Immunosuppression in rabies virus infection mediated by lymphocyte apoptosis.

Previously, we demonstrated a depression of cell-mediated immunity in mice by street rabies virus infection. In the present study, we investigated several events during the course of infection and looked for alterations in the host lymphoid cells for evidence of apoptosis. Total cellular RNA was extracted from muscle tissues at the inoculation site of peripherally infected mice at different intervals after infection. Rabies virus mRNA was monitored by reverse transcription-PCR. The length of virus localization at the site of exposure in the muscle was as long as 5 days post-inoculation before the virus entered the central nervous system. At this inoculation site, the virus disappeared transiently between days 7 and 9 after infection but then was restored thereafter until death. Annexin V-fluorescein isothiocyanate staining of splenocytes and thymocytes from mice revealed apoptotic changes in these cells with a marked increase after day 6 of infection. Rabies virus antigen in the brain became detectable 6 days after infection; this occurred parallel to the appearance of apoptosis in the lymphoid cells. There was atrophy of the spleen and thymus, with no evidence of infection. Our results suggest that the interaction between the rabies virus and infected neurons triggers the process of lymphoid cell apoptosis, which reflects the defective operation of the immune system.

Development of a polymerase chain reaction to distinguish monocellate cobra (Naja khouthia) bites from other common Thai snake species, using both venom extracts and bite-site swabs.

A PCR technique was used in this study to identify and distinguish monocellate cobra snake bites using snake venoms and swab specimens from snake bite-sites in mice from bites by other common Thai snakes. The sequences of nucleotide primers were selected for the cobrotoxin-encoding gene from the Chinese cobra (Naja atra) since the sequences of monocellate cobra (Naja kaouthia) venom are still unknown. However, the 113-bp fragment of cDNA of the cobrotoxin-encoding gene was detected in the monocellate cobra venom using RT-PCR. This gene was not found in the venoms of Ophiophagus hannah (king cobra), Bungarus fasciatus (banded krait), Daboia russelii siamensis (Siamese Russell's Viper, and Calloselasma rhodostoma (Malayan pit viper). Moreover, direct PCR could detect a 665-bp fragment of the cobrotoxin-encoding gene in the monocellate cobra venom but not the other snake venoms. Likewise, this gene was only observed in swab specimens from cobra snake bite-sites in mice. This is the first report demonstrating the ability of PCR to detect the cobrotoxin-encoding gene from snake venoms and swab specimens. Further studies are required for identification of this and other snakes from the bite-sites on human skin.

Effects of different biopsy methods on the development of preimplantation mouse embryos, in vitro and in vivo: implication for preimplantation genetic diagnosis.

The aim of this study was to investigate the efficacy of two different embryo biopsy techniques, direct aspiration and partial zona dissection (PZD)-push, on subsequent in vitro and in vivo development of 8-cell stage mouse embryos. It was found that the rates of normal blastocyst formation and hatching blastocysts of direct aspiration, PZD-push, solution control and control embryos were not significantly different (80.8%, 81.6%, 84.5%, 86.7% and 71.9%, 72.3% and 74.6%) respectively. There was, however, a significant reduction in rate of complete hatching blastocysts (P < 0.1) (72.9% aspiration versus 85.2 per cent solution control and 86.4% control) and rate of live-born fetuses (24.2% aspiration versus 43.3% solution control and 41.2% control) (P < 0.05) in the direct aspiration group but no significant difference in the PZD-push group (80.3% of complete hatched blastocysts and 33.8% of live-born fetuses). These findings indicated that embryo biopsy with PZD-push was superior to the direct aspiration method. This mouse embryo biopsy model was useful in advancing development of biopsy technique for human preimplantation genetic diagnosis.