Elsevier

Experimental Gerontology

Volume 45, Issue 2, February 2010, Pages 158-162
Experimental Gerontology

Short Report
A new cultivation system for studying chemical effects on the lifespan of the fruit fly

https://doi.org/10.1016/j.exger.2009.11.004Get rights and content

Abstract

A side-by-side comparison was made between a conventional vial system and a novel bottle system for cultivating flies and testing the effect of chemical exposure on the lifespan of the flies. While the two cultivation systems yielded very similar results for the effect of DEHP (di[2-ethylhexyl] phthalate) on reducing the lifespan of fruit fly Drosophila melanogaster, the new bottle system has many advantages over the conventional vial system. The bottle system allowed long-term cultivation of flies in the same bottle and thus eliminated the need for transferring of flies between vials. Foods/nutrients were provided as fresh moisture medium coated on a glass slide vertically hanged in the center of the bottle. Fly discharges and dead flies were collected onto a draw horizontally inserted into the bottom of the bottle. These features have resulted in great convenience for cultivating flies and reduced labor and media cost. The effective separation of food from discharge may allow accurate mass balance measurement and thus yield more definitive observations for understanding the actual role of calorie restriction (CR) or dietary-restriction (DR) in fly metabolism and longevity.

Introduction

Flies are frequently used as model organisms for studying physiological and pathological processes affecting lifespan and for understanding the relationship between nutrient metabolism and aging mechanisms (Carey et al., 2002, Fleming et al., 1992, Partridge and Gems, 2007, Piper et al., 2005). Conventionally, flies are cultivated in glass vials which are closed with air-permeable plugs at their top openings. Foods/nutrients are given as a complex medium solidified at the bottom of the vial. To change medium, flies are transferred from vials containing old medium to vials containing fresh medium. This manipulation may introduce opportunities for flies to escape. Whenever that happens, the study result may be jeopardized with some uncertainty as the status and fates of the lost flies are often unknown.

The above problem can be overcome with training of the experimenters performing the transfer or with the anesthesia of the flies before their transfer (Khazaeli et al., 2005). However, the conventional vial cultivation system has another shortcoming that is becoming increasingly significant. The vial cultivation system does not separate intake nutrients from discharge wastes. Thus, it is difficult if not impossible to perform food consumption and waste production measurements in the vial cultivation system. But accurate mass balance may be essential for resolving some controversies surrounding the issues of metabolism and aging (Mair et al., 2005, Molleman et al., 2009, Piper and Partridge, 2007).

For example, many studies have examined the effect of calorie restriction (CR) or dietary-restriction (DR) on longevity (Partridge, 2007, Partridge and Gems, 2007, Partridge et al., 2005a, Partridge et al., 2005b). However, the conclusions of these studies are often based on the differences in the media being provided but not the amount of food actually consumed. Although it is possible that flies may eat all different foods at the same amount and frequency, there are already some studies showing the preference of flies for one type of food over another (Higa and Fuyama, 1993, Nakamura et al., 2008, Nigg et al., 2007). It has also been reported that even slight change in dietary composition affects the result of DR-longevity studies (Zou et al., 2009).

Over the past years, various efforts have been made to overcome the shortcomings of the conventional vial cultivation system. For examples, different sizes and styles of “cages” have been designed and used in various studies (Carey et al., 2008, Flatt et al., 2008, Khazaeli et al., 1995, Pletcher et al., 1998, Promislow and Tatar, 1998). Among which, the square plexiglass cage used by Carey et al. is very interesting because it allows defined quantity of nutrients to be provided (Carey et al., 2008). This was achieved by placing foods and water on top of a slide horizontally inserted into the bottom of the cage. However, this system does not separate food from discharge, a measure essential for defined metabolic study based on accurate mass balance measurements.

A new cultivation system which separates intake food from discharged waste was invented in 2007 and patented in 2008 (Liu, 2008) (Fig. 1). The major advantages claimed for this new cultivation bottle are: (1) it can separate intake from discharge and thus can allow mass balance measurement; (2) it can facilitate the change of medium/food and reduce the chance of losing fly; and (3) it can reduce the wasting of medium/food and allow provision of medium/food in a clean state and defined amount. However, this new invention has not been tested in any real fly cultivation. So its actual performance needs experimental validation.

In this study, we performed side-by-side comparison between the conventional vial system and the new bottle system when used in studying the effect of DEHP (di[2-ethylhexyl] phthalate) on the lifespan of fruit fly Drosophila melanogaster. DEHP is a manufactured chemical commonly added to plastics to make them flexible. It is listed as a probable human carcinogen by the US Environmental Protection Agency (EPA) (ATSDR, 2002) and has been shown with some detrimental effects on the lifespan of fruit fly (Li et al., 2005, Li et al., 2006). This side-by-side evaluation provides some basic results for us to judge the utility of the new cultivation system and showing its potential for its future use in advanced studies on the relationship between metabolism and aging processes.

Section snippets

Reagents and medium preparation

The standard medium was made of the following components and amounts: distilled water 1000 ml; white sandy sugar 65 g; agar 6 g; corn powder (85 g); yeast powder 7.5 g; acetic acid 7 ml. Sandy sugar and agar powder were mixed by stirring while being boiled with water in a pot. After their complete dissolution into a paste, corn powder already made into paste in another pot was added into the sugar–agar pot and mixed with stirring while being continuously heated. Then the paste of yeast powder was

General observations on fruit flies living inside conventional and new bottles

Compared with the widely used fly cultivation vials, the new bottle (Fig. 2) had about 2.6-fold more internal volume so that there is more freedom for active movement by the fruit flies. For the same experiment, the use of nutrient medium by the bottle system was reduced at least 10-folds because only a thin layer of medium was needed on the slide and fresh medium can be provided more at more frequently intervals than the vial system which received thick layer of medium each time of vial change.

Discussion

From our hands-on experience with this side-by-side comparison between a conventional vial system and the new bottle system for fly cultivation, we felt that the newly invented system is much convenient to use and also labor-saving and even cost-saving. The convenience of the bottle system for long-term fly cultivation is afforded by its intrinsic advantages of allowing changing food/nutrient supplies and collecting waste discharge and dead body without any need of transferring live flies. This

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