FIELD VACCINATION WITH PALM OIL ADJUVANTED FEED-BASED STREPTOCOCCUS AGALACTIAE BACTERIN PROTECTS TILAPIA FROM STREPTOCOCCOSIS

Mohd Syafiq Mohammad Ridzuan^1,2, Azila Abdullah², Norazsida Ramli³, Hanan Mohd Yusof⁴, Mohd Firdaus-Nawi^1,*

¹Department of Marine Science, Kulliyyah of Science, International Islamic University Malaysia, Bandar Indera Mahkota, 25200 Kuantan, Pahang, Malaysia

²National Fish Health Research Centre (NaFisH), Fisheries Research Institute (FRI) Batu Maung, Department of Fisheries Malaysia, 11960 Batu Maung, Penang, Malaysia

³Kulliyyah of Allied Health Science, International Islamic University Malaysia, Bandar Indera Mahkota, 25200 Kuantan, Pahang, Malaysia

⁴Fisheries Research Institute (FRI) Glami Lemi, Department of Fisheries Malaysia, 71660 Titi Jelebu, Negeri Sembilan, Malaysia

^*Corresponding author: firdausn@iium.edu.my

Tilapia (Oreochromis sp.) is one of the most important globally traded commodities in freshwater aquaculture industry. Intensive tilapia farming is, however, threatened by a bacterial infection caused by Streptococcus agalactiae, the causative agent of streptococcosis. Therefore, a tilapia farm that experiencing the endemic of the disease was selected to assess the efficacy of palm oil adjuvanted feed-based streptococcosis vaccine on naturally occurring streptococcosis. A total of 6,000 red tilapias were selected and divided evenly into six cages. Fish from Group 1 in cages number 1 and 2 were not vaccinated to represent negative control. Group 2 in cages number 3 and 4 were vaccinated twice on week 0 and week 2 (single booster), while Group 3 in number cages 5 and 6 were vaccinated thrice on week 0, week 2, and week 6 (double booster). Vaccination was done by oral administration of the palm oil adjuvanted feed-based bacterin vaccine at an average of 5% body weight. Organ samples of eye, brain, and kidney were collected to isolate the bacteria at two-weeks intervals until the end of the 16-week trial period. From the study, S. agalactiae was isolated as early as week 2 at an average of 11.1 ± 19.2% of the sampled fish but increased to 21.1 ± 5.1% in week 4, to 12.2 ± 21.2% in week 6, to 30.0 ± 26.5% in week 10 and 21.1 ± 10.2% in week 14. The average isolation rate was 19.6 ± 20.2%, 11.5 ± 11.3%, and 8.2 ± 9.4% of the fish sampled from unvaccinated, single, and double booster groups, respectively. At the end of the study period, the mortality rate was 23.6 ± 1.8% for unvaccinated, 14.5 ± 5.9% for single booster and 6.4 ± 0.4% for double booster groups. These results indicate that a double booster vaccination regime with palm oil adjuvanted vaccine can reduce the incidence of streptococcosis to 11.4% and improve the survival rate to 93.6%. Given current tilapia farming practices and economic constraints, oral administration may be the most suitable approach for vaccination.

Keywords: Oral vaccination, Field trial, Tilapia, Streptococcosis, Streptococcus agalactiae

PRELIMINARY STUDY ON ORAL ADMINISTRATION OF NA-ALGINATE AND MULTIVITAMIN TO IMPROVE THE NON-SPECIFIC IMMUNE SYSTEM IN RED TILAPIA

Muhammad Fauzi Azhari, Ridho Nasih Adi Kurniawan, Alim Isnansetyo*

Department of Fisheries, Faculty of Agriculture, Universitas Gadjah Mada, Jl. Flora, Bulaksumur, Yogyakarta

*corresponding author: isnansetyo@ugm.ac.id

Alginate from Sargassum sp. and multivitamin stimulate immune response of fish, but their effective combination has not been reported yet. The objectives of this study were to evaluate the effect of Na-alginate and multivitamin by oral administration on the non-specific humoral and cellular immune of tilapia, and to determine the their effective doses. This study used Completely Randomized Design (CRD) with 6 treatments in triplicates. The treatments were negative controls (without Na-alginate and multivitamins) (T1), Na-alginate at 4 g / kg of feed (T2), multivitamin 2.5 g / kg) (T3), Na-alginate 1g / kg and multivitamin 0.625 g / kg of feed (T4), Na-alginate 2 g / kg and multivitamin 1.25 g / kg (T5), Na-alginate 4 g / kg and multivitamin 2.5 g / kg of feed (T6). The fish were kept for 12 days and the tests were performed on days 0, 4, 8, and 12. Feeding was done twice daily at a feeding rate of 3% biomass. The observed parameters included phagocytosis activity, phagocytosis index, superoxide dismutase (SOD), extracellular respiratory burst, leukocyte differentiation, hematocrit, leukocrit, total plasma protein, natural agglutination, serum antibacterial activity, hematocrit and leucocrite, and water quality. The results showed that oral administration of Na-alginate increased respiratory burst, phagocytosis index, lymphocyte percentage, serum bacterial activity against A. hydrophila and natural agglutination. Multivitamins increased the percentage of lymphocytes, but did not influence humoral non-specific immune parameters. Combination of Na-alginate and multivitamins orally increased the parameters of phagocytosis activity, phagocytosis index, SOD, respiratory burst, monocyte count, neutrophil, total plasma protein and serum bacterial activity against A. hydrophila. Hematocrit and leukocrit did not affected by the addition of Na-alginate, multivitamin combination. The most effective dose of Na-alginate and multivitamin to enhance non-specific humoral immune on tilapia are 2 and multivitamin 1,25 g/kg diet, This results give a new perspective in application of Na-alginate and multivitamin for fish health management.

Keywords: humoral non-specific, cellular non-specific, phagocytic, agglutination, serum bacterial activity.

FEEDING PELLET ENRICHED WITH BACTERIA OVER-PRODUCING HEAT SHOCK PROTEINS (HSPs) MODULATES THE SYNTHESIS OF mRNA ENCODING IMMUNE-RELATED PROTEINS IN WHITE LEG SHRIMP, PENAEUS VANNAMEI JUVENILE

Saranya Sinnasamy¹, Noordiyana Mat Noordin², Muhd Danish Daniel Muhammad¹_, Yeong Yik Sung^1*

¹Institute of Marine Biotechnology, University Malaysia Terengganu (UMT), 21030, Kuala Terengganu, Malaysia

²School of Fisheries and Aquaculture Sciences, University Malaysia Terengganu (UMT), 21030, Kuala Terengganu, Malaysia

^*Corresponding author: yeong@umt.edu.my

Feeding aquatic animals bacterial-encapsulated heat-shock proteins (HSPs) might be potential way to prevent vibriosis, a serious disease that affects aquatic animals used in aquaculture. Shrimp pellets were formulated and enriched with Escherichia coli overexpressing DnaK-DnaJ-GrpE, the prokaryotic equivalents of HSP70-HSP40-HSP20, and the stability of bacterial HSPs on the pellet was assessed during storage. Pellets coated with bacterial HSPs were fed to juveniles of the white leg shrimp Penaeus vannamei to test their resistance to pathogenic Vibrio. Maintaining pellets at different temperatures for variable amounts of time reduced the quantity of bacterial HSP70 on the pellet, suggesting that storage adversely effected the stability of bacterial HSP70 on the pellet. P. vannamei being fed E. coli cells enriched pellets did not affect their survival, demonstrating that the bacteria were not harmful to shrimp, but it did improve survival against Vibrio, indicating that bacterial heat shock proteins play an important role in Vibrio tolerance. The immunological response of this Penaied shrimp species after feeding with bacterial HSPs-enriched pellets was examined to better understand how protection develops. Bacterial HSPs intake influenced the expression of hemolymph mRNAs encoding penaedin, peroxinectin, crustin, transglutaminase, and prophenoloxidase, all of which are immune-related proteins required for P. vannamei to resist infection. After feeding bacterial HSPs, the production of mRNAs encoding all five immune-related proteins was regulated in hemolymph. This study's findings offered a fundamental knowledge of how bacterial HSPs work in the shrimp's innate immune system, enabling for the future development of viable Vibriosis-control techniques in aquaculture.

Keywords: Heat shock protein; shrimp; P. vannamei; immunity; Vibrio tolerance

OZONE GAS EXPOSURE EFFECTS ON VIBRIO LOAD IN THE BRINE SHRIMP ARTEMIA

Asmidar Mohamad1, 2, *Yeong Yik Sung1

1Institute of Marine Biotechnology, Universiti Malaysia Terengganu, 21030 Kuala Nerus, Terengganu, Malaysia

2Institute of Tropical Aquaculture and Fisheries, Universiti Malaysia Terengganu, 21030

Kuala Nerus Terengganu, Malaysia

*Corresponding author : yeong@umt.edu.my, asmidarmohamad18@gmail.com

Artemia, often known as brine shrimp, is an essential live food used in aquaculture. Although Artemia are excellent, there are certain drawbacks to utilising them as food. Artemia's typical feeding properties, including as non-selective filter feeding, allow for the ingestion of bacterial Vibrios that are harmful to aquatic species. In this context, finding an efficient sterilising procedure to eliminate and minimise Vibrio load in Artemia is essential. Several disinfection techniques have been explored so far, including UV irradiation, formaldehyde, ozonation, and bioactive chemicals. The effectiveness of ozone gas in reducing Vibrio load in Artemia was investigated in this study, which further examined at their impact on survival. Ozone exposure did not result in death, demonstrating that this gas was not lethal to Artemia. Agar plating demonstrated that a 5 minute exposure reduced the Vibrio load in Artemia from 61.0x10⁻³ CFU to 3.5x10⁻³ CFU. The elimination of Vibrio was boosted further by increasing the exposure period, with 1.0x10⁻³ CFU remaining after 30 minutes of exposure, the maximum time examined in this study. These findings suggest that exposing Artemia to ozone gas is a viable strategy for reducing Vibrio load, and further improvement of the procedure may aid in the production of a Vibrio-free Artemia product for use in aquaculture.

Keywords : Artemia biomass, ozonization, Palm kernel cake, Vibrio      

LYMPHOCYSTIS DISEASE VIRUS INFECTION ON ORANGE CLOWNFISH AMPHIPRION PERCULA  IN BATAM INDONESIA

Murwantoko¹, Endang Wijayanti², Sri Agustatik², Harshelly Valianti², Lalla Kumala Yulanda¹,

Nur Lailatul Fitrotun Nikmah¹

1. Department of Fisheries Faculty of Agriculture Universitas Gadjah Mada  Flora  Bulaksumur Yogyakarta 55281  INDONESIA
2. Batam Mariculture Development Center, Batam, INDONESIA

Email : murwantoko@ugm.ac.id

The clown anemone fish ‘Nemo’ is the most popular fish species in the global marine ornamental trade. As an ornamental fish, the appearance of the body is important. There is case  several orange clown fishes Amphiprion percula  at Batam Mariculture Development Center showed the presence of wart-like nodules on fins and looked unaesthetic. This study aims to detect the disease agent on the clownfish. The wart-like nodule tissues were collected and fixed in ethanol. The DNA was extracted from the tissues and used as template for PCR using the Lymphocystis disease virus (LCDV) major capsid protein (MCP), myristylated membrane protein (MMP) and DNA Polymerase (DNAPol) primers. The nucleotide sequences of the PCR products were analyzed using Basic Local Alignment Search Tool (BLAST) to look at the homology with data in Genbank. Multiple alignment of the amino acid sequence of MCP was performed using MEGA to determine the genotype.  The PCR produced the expected size products on the detection of MCP, MMP and DNAPol genes. Those results indicated the presence of Lymphocystis disease virus (LCDV) and designated as LCDV-Oc-Btm.  The sequencing could determine 1221 nucleotides of MCP and 407 nucleotides of MMP genes. The BLAST analysis from those sequences showed the highest homology with the species of LCDV-1 (LCDV-Sa strain) at 91.04 % and 88.19 % for MCP and MMP respectively.  The UPGMA phylogenetic tree showed LCDV-Oc-Btm is differ from the existing genotype.

Keywords : genotype, LCDV, major capsid protein, myristylated membrane protein, wart-like nodule

COMPARATIVE HEALTH ANALYSIS BETWEEN LIMNOGONUS SP. AND ALPHEUS SP., AS SENTINEL SPECIES FROM THE SEAGRASS MEADOWS, LIBONG ISLAND, THAILAND

Nisreen Dahlan1,*, Chanyut Sudtongkong1, Pornthep Wirachwong1, Supparat Kong-Oh1, Narit Thaochan2, Ezra Mongkolchaichana3, Nattawut Charoenphon4, Gen Kaneko5, Piyamat Kongtueng6, Sinlapachai Senarat1

1Department of Marine Science and Environment, Faculty of Science and Fisheries Technology, Rajamangala University of Technology Srivijaya, Trang, Thailand Agriculture 2Innovation and Management Division (Pest Management), Faculty of Natural Resources, Prince of Songkla University, Hat Yai, Thailand

3Department of General Education, Faculty of Science and Health Technology, Navamindradhiraj University, Bangkok, Thailand

4Department of Anatomy, Faculty of Medical Science, Naresuan University,Thailand 5College of Natural and Applied Science, University of Houston-Victoria, Victoria, Texas, USA

6Central Laboratory, Faculty of Veterinary Medicine, Chiang Mai University, Chiang Mai, Thailand

*Corresponding author: nisreen4062@gmail.com

The regenerated seagrass beds in Libong Island, Thailand, which presently is considered as a habitat of a large group of dugongs, is rapidly decreasing. Monitoring of marine species living in this habitat is necessary, but such studies are missing. We hence selected two common species Pleciogonus sp. as a pelagic species and Alpheus sp. as a benthic species, and used them as sentinel species to assess the environmental health of the seagrass meadows in Libong island. Their comparative health analysis was conducted using the accurate histopathological methods using several biomarkers. During January to March 2021, all samples were collected from the healthy and unhealthy conditions of seagrass areas of Libong island. Histological observation demonstrated that all body systems were normal in Pleciogonus sp. irrespective of sampling site; however, some histopathological alterations were seen in Alpheus sp. including the gill disorganization and the hepatopancreatic degeneration, especially those from unhealthy seagrass areas. The observed abnormalities in Alpheus sp. might be related to environmental stresses, which may be more severe in the benthic than pelagic habitat in the seagrass area. The present study also showed that the above aquatic animals can be used as a sentinel species with certain histopathology biomarkers to assess the environmental health.

Keywords: Aquatic animal health, Histopathology, Seagrass, Thailand