Production of the baculovirus-expressed dengue virus glycoprotein NS1 can be improved dramatically with optimised regimes for fed-batch cultures and the addition of the insect moulting hormone, 20-Hydroxyecdysone

A perennial problem in recombinant protein expression is low yield of the product of interest. A strategy which has been shown to increase the production of baculovirus-expressed proteins is to utilise fed-batch cultures. One disadvantage of this approach is the time-consuming task of optimising the feeding strategy. Previously, a statistical optimisation routine was applied to develop a feeding strategy that increased the yield of beta-Galactosidase (beta-Gal) by 2.4-fold (Biotechnol. Bioeng, 59 (1998) 178). This involves the single addition of nutrient concentrates (amino acids, lipids. glucose and yeastolate ultrafiltrate) into Sf9 cell cultures grown in SF900II medium. In this study, it is demonstrated that this optimised fed-batch strategy developed for a high-yielding intracellular product beta-Gal could be applied successfully to a relatively low-yielding glycosylated and secreted product such as the dengue virus glycoprotein NS1. Optimised batch infections yielded 4 mug/ml of NS1 at a peak cell density of 4.2 x 10(6) cells/ml. In contrast. optimised fed-batch infections exhibited a 3-fold improvement in yield, with 12 mug ml of NS1 produced at a peak cell density of 11.3 x 10(6) cells/ml. No further improvements in yield were recorded when the feed volumes were doubled and the peak cell density was increased to 23 x 10(6) cells/ml, unless the cultures were stimulated by the addition of 4 mug/ml of 20-Hydroxyecdysone (an insect moulting hormone). In this case, the NS1 yield was increased to 20 mug/ml. which was nearly 5-fold higher than optimised batch cultures. (C) 2002 Elsevier Science B.V. All rights reserved.

Production of mosquitoes in rainwater tanks and wells on Yorke Island, Torres Strait: Preliminary study

The Torres Strait in northernmost Queensland, Australia, is subject to periodic outbreaks of dengue. A large outbreak of dengue 2 in 1996-97 affected five islands, resulting in 200 confirmed cases. On most of the affected islands, rainwater tanks were a common breeding site for vector mosquitoes. Rainwater tanks, wells and household containers filled with water are the most common breeding sites for dengue mosquitoes (Aedes aegypti), the primary vector of dengue in Queensland. We report on surveys conducted in February 2002 to measure the productivity of rainwater tanks and wells on Yorke Is. (Torres Strait), the first time the productivity of rainwater tanks has been measured in Australia. Of 60 rainwater tanks sampled, 10 had broken screens. Using a sticky emergence trap, 179 adult mosquitoes were collected, consisting of 63 Aedes scutellaris and 116 Culex quinquefasciatus. One unscreened tank produced 177 (99%) of the adults. A plankton net was used to sample 16 wells; 12 positive wells yielded 111 immature (larvae and pupae) mosquitoes, consisting of 57% and 43% Ae. scutellaris and Cx. quinquefasciatus, respectively. The apparent displacement of Ae. aegypti by Ae. scutellaris is discussed. Measures to reduce the likelihood of future dengue outbreaks are recommended.

The over-production of US sports and the new International Division of Cultural Labor

This article focuses on how US professional sports utilize the New International Division of Cultural Labor to supplement an overly costly local labor pool and over-supplied local market. We argue that while the classic problem of over-production is slowly eroding the sealed-off nature of US culture, the forces of its hyper-protectionist capitalism continue to characterize sports, precluding equal exchange.

Production of proteinases by psychrotrophic bacteria in raw milk stored at low temperature

Raw milk samples from two different sources were stored at 2degreesC, 4degreesC and 7degreesC for 10 days and the growth of psychrotrophic bacteria, production of proteinase and proteolysis in the milks were measured during storage. Peptide analyses by the fluorescamine method and RP-HPLC were used in determination of proteolysis and proteinase activity. The average times taken for the psychrotroph counts to reach 10(7) cfu/mL at 2degreesC, 4degreesC and 7degreesC were approximately 9, 7 and 4 days, although there was considerable variation in growth rates in the different milks. There was little correlation between psychrotroph counts and either proteolysis or proteinase activity levels. At 2degreesC, no milk stored showed significant proteolysis by the fluorescamine method after 10 days' storage, but significant proteinase activity could be measured in some of these milks at 8 and 10 days. RP-HPLC analysis was a more sensitive means of detecting peptides than the fluorescamine method.