Two forms of vitellogenin were isolated by DEAE agarose ion-exchange chromatography from plasma of the tilapia, Oreochromis mossambicus. The monomers have apparent molecular masses of 200 and 130 kDa, as indicated by SDS-polyacrylamide gel electrophoresis (SDS-PAGE),
and a total amount of phosphorus of 1.7 and 0.1%, respectively. Antibodies specific to the two forms, designated tVTG-200
and tVTG-130, were generated in rabbits and used to develop enzyme-linked immunosorbent assays (ELISAs) and in Western blot
analyses of plasma and oocyte extract. SDS-PAGE of the oocyte extract showed a major protein band at 106.6, minor bands at
26.6, 24.2, and 23.7 kDa, and very faint bands at 83.4 and 17.5 kDa. Western blots of the oocyte extract revealed that the
antiserum to tVTG-200 recognized strongly the protein bands at 24.2 and 23.7 kDa, and less strongly the bands at 25.1 and
22.6 kDa, whereas the antiserum to tVTG-130 recognized mainly the protein band at 106.6 kDa. The presence of both VTGs in
untreated male tilapia was detected with the ELISAs using relatively high plasma volumes. Their presence in males was confirmed
by VTG-like immunoreactive materials eluting from the ion-exchange column at the same positions as tVTG-200 and tVTG-130.
The concentrations of the VTGs in males were several orders of magnitude lower than in vitellogenic females. Treatment of
male tilapia with estradiol-17β (E2) induced both VTGs within 24h. After 7 days, tVTG-130 reached a maximum concentration in plasma, whereas tVTG-200 continued
to increase. Our findings demonstrate that the two vitellogenins are biochemically distinct, possibly differentially regulated,
and made by both sexes. 相似文献
The present study aimed to determine whether protection is conferred by immunization of grouper, Epinephelus coioides, against a protozoan parasite, Cryptocaryon irritans. The immunization of E. coioides was carried out by a low level exposure of fish to live C. irritans theronts from predetermined number of tomonts and by an intraperitoneal injection of a vaccine consisting of formalin-killed C. irritans theronts.
Mucus titers detected by ELISA were significantly higher in fingerling and adult grouper subjected to the low level of exposure to C. irritans theronts at 3-week post-exposure compared to fish that had no previous exposure. In addition, significantly smaller tomonts were produced from adult grouper after three successive exposures than the tomonts produced after a single exposure to the parasite.
In the vaccine-immunization experiment, no mortality was monitored in fish that received high dose vaccine (100 μg/fish), while 40% cumulative mortality and 100% cumulative mortality were recorded in low dose group (10 μg/fish) and control group (PBS-injected), respectively. In the succeeding replicate, the vaccine-immunized group (high dose) had 37.5% cumulative mortality and 100% cumulative mortality for the control. In addition, a total of 1830 tomonts were collected at 5-day post-challenge from the control group while none from the vaccine-immunized group. Significantly fewer trophonts and tomonts were enumerated at 5-day and 7-day post-challenge, respectively, in the vaccine-immunized group than the control.
Results suggest that a protective immunity has been conferred on the immunized grouper as indicated by high antibody titers in the mucus of C. irritans-exposed fish and higher survival and fewer parasites in vaccine-immunized fish than the control groups. The conferred immunity played a major role in preventing or limiting the adhesion, invasion, and development of C. irritans theronts on the skin of the immunized grouper. 相似文献
