A High-Throughput Method For Zebrafish Sperm Cryopreservation and In Vitro Fertilization

This is a method for zebrafish sperm cryopreservation that is an adaptation of the Harvey method (Harvey et al., 1982). We have introduced two changes to the original protocol that both streamline the procedure and increase sample uniformity. First, we normalize all sperm volumes using freezing media that does not contain the cryoprotectant. Second, cryopreserved sperm are stored in cryovials instead of capillary tubes. The rates of sperm freezing and thawing (δ°C/time) are probably the two most critical variables to control in this procedure. For this reason, do not substitute different tubes for those specified. Working in teams of 2 it is possible to freeze the sperm of 100 males per team in ~2 hrs. Sperm cryopreserved using this protocol has an average of 25% fertility (measured as the number of viable embryos generated in an in vitro fertilization divided by the total number of eggs fertilized) and this percent fertility is stable over many years

Ciliate predation by nematodes in tidal flat sediments

The present study investigates the possibility of a trophic link between ciliates and nematodes in fine sandy sediments of the Molenplaat intertidal flat (Schelde estuary, SW Netherlands). Grazing experiments were conducted under controlled laboratory conditions, with ciliate species isolated from enrichment cultures and nematodes directly collected from the field. Significant reductions in ciliate numbers were found in the presence of the predatory nematode Enoploides longispiculosus, a prominent species (and genus) in fine to medium sandy sediments of the North Sea and adjacent estuaries. No such effects were found when ciliates were inoculated with a mix of mainly deposit-feeding nematodes from the same sampling site. Based on these results, ciliate predation by E. longispiculosus was tested for several benthic ciliate species and abundances, at a range of predator abundances and temperatures, and in the presence of alternative prey (in casu nematodes). E. longispiculosus was capable of significantly reducing densities of 5 out of 6 ciliate species offered as prey. Depending on the experimental conditions and the prey species, predation rates ranged from 0.19 to 10.8 ciliates predator-1 hour-1, corresponding to a biomass consumption of 0.001-0.33 µg C predator-1 day-1. An overall positive relation between available ciliate biomass and predation rate was found. Comparison of experimental data with field conditions suggests that a considerable part of the ciliate production in fine sandy sediments of the Molenplaat is likely to be consumed by E. longispiculosus, which largely dominates meiofaunal biomass there. Estimated carbon requirements for the predator and production estimates of ciliate and nematode prey at the study site, strongly suggest that ciliates are probably a far more important carbon source for E. longispiculosus than nematode prey, at least between late spring and autumn. This implies that carbon transfer from primary producers and bacteria to predatory nematodes may to a large extent be mediated through the microbial food web. In view of the generally high densities/biomasses of ciliates as well as predacious nematodes in fine sandy sediments, similar patterns are to be expected in many other estuarine and marine sediments

Trophic interactions between ciliates and nematodes from an intertidal flat

The present study investigated the possibility of a trophic link between ciliates and nematodes in fine sandy sediments of the Molenplaat intertidal flat (Schelde estuary, SW Netherlands). Grazing experiments were conducted under controlled laboratory conditions, with ciliate species isolated from enrichment cultures and nematodes collected directly from the field. Significant reductions in ciliate numbers were found in the presence of the predatory nematode Enoploides longispiculosus, a prominent species (and genus) in fine to medium sandy sediments of the North Sea and adjacent estuaries. No such effects were found when ciliates were inoculated with a mix of mainly deposit-feeding nematodes from the same sampling site. On the basis of these results, ciliate predation by E. longispiculosus was tested for several benthic ciliate species and abundances, at a range of predator abundances and temperatures, and in the presence of alternative prey (in casu nematodes). E. longispiculosus significantly reduced the densities of 5 out of 6 ciliate species offered as prey. Depending on the experimental conditions and the prey species, predation rates ranged from 0.19 to 10.8 ciliates predator-1 h-1, corresponding to a biomass consumption of 0.001 to 0.33 µg C predator-1 d-1. An overall positive relation between available ciliate biomass and predation rate was found. Comparison of experimental data with field conditions suggests that a considerable part of the ciliate production in fine sandy sediments of the Molenplaat is likely to be consumed by E. longispiculosus, which largely dominates meiofaunal biomass there. Estimated carbon requirements for the predator and production estimates of ciliate and nematode prey at the study site strongly suggest that ciliates are probably a far more important carbon source for E. longispiculosus than nematode prey, at least between late spring and autumn. This implies that carbon transfer from primary producers and bacteria to predatory nematodes may to a large extent be mediated through the microbial food web. In view of the generally high densities and biomasses of ciliates as well as predacious nematodes in fine sandy sediments, similar patterns are to be expected in many other estuarine and marine sediments

Migrating medical communications software to a multi-tenant cloud environment

The rise of cloud computing has paved the way for many new applications. Many of these new cloud applications are also multi-tenant, ensuring multiple end users can make use of the same application instance. While these technologies make it possible to create many new applications, many legacy applications can also benefit from the added flexibility and cost-savings of cloud computing and multi-tenancy. In this paper, we describe the steps required to migrate a. NET-based medical communications application to the Windows Azure public cloud environment, and the steps required to add multi-tenancy to the application. We then discuss the advantages and disadvantages of our migration approach. We found that the migration to the cloud itself requires only a limited amount of changes to the application, but that this also limited the benefits, as individual instances would only be partially used. Adding multi-tenancy requires more changes, but when this is done, it has the potential to greatly reduce the cost of running the application

Anthelmintic tolerance in free-living and facultative parasitic isolates of Halicephalobus (Panagrolaimidae)

Studies on anthelmintic resistance in equine parasites do not include facultative parasites. Halicephalobus gingivalis is a free-living bacteriovorous nematode and a known facultative parasite of horses with a strong indication of some form of tolerance to common anthelmintic drugs. This research presents the results of an in vitro study on the anthelmintic tolerance of several isolates of Halicephalobus to thiabendazole and ivermectin using an adaptation of the Micro-Agar Larval Development Test hereby focusing on egg hatching and larval development. Panagrellus redivivus and Panagrolaimus superbus were included as a positive control. The results generally show that the anthelmintic tolerance of Halicephalobus to both thiabendazole and ivermectin was considerably higher than that of the closely related Panagrolaimidae and, comparing to other studies, than that of obligatory equine parasites. Our results further reveal a remarkable trend of increasing tolerance from fully free-living isolates towards horse-associated isolates. In vitro anthelmintic testing with free-living and facultative parasitic nematodes offers the advantage of observing drug effect on the complete lifecycle as opposed to obligatory parasites which can only be followed until the third larval stage. We therefore propose Halicephalobus gingivalis as an experimental tool to deepen our understanding of the biology of anthelmintic tolerance