b towards the VEGFR-2 active site were performed. Compound 5b displayed high inhibitory activity against VEGFR-2 (IC₅₀ = 0.20 µM). It extremely reduced the HUVECs migration potential exhibiting deeply reduced wound healing patterns after 72 h. It induced apoptosis in HCT-15 cells (52.72-fold). This evidence was supported by an increase in the level of apoptotic caspases-3, -8, and -9 by 7.808-, 1.867-, and 7.622-fold, respectively. Compound 5b arrested the cell cycle in the G0/G1 phase. Furthermore, the ADME studies showed that compound 5b possessed promising pharmacokinetic properties.     

Development of liver microtissues with functional biliary ductular network

Liver tissue engineering aims to create transplantable liver grafts that can serve as substitutes for donor's livers. One major challenge in creating a fully functional liver tissue has been to recreate the biliary drainage in an engineered liver construct through integration of bile canaliculi (BC) with the biliary ductular network that would enable the clearance of bile from the hepatocytes to the host duodenum. In this study, we show the formation of such a hepatic microtissue by coculturing rat primary hepatocytes with cholangiocytes and stromal cells. Our results indicate that within the spheroids, hepatocytes maintained viability and function for up to 7 days. Viable hepatocytes became polarized by forming BC with the presence of tight junctions. Morphologically, hepatocytes formed the core of the spheroids, while cholangiocytes resided at the periphery forming a monolayer microcysts and tubular structures extending outward. The spheroids were subsequently cultured in clusters to create a higher order ductular network resembling hepatic lobule. The cholangiocytes formed functional biliary ductular channels in between hepatic spheroids that were able to collect, transport, and secrete bile. Our results constitute the first step to recreate hepatic building blocks with biliary drainage for repopulating the whole liver scaffolds to be used as transplantable liver grafts.     

Metabolic engineering of Propionibacterium freudenreichii for n-propanol production

Propionibacteria are widely used in industry for manufacturing of Swiss cheese, vitamin B₁₂, and propionic acid. However, little is known about their genetics and only a few reports are available on the metabolic engineering of propionibacteria aiming at enhancing fermentative production of vitamin B₁₂ and propionic acid. n-Propanol is a common solvent, an intermediate in many industrial applications, and a promising biofuel. To date, no wild-type microorganism is known to produce n-propanol in sufficient quantities for industrial application purposes. In this study, a bifunctional aldehyde/alcohol dehydrogenase (adhE) was cloned from Escherichia coli and expressed in Propionibacterium freudenreichii. The mutants expressing the adhE gene converted propionyl- coenzyme A, which is the precursor for propionic acid biosynthesis, to n-propanol. The production of n-propanol was limited by NADH availability, which was improved significantly by using glycerol as the carbon source. Interestingly, the improved propanol production was accompanied by a significant increase in propionic acid productivity, indicating a positive effect of n-propanol biosynthesis on propionic acid fermentative production. To our best knowledge, this is the first report on producing n-propanol by metabolically engineered propionibacteria, which offers a novel route to produce n-propanol from renewable feedstock, and possibly a new way to boost propionic acid fermentation.     

Image cytometric analysis of p53 and mdm-2 expression in primary and recurrent mucoepidermoid carcinoma of parotid gland: immunohistochemical study

Aims and Objectives

This study aims to analyze immunocytochemically p53 aberrant expression and mdm-2 expression in primary and recurrent mucoepidermoid carcinoma (MEC) of parotid gland and to ascertain if expression of these markers correlates with tumor behavior, clinical outcome, histological grade and local recurrence.

Methods

20 cases histologically diagnosed as primary MEC with different grades were included in the study. Out of 20 cases, 7 were classified as grade I, 8 as grade II and 5 as grade III. Immunohistochemical staining of these 20 primary cases as well as 6 recurrent cases with anti-p53 and anti-mdm-2 antibodies was carried out. Area fraction of immunopositivity was estimated by image analysis software.

Results

16/20 primary cases were p53 +ve (80%). The p53 positive cases included 3 cases classified as grade (I), 8 cases as grade (II) and 5 cases as grade (III). All 6 recurrent cases were p53 +ve. On the other hand, 14/20 primary and only 2/6 recurrent cases were mdm-2 +ve. The mdm-2 +ve primary cases included 2 classified as grade (I), 7 as grade (II) and 5 as grade (III). 12 primary MEC showed co-expression of both p53 and mdm-2 of which 2 cases showed local recurrence.

Conclusions

these data suggested that expression of p53 and mdm-2 in primary and recurrent MEC correlates with the high histological grade. P53 aberrant expression is not only considered as an early event in MEC carcinogenesis but also correlates to tumor behavior and local recurrence. Mdm-2 overexpression is correlated to pathogenesis of MEC. However, no strong evidence was found between mdm-2 expression and MEC local recurrence.