Structural and Functional Characterization of Liposomal Recombinant Hepatitis B Vaccine

Abstract

Liposomal hepatitis B vaccine was prepared by encapsulating recombinant 22-nm hepatitis B surface antigen (HBsAg) particles derived from a Chinese hamster ovary (CHO) cell line in multilammelar lipid vesicles (MLV) composed of 9:1 dimyristoyl phosphatidyl-choline/dimyristoyl phosphatidylglycerol. The CHO-derived HBsAg particles reveal 6 bands in polyacrylamide gel electrophoresis related to the presence of 3 peptides (S, M, & L). Four different methods were used to prepare the MLV vaccine, each resulting in freeze-dried powder which upon hydration gave MLV of a similar mean size, 4.5 μm. The humoral response to these 4 liposomal vaccines in mice was dependent on the method of preparation, but for all of them it was better than the response to alum-based vaccine (especially at a low dose of antigen). Comparison of vaccination using “naked” HBsAg particles, particles adsorbed to alum, and particles encapsulated in liposomes demonstrated that at low dose of antigen the liposomal vaccine was superior in eliciting humoral response. Encapsulation in liposomes did not improve specific cytotoxic T lymphocyte (CTL) response. The alum in the vaccine completely eliminates CTL response, though it improved the humoral response by increasing the linear range in the antigen dose-response curve (increasing the antibody titer at high antigen dose). A similar response profile was obtained with recombinant yeast (Hansenula) 22-nm particles composed of a single non-glycosylated (p24) peptide and lipids. The similarity in the response to the mammalian cell and yeast derived vaccine suggests that the physical nature of the vaccine, more than the exact composition, determines the balance between humoral and CTL responses.     

Maternal antibody blocks humoral but not T cell responses to BVDV.

Bovine viral diarrhoea virus (BVDV) contributes significantly to health-related economic losses in the beef and dairy industry. Antibodies of maternal origin can be protective against BVDV infection, however, calves with low titres of maternal antibody or that do not receive colostrum may be at risk for acute BVDV infection. Interference by high titres of maternal antibodies prevents the development of an antibody response following vaccination with either a killed or attenuated BVDV vaccine. However, the T cell mediated immune response to BVDV may be generated in the absence of a detectable serum neutralizing antibody response. Two trials were conducted to evaluate the potential to elicit T cell mediated immune responses to BVDV in calves with circulating maternal antibody to BVDV. In the first trial, calves with high levels of circulating maternal antibody to BVDV 1 and BVDV 2 were experimentally infected with BVDV 2 (strain 1373) at two to five weeks of age. The T-cell mediated immune responses of the experimentally infected calves and non-infected calves were monitored monthly until circulating maternal antibody was no longer detectable in either treatment group. Calves experimentally infected with BVDV developed BVDV specific CD4(+), CD8(+), and delta T cell responses while high levels of maternal antibody were circulating. A second challenge with BVDV 2 (strain 1373) was performed in the experimentally infected and control calves once maternal antibody could no longer be detected. Previous exposure to BVDV in the presence of maternal antibody protected calves from clinical signs of acute BVDV infection compared to the control calves. In the second trial, three groups of calves with circulating maternal antibody to BVDV were given either a modified live vaccine (MLV) containing BVDV 1 and BVDV 2, a killed vaccine containing BVDV 1 and BVDV 2, or no vaccine, at seven weeks of age. Serum neutralizing antibody levels and antigen specific T cell responses were monitored for 14 weeks following vaccination. Calves vaccinated with MLV BVDV developed BVDV 1 and BVDV 2 specific CD4(+)T cell responses, and BVDV 2 specific gammadelta T cell responses, in the presence of maternal antibody. Vaccination with killed BVDV did not result in the generation of measurable antigen specific T cell immune responses. In this trial, a second vaccination was performed at 14 weeks to determine whether an anamnestic antibody response could be generated when calves were vaccinated in the presence of maternal antibody. Calves vaccinated with either a MLV or killed BVDV vaccine while they had maternal antibody developed an anamnestic antibody response to BVDV 2 upon subsequent vaccination. The results of these trials indicate that vaccinating young calves against BVD while maternal antibody is present may generate BVDV specific memory T and B cells. The data also demonstrated that seronegative calves with memory T and B cells specific for BVDV may be immune to challenge with virulent BVDV.     

Milk intake in blue fox (Alopex lagopus) and silver fox (Vulpes vulpes) cubs in the early suckling period

Milk intake of fox cubs (2-16 days of age; body weight, 96-649 g) in ten blue fox litters and ten silver fox litters were measured by the water isotope dilution (WID) technique following a single intraperitoneal injection of tritiated water (3HHO). Litter size varied from four to 14 in blue foxes and from three to eight in silver foxes. Silver fox cubs had higher birth weights than blue foxes. Inter-species body weights and growth rates were apparently dependent on litter size and the dam's constitution. In both species growth rate increased with age and body weight (7-35 g per day). In the cubs, the biological half-life of body water turnover (BWT) rose from 1.5 days at 2-3 days of age to 2.5 days at 13-16 days of age, although a considerable scatter was seen. The mean daily milk intake of the cubs varied with body weight, from 31 to 193 g per day, whereas daily milk intake per unit of body mass remained stable at 30-35 g per 100 g body weight. The ratio of milk intake to body weight gain varied considerably among cubs, averaging 4.5 g/g during the 3-week experimental period. In suckling fox cubs, the calculated daily intake of metabolically energy (ME) corresponded fairly with the estimated energy requirements for growth and maintenance of the young. Finally, the applicability and the accuracy of the WID technique was evaluated in ten 3-week-old fox cubs, by tube-feeding with a milk replacer for 48 h, which documented that the daily rates of milk intake and water turnover can be accurately measured in suckling fox cubs by the WID technique following a single injection of 3HHO.     

No evidence of murine leukemia virus-related viruses in live attenuated human vaccines

Background

The association of xenotropic murine leukemia virus (MLV)-related virus (XMRV) in prostate cancer and chronic fatigue syndrome reported in previous studies remains controversial as these results have been questioned by recent data. Nonetheless, concerns have been raised regarding contamination of human vaccines as a possible source of introduction of XMRV and MLV into human populations. To address this possibility, we tested eight live attenuated human vaccines using generic PCR for XMRV and MLV sequences. Viral metagenomics using deep sequencing was also done to identify the possibility of other adventitious agents.

Results

All eight live attenuated vaccines, including Japanese encephalitis virus (JEV) (SA-14-14-2), varicella (Varivax), measles, mumps, and rubella (MMR-II), measles (Attenuvax), rubella (Meruvax-II), rotavirus (Rotateq and Rotarix), and yellow fever virus were negative for XMRV and highly related MLV sequences. However, residual hamster DNA, but not RNA, containing novel endogenous gammaretrovirus sequences was detected in the JEV vaccine using PCR. Metagenomics analysis did not detect any adventitious viral sequences of public health concern. Intracisternal A particle sequences closest to those present in Syrian hamsters and not mice were also detected in the JEV SA-14-14-2 vaccine. Combined, these results are consistent with the production of the JEV vaccine in Syrian hamster cells.

Conclusions

We found no evidence of XMRV and MLV in eight live attenuated human vaccines further supporting the safety of these vaccines. Our findings suggest that vaccines are an unlikely source of XMRV and MLV exposure in humans and are consistent with the mounting evidence on the absence of these viruses in humans.     

Behavioural Differences between Single Scandinavian Brown Bears (Ursus arctos) and Females with Dependent Young When Experimentally Approached by Humans

Carnivore-human encounters that result in human injury present a conservation and management challenge and it is therefore important to understand under what conditions such incidents occur. Females with cubs are often involved when humans are injured by brown bears Ursus arctos. In Scandinavia, this is particularly true for unarmed recreational forest users. Our aim was to document behavioural differences between single bears and females with cubs in order to develop recommendations to minimize the risk of injuries to recreational forest users. We documented the reactions of GPS-collared females with cubs and single brown bears to experimental approaches by humans to 50 m from the bear on 42 and 108 occasions, respectively. The majority of females with cubs (95%) and single bears (89%) left when approached. Bears that left were passed at shorter distances and were in more open areas than those that stayed. Both groups had similar flight initiation distances, which were longer for bears that were active at the time of the disturbance. Females with cubs selected more open habitat than single bears, also for the new site they selected following disturbance. Females with cubs, particularly active females with cubs of the year, moved greater distances and spent more time active following the approach. Females with cubs and single bears were seen or heard in 26% and 14% of the approaches, respectively. None of the bears displayed any aggressive behaviour during the approaches. Females with cubs selected more open habitat, perhaps predisposing them to encountering people that are not involved in hunting activities, which might be the primary explanation why females with cubs are most frequently involved when unarmed people are injured by bears in Scandinavia. To mitigate injury risks, one must consider factors that bring bears closer to human activity in the first place.     

Selection of maternity dens by female polar bears in western Hudson Bay,Canada and the effects of human disturbance

We deployed 19 satellite radio collars on adult female polar bears (Ursus maritimus) in western Hudson Bay to study how pregnant females select maternity-den sites and the degree to which human disturbance might influence this process. To eliminate the possibility that being handled while pregnant might influence the final choice of a suitable site, the collars were attached 1 year before the females were expected to select dens. Our results indicated that pregnant females moved inland and selected an area to den shortly after coming ashore in the summer and remained there if not disturbed. We analyzed 24 years of consecutive data (1979–2002) to examine whether disturbance of pregnant females in the denning area in the autumn affected litter size or mass of cubs the following spring. We defined three levels of disturbance (none—bear not seen; low—bear seen but not handled; high—bear seen and handled), and found that disturbance had no effect on either litter size or the mass of male cubs. However, we did find that females handled in the autumn had significantly lighter female cubs than females that were not seen. Our disturbance of pregnant females occurred in August and September, which is only a few months prior to parturition; it is not known what the effects of disturbance might be in late October or early November, closer to the time when cubs are born.     

Growth rate of 21 captive-born,mother-raised cheetah cubs

This paper presents the growth rate of 21 clinically normal, mother-raised. captive cheetah cubs from birth through 45 days of age. The development of a growth curve for healthy, mother-raised cheetah cubs provides a diagnostic tool for individuals involved in cheetah propagation. Use of the curve may alert caretakers to problems early and thus help reduce the high neonatal mortality rate seen in captive-born cheetah cubs. The growth curve was constructed using 21 (11 males and 10 females) captiveborn cheetah cubs from six litters (offspring of three unrelated adult males and three unrelated adult females) born at the Columbus Zoo from September 1985 through December 1989. Each cub was weighed to the nearest gram the morning after birth and approximately the same time every consecutive morning for 45 days. The mean weight the morning after birth was 463 g (range 385–542 g). The average litter size was 3.5 (range 2–4, n = 6). The daily weight gain was 40–50 g/cub/day.     

Retroviral vectors for vaccine development: induction of HIV-1-specific humoral and cellular immune responses in rhesus macaques using a novel MLV(HIV-1) pseudotype vector

Retroviral vectors have yet not been tested for their potential as vaccines despite their frequent utilization in gene therapy allowing for highly efficient gene transfer into a number of cell types and their suitability for large-scale production in biotechnology. To investigate MLV-based vectors suitability for inducing immune response against HIV-1-antigens, we generated a MLV(HIV-1) pseudotype vector enabling CD4-specific transduction of HIV-1 genes env, vpu, tat and rev originating from the pathogenic SHIV-89.6P. Functional expression of the lentiviral genes in packaging cells, human and rhesus CD4+ target cells was demonstrated by various assays. Following highly efficient ex vivo transduction, up to 3.4x10(7) autologous, transfer vector-positive rhesus peripheral blood mononuclear cells (rhPBMCs) were re-inoculated into a rhesus macaque. Five weeks after the initial inoculation HIV-1 Env-specific antibodies were detected using ELISA. ELIspot-assay revealed the induction of a HIV-1 Rev and Env-specific CTL-response 7.5 weeks after immunization. Thus, these novel MLV(HIV-1) vectors facilitate efficient transduction and subsequent expression of HIV-1-genes in CD4-positive host cells. Induction of both humoral and cellular HIV-1-specific immune responses in vivo confirmed their potential as an effective HIV-1 vaccine to be further studied in SHIV/rhesus macaque model of lentivirus infection.     

Comparison of molecular and biological characteristics of a modified live porcine reproductive and respiratory syndrome virus (PRRSV) vaccine (ingelvac PRRS MLV), the parent strain of the vaccine (ATCC VR2332), ATCC VR2385, and two recent field isolates of PRRSV

The objectives of this study were to compare the molecular and biological characteristics of recent porcine reproductive and respiratory syndrome virus (PRRSV) field isolates to those of a modified live virus (MLV) PRRS vaccine and its parent strain. One hundred seventeen, 4-week-old pigs were randomly assigned to six groups. Group 1 (n = 20) served as sham-inoculated negative controls, group 2 (n = 19) was inoculated with Ingelvac PRRS MLV vaccine, group 3 (n = 20) was inoculated with the parent strain of the vaccine (ATCC VR2332), group 4 (n = 19) was inoculated with vaccine-like PRRSV field isolate 98-38803, group 5 (n = 19) was inoculated with PRRSV field isolate 98-37120, and group 6 (n = 20) was inoculated with known high-virulence PRRSV isolate ATCC VR2385. The levels of severity of gross lung lesions (0 to 100%) among the groups were significantly different at both 10 (P < 0.0001) and 28 days postinoculation (p.i.) (P = 0.002). At 10 days p.i., VR2332 (26.5% +/- 4.64%) and VR2385 (36.4% +/- 6.51%) induced gross lesions of significantly greater severity than 98-38803 (0.0% +/- 0.0%), 98-37120 (0.8% +/- 0.42%), Ingelvac PRRS MLV (0.9% +/- 0.46%), and negative controls (2.3% +/- 1.26%). At 28 days p.i., 98-37120 (17.2% +/- 6.51%) induced gross lesions of significantly greater severity than any of the other viruses. Analyses of the microscopic-interstitial-pneumonia-lesion scores (0 to 6) revealed that VR2332 (2.9 +/- 0.23) and VR2385 (3.1 +/- 0.35) induced significantly more severe lesions at 10 days p.i. At 28 days p.i., VR2385 (2.5 +/- 0.27), VR2332 (2.3 +/- 0.21), 98-38803 (2.6 +/- 0.29), and 98-37120 (3.0 +/- 0.41) induced significantly more severe lesions than Ingelvac PRRS MLV (0.7 +/- 0.17) and controls (0.7 +/- 0.15). The molecular analyses and biological characterizations suggest that the vaccine-like isolate 98-38803 (99.5% amino acid homology based on the ORF5 gene) induces microscopic pneumonia lesions similar in type to, but different in severity and time of onset from, those observed with virulent strains VR2385 and the parent strain of the vaccine. Our data strongly suggest that isolate 98-38803 is a derivative of Ingelvac PRRS MLV and that the isolate is pneumovirulent.     

Combination therapy with a synthetic peptide of C-reactive protein and interleukin 2: augmented survival and eradication of pulmonary metastases

A synthetic peptide (RS-83277) derived from the structure of human C-reactive protein (CRP) was previously shown to have antitumor activity in three different murine tumor models when administered in multilamellar vesicles (MLV). The therapeutic effects were comparable to those seen with MLV-encapsulated native CRP. The present study evaluated the therapeutic and immunomodulatory effects of administering CRP peptide RS-83277 MLV simultaneously with low-dose recombinant interleukin-2 (IL-2) to C57B1/6 mice bearing established pulmonary metastases of fibrosarcoma T241. Results demonstrated that the capacity of RS-83277 MLV to inhibit tumor metastases and prolong survival was significantly augmented by combination with 10 000 U/day IL-2 i.p. Treated animals showed no evidence of toxicity. By immunohistochemistry, increased Thy 1.2⁺ cells were detectable in lungs of RS-83277 MLV/IL-2-treated animals compared to those receiving RS-83277 MLV alone. Circulating tumor necrosis factor (TNF) and interferon (IFN) were not detectable in animals receiving RS-83277 MLV alone, but TNF was significantly elevated in animals receiving IL-2. In the presence of combination therapy, however, circulating TNF was not detectable. Results suggest that the combination of synthetic CRP peptide RS-83277 MLV and low-dose IL-2 offers a therapeutic advantage over either agent alone.This work was supported in part by grant CA49950 from the National Cancer Institute (B.P.B.), and grant 43618 from the National American Cancer Society (S.D.D.)     

Aviation noise does not impair the reproductive success of farmed blue foxes

The aim of our study was to assess the effects of aviation noise on reproduction and cub mortality in farmed blue foxes. Eighty artificially inseminated blue fox vixens (45 primiparous and 35 multiparous) were exposed to aviation noise on 5 days when they were pregnant or had cubs. The noise during the exposures varied from 85 to 121 dB (L(AFmax)). Vixens (45 primiparous and 34 multiparous) on a farm without flight action acted as controls. Cubs were counted 1, 3, 7, 14 and 49 days postpartum and at the beginning of July. Litter size (cubs per whelped vixen), reproductive performance (cubs per mated vixen) and cub losses (lost cubs per whelped vixen) were analyzed from both experimental farms (A and C). The flight action had no effect on reproductive success. Reproductive performance in primiparous vixens was 4.2+/-3.8 and 4.3+/-3.6 cubs (ns, Mann-Whitney U-test) in the control and aviation group, respectively, while in multiparous vixens the corresponding figures were 7.1+/-4.4 and 7.3+/-3.8 cubs (ns). In general, litter size declined from birth to weaning (in primiparous vixens from 8.1+/-3.8 to 5.4+/-3.2 cubs, and in multiparous from 9.7+/-3.8 to 7.2+/-3.8 cubs, P<0.001, GLM for repeated measures). The decline was greater in primiparous than in multiparous vixens (P<0.01). There were no differences in total cub losses between the experimental groups (ns). Accordingly, the present results show that exposure to severe and repeated aviation noise does not impair the reproductive success of farmed blue foxes.     

Body composition of three polar bear (Ursus maritimus) cubs found dead at Svalbard

We determined the body composition from the carcasses of three 3- to 4-months-old polar bear cubs found dead at Svalbard, Norway. Two of the cubs were killed by a male bear at the den and one cub died from starvation. These three carcasses weighed 70-77% of total body mass of cubs. The ash and protein content of the three carcasses were similar but the starving cub had higher water content and lower fat content than the two killed cubs. The fat content varied from 1.3% (starving cub) to 12.9% while the content of water and fat was similar (about 76%) in the three cubs. More data on the body composition of polar bear cubs at time of emergence from the den are needed to understand the breeding ecology and reproductive energetics of female polar bears.     

Short-term behavioural response of polar bears (<Emphasis Type="Italic">Ursus maritimus</Emphasis>) to snowmobile disturbance

In this study the distance, at which polar bears detected and actively responded to approaching snowmobiles was measured and the behavioural response was recorded. The study was performed on Svalbard, an arctic island where human traffic has increased substantially in recent years. Fieldwork was conduced in April and/or May during the years 2003–2005. Polar bears were observed on ice with telescopes and binoculars. Undisturbed polar bears were observed continuously and their behaviours recorded, during the time when two snowmobiles moved toward the bear(s). Distances between the bear, the observer, and the approaching snowmobiles were measured using GPS positions taken on the track towards the bear. Data on the behavioural response of 20 encounters with bears were collected. On average, bears were alerted to the snowmobiles at 1,164 m. Mean distance at which the locomotive response occurred was 843 m, and there was a statistical significant difference in distance between sex and age classes [326 m (95% CI = 138–496 m) for adult males; 1,534 m (95% CI = 508–2,768 m) for adult females with cubs; 164 m (95% CI = 49–543 m) for two adult females without cubs; and 1,160 m (95% CI = 375–1,353 m) for single medium sized bears]. The responses of the polar bears to the snowmobiles were categorized according to intensity and persistence of reactions. Females with cubs and single medium sized bears tended to show more intense responses than adult males and lone adult females. Wind direction affects sound and odour transmission, and although an effect on response distance was not found, the response intensity was affected by wind direction. We conclude that female polar bears with small cubs in particular may have a greater risk to be disturbed, since they react at greater distances with amplified reactions; thus, users of snowmobiles should take particular care in areas where females with cubs are present.     

Dendritic cell-mediated HIV-1 transmission to T cells of LAD-1 patients is impaired due to the defect in LFA-1

Background

Murine Leukemia Virus (MLV) assembly has been long thought to occur exclusively at the plasma membrane. Current models of retroviral particle assembly describe the recruitment of the host vacuolar protein sorting machinery to the cell surface to induce the budding of new particles. Previous fluorescence microscopy study reported the vesicular traffic of the MLV components (Gag, Env and RNA). Here, electron microscopy (EM) associated with immunolabeling approaches were used to go deeply into the assembly of the "prototypic" MLV in chronically infected NIH3T3 cells.

Results

Beside the virus budding events seen at the cell surface of infected cells, we observed that intracellular budding events could also occur inside the intracellular vacuoles in which many VLPs accumulated. EM in situ hybridization and immunolabeling analyses confirmed that these latter were MLV particles. Similar intracellular particles were detected in cells expressing MLV Gag alone. Compartments containing the MLV particles were identified as late endosomes using Lamp1 endosomal/lysosomal marker and BSA-gold pulse-chase experiments. In addition, infectious activity was detected in lysates of infected cells.

Conclusion

Altogether, our results showed that assembly of MLV could occur in part in intracellular compartments of infected murine cells and participate in the production of infectious viruses. These observations suggested that MLV budding could present similarities with the particular intracellular budding of HIV in infected macrophages.     

Heterogeneous antibody response to polyvalent melanoma vaccines in syngeneic mice

In this study, a human melanoma vaccine induced antibody responses in mice that varied significantly from animal to animal. BALB/c mice were immunized to a xenogenic human polyvalent melanoma vaccine that has been used in phase II clinical trials in over 600 patients. Mice were bled biweekly for up to 6 weeks to measure antibody responses. IgG antibody responses to the melanoma vaccine components were detectable within 2 weeks but were much stronger at 4 and 6 weeks. When the pooled sera were further analyzed by Western blot, a complex pattern of antigens was detected. When individual sera from identically immunized mice were assayed by Western blot, a consistent, reproducible pattern of antigen recognition was not seen. Rather, we found significantly different antibody responses among the mice. Both the intensity of antibody responses and the pattern of antigens recognized varied from animal to animal. Although there appeared to be immunodominant antigens that produced antibody responses in most mice, no single antigen induced antibody responses in all mice. These results demonstrate that polyvalent vaccines induce heterogeneous antibody responses in mice treated identically. Analysis of the response of selected melanoma patients immunized to the same vaccine revealed similar antibody responses to the antigens in the melanoma vaccine. Heterogeneity may hamper interpretation of vaccine immunogenicity and relevant tumor antigens in humans.     

Vaccination of pigs against swine influenza viruses by using an NS1-truncated modified live-virus vaccine

Swine influenza viruses (SIV) naturally infect pigs and can be transmitted to humans. In the pig, genetic reassortment to create novel influenza subtypes by mixing avian, human, and swine influenza viruses is possible. An SIV vaccine inducing cross-protective immunity between different subtypes and strains circulating in pigs is highly desirable. Previously, we have shown that an H3N2 SIV (A/swine/Texas/4199-2/98 [TX98]) containing a deleted NS1 gene expressing a truncated NS1 protein of 126 amino acids, NS1black triangle126, was attenuated in swine. In this study, 4-week-old pigs were vaccinated with the TX98 NS1black triangle126 modified live virus (MLV). Ten days after boosting, pigs were challenged with wild-type homologous H3N2 or heterosubtypic H1N1 SIV and sacrificed 5 days later. The MLV was highly attenuated and completely protected against challenge with the homologous virus. Vaccinated pigs challenged with the heterosubtypic H1N1 virus demonstrated macroscopic lung lesions similar to those of the unvaccinated H1N1 control pigs. Remarkably, vaccinated pigs challenged with the H1N1 SIV had significantly lower microscopic lung lesions and less virus shedding from the respiratory tract than did unvaccinated, H1N1-challenged pigs. All vaccinated pigs developed significant levels of hemagglutination inhibition and enzyme-linked immunosorbent assay titers in serum and mucosal immunoglobulin A antibodies against H3N2 SIV antigens. Vaccinated pigs were seronegative for NS1, indicating the potential use of the TX98 NS1black triangle126 MLV as a vaccine to differentiate infected from vaccinated animals.     

Milk composition in free-ranging polar bears (Ursus maritimus) as a model for captive rearing milk formula

The goals of this study were to have an improved understanding of milk composition and to help create a suitable milk formula for cubs raised in captivity. Milk samples were evaluated for fat, fatty acids, carbohydrate, vitamin D(3), 25(OH)D(3), vitamin A (retinol), vitamin E (α-tocopherol), protein, and amino acids. Total lipids in milk did not differ for cubs (mean ± SEM = 26.60 ± 1.88 g/100 ml vs. yearlings 27.80 ± 2.20 g/100 ml). Milk lipids were of 23.6% saturated fatty acid for cubs and 22.4% for yearlings. Milk consumed by cubs and yearlings contained 43.8 and 42.0% mono-unsaturated fatty acids and 23.4 and 21.9% polyunsaturated fatty acids, respectively. Carbohydrate content was higher in milk for cubs (4.60 ± 0.64 g/100 ml) than for yearlings (2.60 ± 0.40 g/100 ml). Vitamin D(3) concentration of milk was 18.40 ± 5.00 ng/ml in early lactation compared with 7.60 ± 2.00 ng/ml for mid-lactation. 25(OH)D(3) was lower in milk consumed by cubs (162.00 ± 6.70 pg/ml) than in milk consumed by yearlings (205.00 ± 45.70 pg/ml). Vitamin A concentrations were 0.06 ± 0.01 and 0.03 ± 0.01 μg/ml for cubs and yearlings, respectively. Vitamin E was higher in milk consumed by cubs (20.16 ± 4.46 μg/ml) than by yearlings (7.30 ± 1.50 μg/ml). Protein content did not differ in milk available to cubs (11.40 ± 0.80 g/100 ml compared with milk for yearlings 11.80 ± 0.40 g/100 ml). Taurine was the most abundant free amino acid at 3,165.90 ± 192.90 nmol/ml (0.04% as fed basis).     

圈养大熊猫同时哺育双胞胎行为观察

2003 年,圈养大熊猫“梅梅”首例哺育成活一胎二仔,通过对其近半年的育幼行为观察,结果发现:1) 母兽主要以同时衔2 仔、同时衔和抱2 仔二种方式将幼仔抱入怀中哺育;60 d 内, 育幼姿势以坐位为主,倦卧其次, 其它姿势更少, 其中坐位随日龄增加逐渐减少, 倦卧变化不大。2) 母兽活动时间在产仔当天最多, 之后显著下降并维持在35.2 ± 0.6% 的低水平, 47 d 后再缓慢上升到108 d 后的54.8 ± 0.9% 。3) 双胞胎幼仔间哺乳的日均次数和时间无显著差异。4) 7 d 内2 幼仔“仔在母身上” 的时间占100%, 21 ～ 23 d 后显著减少, 而“母体盖仔”、“仔在母身边”和“母仔自然分离”的时间显著增加, 但“母体盖仔” 的时间在32 d 左右后又显著减少;双胞胎分别在与母兽的此四种位置变化的时间上无显著差异。5) 母兽的活动、幼仔哺乳日均次数、“母仔自然分离”在全天的日均时间分布有峰、谷变化。6) 随幼仔活动能力的逐渐增强,幼仔离“育幼窝”的距离也逐渐增加,双胞胎幼仔离“育幼窝”的远近也有差别。7)母兽分别与其雄性双胞胎幼仔玩耍的时间有显著差异,而两幼仔自玩的时间无差异,此两双胞胎自玩和一起玩耍所用时间远大于分别与母兽玩耍的时间。该研究丰富了大熊猫育幼行为内容,并为以后的大熊猫双胞胎育幼提供了可供参考的行为资料。     

Economic Analysis of Vaccination Strategies for PRRS Control

Porcine reproductive and respiratory syndrome virus (PRRSv) is a swine-specific pathogen that causes significant increases in production costs. When a breeding herd becomes infected, in an attempt to hasten control and elimination of PRRSv, some veterinarians have adopted a strategy called load-close-expose which consists of interrupting replacement pig introductions into the herd for several weeks (herd closure) and exposing the whole herd to a replicating PRRSv to boost herd immunity. Either modified-live virus (MLV) vaccine or live field-virus inoculation (FVI) is used. This study consisted of partial budget analyses to compare MLV to FVI as the exposure method of load-close-expose program to control and eliminate PRRSv from infected breeding herds, and secondly to estimate benefit / cost of vaccinating sow herds preventatively. Under the assumptions used in this study, MLV held economic advantage over FVI. However, sensitivity analysis revealed that decreasing margin over variable costs below $ 47.32, or increasing PRRSv-attributed cost above $18.89 or achieving time-to-stability before 25 weeks resulted in advantage of FVI over MLV. Preventive vaccination of sow herds was beneficial when the frequency of PRRSv infection was at least every 2.1 years. The economics of preventative vaccination was minimally affected by cost attributed to field-type PRRSv infection on growing pigs or by the breeding herd productivity level. The models developed and described in this paper provide valuable tools to assist veterinarians in their efforts to control PRRSv.     

A note on the reproductive success of primiparous blue fox vixens in social groups

Our aim was to compare traditional breeding system, i.e. artificial insemination with singly-housing, to alternative breeding systems in farmed blue fox (Alopex lagopus or Vulpes lagopus) vixens. At the age of 7 weeks (i.e. at weaning), 48 randomly selected female blue fox cubs were divided into four experimental groups: (1) artificially inseminated singly-housed vixens in cages (AI-SC), (2) artificially inseminated pair-housed vixens in double-cages (AI-PC), (3) naturally bred pair-housed vixens with a male in triple-cages (NB-PC), and (4) naturally bred pair-housed vixens with a male in outdoor enclosures (NB-PE). The cubs were counted on days 1, 2, 3, 7, 14 and 49 postpartum and the reproductive performance per breeding (RPB) and per mated vixens (RPM) with its subcomponents were formed from these data. RPB was zero in both NB groups. In AI-PC and AI-SC, RPB was 1.3+/-2.5 and 4.1+/-4.7 cubs, respectively. RPB and the percentage of vixens that weaned cubs were lower in NB-PC and NB-PE than in AI-PC. In AI groups, both RPB and RPM at weaning tended to be better in singly-housed than in pair-housed blue fox vixens. No statistically significant differences were found between AI groups in the percentage of vixens without oestrus, barren vixens, vixens that lost all cubs or weaned at least one cub. Only in one pair (AI-PC) both vixens displayed communal breeding and weaned cubs. The present results show that vixens in traditional breeding system had better RPM than in any of the studied pair-housing systems. The current blue fox population has been effectively selected for cage-breeding and artificial reproduction, and therefore blue foxes generally reproduce well with artificial insemination after careful detection of oestrus in traditional breeding conditions.