PRIMARY CELL CULTURE OF AEDES ALBOPICTUS MIDGUT CELLS: A PROSPECTIVE MODEL FOR IN VITRO STUDY OF ARBOVIRUSES

  Objective: Midgut cells play a key role in the propagation of mosquito borne Arboviruses. The existing mosquito cell lines for studying viral pathogenesis are derived either from larvae or from eggs since there is no cell line available from the mosquito midgut. Therefore, to delineate the in situ viral interaction which naturally occurs within the mosquito midgut and represent cellular pathogenesis in human beings, the present work was aimed to develop a primary cell line from the midgut cells of Aedes albopictus.Methods: The midgut cells of A. albopictus were collected, cultured and incubated at 28°C to study the growth after every 24 hrs for 7 days.Result: The primary cell culture showed an increasing growth pattern of columnar cells up to 48 hrs followed by decrease in cell population afterward. However, the number of stem cells increased significantly throughout the study period, and their population outnumbered the columnar cells after 72 hrs. There was no significant change of goblet cells and regenerating cells which were scanty in number throughout the experiment.Conclusion: The present method will help to develop the individual cell lines from mosquito midgut and study the host pathogen interaction in arboviral diseases in future

An observation on direct changes in Aedes albopictus midgut cells by Rhus tox 6C in relation to dengue virus infection

Background and Objectives: In mosquito vectors, dengue virus (DENV) invasion occurs through midgut cells, but available mosquito cell lines for in vitro study of DENV are prepared from eggs or larvae, which are not appropriate models, to study its infectivity. Hence, we developed a new primary cell culture, from Aedes albopictus mosquito midgut, and standardized it for in vitro study of DENV, with an aim to find out any possible role of homoeopathic medicines, in preventing or reducing DENV invasiveness in these midgut cells. This midgut primary cell culture demonstrated prominent cytopathic effects on infection with wild DENV isolated from dengue-infected patients in viremic phase. Materials and Methods: In this paper, we observed the direct effect of homoeopathic medicine Rhus toxicodendron 6C (Rhus tox 6C) (ultra dilution of 10−12 ) on this primary cell culture, to find out significant changes, to be used as baseline data in future experiments to observe possible role of Rhus tox 6C against DENV infection in these cells. Hence, these direct changes may be a prerequisite for the action of this medicine against DENV invasion; as this is one of common repertoire homoeopathic medicines used against dengue fever. Conclusion and Discussion: In our experiments, we found that Rhus tox 6C could increase cell size and help organization of cells on the solid surface as observed under scanning electron microscope although the total number of cells was decreased. Moreover, Rhus tox 6C treated cells were healthier as indicated by less number of deformed, clump, and diploform cells

Changes in viral load in different organs of Japanese Encephalitis virus-infected chick embryo under the influence of Belladonna 200C

Background: Japanese encephalitis(JE) is highly prevalent in many states of India. Belladonna 200C is widely used in the prevention and treatment of JE.The effect of Belladonna 200C in virus replication inside different tissues has not been studied. Objective: To study the effect of Belladonna 200C in virus replication inside different tissues utilising chick embryo model. Materials and Methods: Twelve-day-old fertilised eggs of Black Australorp were inoculated with JE via chorioallantoic membrane (CAM) route in different experimental sets: infection, Belladonna 200C treated and vehicle control, keeping matched blank sets. All experimental sets were incubated for 48 hours. After incubation, viable eggs were sacrificed humanly and different tissues were observed and collected for viral load determination by real-time-polymerase chain reaction (PCR). Results: The control group showed visible pocks over the CAM; brains were liquefied due to haemorrhagic liquefactive necrosis and white patches were found over the liver. However, the medicine-treated group was apparently normal; there were no visible changes in the brain and the liver was healthy like control. Real-time-PCR results showed high viral load in CAM and brain with absence of viral RNA in liver of the virus-infected group. Pre-treatment with Belladonna 200C significantly reduced the overall load (P < 0.05) in CAM and brain which correlated with the morbid pathological changes of the organs. Conclusion: Although Belladonna 200C did not completely inhibit JE viral replication in the brain, it reduced the severity of JE by diminishing the viral loads in this tissue