Pengenalpastian dan pengkulturan secara in vitro patogen protozoa, cryptocaryon irritans daripada ikan siakap (lates calcar/fer) untuk tujuan penghasilan vaksin
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Date
2002-06
Authors
Beng Chu, Kua
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Abstract
Cryptocaryoniasis is a major parasitic disease causing high mortalities in
cultured marine and aquarium fishes. Eradication of the protozoal Cryptocaryon irritans
is difficult, as the encysting stage is resistant to drugs while the trophonts are often
buried deep in fish tissue. Therefore, research on effective control methods to prevent
outbreaks of this pathogenic ciliate is urgently needed.
This study has shown that cryptocaryoniasis could occur within 7 to 31 days in
new fish stock exposed to sudden changes of temperature ranging from 16 - 3 8°C and
salinity 27 - 30 ppt. Range 76 - 280 hrs are needed for C. irritans to complete its life
cycle at room temperature (27°C). Studies also shown that the best time to break the life
cycle of C. irritans is 0 - 68 hrs immediately after fish infection. In vitro theronts were
able to infect the seabass within an average 40.00 ± 14.28 hrs. High identity(99%)
amongst the sequence of 16S rDNA proved that the parasite isolated was C. irritans
whereby the comparison amongst the sequence of ITS-I region showed that the local
isolate was of different strain from C. irritans isolated from Australia, Israel and USA.
Studies on the survival of the trophont and theront stages of C. irritans in
different nutrient media, temperature and salinity showed that these three factors could
influence its life span. Trophont had higher number of cyst formation (68.00 ± 5.29)
and production of theront (22.00 ± 2.64) at 30°C. Temperature as high as 40°C attained
43.00 ± 11.79 number of cyst formation but failed to produce the theront stage. As for
the salinity, the highest number of cyst formation was obtained at 20 ppt (24.00 ± 0.00)
as compared to control ( 16.00 ± 1. 73). The survival rate of trophont in different media
showed that only HBSS and sea water media could support all stages of C. irritans.
Studies on survival rate of theronts showed that their mean life span are longer at 30°C
(11.38 ± 1.01 hrs) and in 20 ppt (33.60 ± 0.50 hrs). HBSS medium showed a longer
mean life span, 62.40 ± 23.02 hrs as compared to control seawater medium (29.23 ±
6.81 hrs).
Pathological studies on C. irritans theronts showed that 200-400 theronts would
cause 30% fish mortality within 10 days. Hundred percentage of mortality was also
observed within 5 days after the fish was exposed to 20,000-40,000 theronts. Fish
exposed to 1000-2000 theronts recorded an increase of 30% to 95% mortality within 10
days.
Studies on the immune response in vaccinated fish showed that the antibody titre
was enhanced against live theronts (endpoint titer 24 to 28
) compared with control serum
(titer 22
). It also caused a significant increase in the splenosomatic index (0.36 x 1 o-2
)
compared to control fish (0.03 x I o-2
). Injection of Cryptocaryon antigen was able to
protect fish against the challenge of live theronts [survival rate of 25% - 75% compared
· to control (0%)]. The highest survival at 75% was obtained in fish receiving 2 booster
shots of FTh (Formalin-killed theront) on the ?'h and 141
h day post-vaccination.
Vaccination studies showed that injection of antigen FTh C. irritans was effective
against cryptocaryoniasis in seabass.
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Keywords
Pengkulturan secara in vitro , Cryptocaryon irritans daripada ikan siakap , (Lates calcar/fer) , Penghasilan vaksin