Effect of Dietary Supplement of Probiotic (Lactobacillus plantarum) on Growth Performance, Feed Utilization and Survival Rate in Bocourti Catfish (Pangasius bocourti Sauvage, 1880)

This study was investigated the effect of Probiotic (L. plantarum) supplementation at 0 , 1x107,1x108,1x109 cfug-1 diet on growth performance, feed utilization, survival rate and carcass quality in Bocourti’s catfish (Pangasius bocourti). An average initial weight of fish 132.59 ±1.20 gfish-1were divided into four treatments, three replication and fed for 8 weeks. The results showed that body weight, weight gain, daily growth rate, specific growth rate, survival rate, feed conversion ratio and feed conversion efficiency were not significant difference in all groups (P>0.05).However, Fish fed dietary supplementation of probitic at 1x107 cfug-1 diet had trendy high in body weight , weight gain, daily growth rate and specific growth rate with 207.16 ± 16.31 g., 74.30 ± 15.13 gfish-1, 1.33 ± 0.27gday-1 and 0.79 ± 0.13%day-1respectively. The biochemical parameters in serum and carcass quality were did not significantly different for any of the diet treatment. Therefore, this study indicated that the optimum level of L. plantarum in diet for bocourti catfish culturing could be used for contains 1x107 cfug-1 diet.

Bocourti catfish or Basa or Pla-mong (Thai name) is common name of Pangasius bocourti Sauvage 1880 that number is Pangasiidae. This fishes are native species in Mekong and Chao Phraya basins in Indochina. In Thailand found in Mae Kong river, Chao Phraya river and some other smaller river such as Mun river and Chi river and other river in Northeastern in Thailand 23, 25 . This fish has a good taste and has been accepted of consumers and the demanded of globe has been increased in America and Europe 15 . In both foreign markets are mostly demanded in filet forms and has high price. In Thailand markets had price depended on size of fish, e.g, 0.7-1.0 kg had about up to 50 baht and 1.5 to 2.0 kg had about 150 baht 21 .
Recently, Aquaculture is important sector and spreading area to providing human consumption and fisheries. The growing of aquaculture practices that from many factor :1) the demand of market because protein form the aquatic animals is good quality, 2) the security and safety food of consumer and 3) make income from aquaculture 5 . However, aquaculture practices in intensive system requires cultivation at high densities, which has caused destroyed the environment because the effluent water from the ponds containing organic wastes in high volume, effect on water quality such as reducing dissolved oxygen, giving the ammonia, nitrites and hydrogen sulfide from the metabolites of organics degradation that often are responsible for mortality 27 . And, under these conditions of intensive system, aquatic animals has to high-stress, that easily infection by pathogens and high mortality rate and causing a decreasing productivity 3 . So, the owner of farm using antibiotic drug for preventing and treating which has seen the previous research that can be used to prevent infection. But, the using antibiotics will kill the bacteria in the digestive tract of aquatic animals and the accumulation of antibiotic residues in fish products to be harmful for human consumption. Form the this problem, the farmer has turned to use probiotics in aquaculture that it is non-pathogenic microorganisms in aquatic animals that are many proportion , such as improved growth of fish , preventing the pathogen , increasing utilization of nutrient , treat the water quality in pond, reduce stress of fish and improved reproductive system of aquatic animals 5 .
Probiotic is a relatively new term which is used to name microorganisms that are associated with the beneficial effects for the host. Probiotics have been introduced as a growth stimulant. Such as the study of Lara et al. 17 that use of Streptococcus as growth promoters on Nile tilapia (Oreochromis niloticus) the result showed probiotic strain ,increasing significantly the content of crude protein and crude lipid in the fish, also weight has increased from 0.154 g to 6.164 g. in 9 weeks of culture. And ornamental fishes include swordtail (Xiphophorus helleri, X. maculatus) and guppy, (Poecilia reticulate, P. sphenops), their feed was supplemented with Bacillus subtilis and Streptomyces, can be increases in growth and survival of Xiphophorus and Poecilia after 90 and 50 days of experimental period, respectively 7,12 .
Lactic acid bacteria (LAB) are most applied probiotics in for use enhanced growth or immune system for aquatic species 8,9,24 . And they are producers of bacteriocins and organic acids (lactic and acetic acids) which have inhibitory effects in vitro on the growth of some pathogens in fish 20 . And they have the previous study for use enhanced growth or immune system in fish . e.g Nile tilapia,Oreochromis niloticus 1 , Grouper, Epinephalus coioides 26 , Cobia, Rachycentron canadum 11 and European sea bass, Dientrachus labrax 19 . Therefore, The aim of this study was to determine the effect of Lactobacillus plantarum supplementation on growth performance, survival rate, serum biochemical and carcass quality in Bocourti catfish (Pangasius bocourti Sauvage, 1880)

Fish, diets and feeding protocol
This study was carried out with the used four different levels of Lactobacillus plantarum ( 0, 1x10 7 , 1x10 8 and 1x10 9 cfug -1 diet ). A Completely Randomized Design (CRD) with three replication. The basal diet was developed from formula diet of Boonarsa and Doolgidachbaporn 4 and the nutritional value of basal diet had a crude protein and gross energy with 29.89% dry matter and 424.65 kcal per 100 g diet( Table 1). The experimental diets were prepared by mixing the dry ingredients with soybean oil and water by hands and then the wet dough was placed in a grinder and pelleted by mincer to pass through a 2-mm and finally the moist pellets were dried in hot air oven at 60 °C for 12 hr and then stored in the fridge at -20 0 C for further uses.
Lactobacillus plantarum CR1T5 were provided from Department of Microbiology, Faculty of science, Khon Kaen Unversity, Thailand. The bacterial suspension content was about 3.4x10 9 cfuml -1 and the bacterial were prepared stocking every twice a weeks. After that diluted the bacteria in concentration of experimental by used sterilized MRS broth is dilutant. The dilution of bacteria sprayed on basal diet with 0, 1x10 7 , 1x10 8 and 1x10 9 cfug -1 diet that test diets were daily prepared.
Bocourti catfish (P. bocourti) juvenile obtained from Phayao Inland Fisheries Research and Development Center, Phayao Province, Thailand . Fish were kept in 1000L fiber tank for acclimatization with commercial diet with 40% protein for four weeks. Then protein content in diet was decreased to 30 and fed for 8 weeks prior to feeding the experimental diets. After the acclimatization fish were randomly distributed into four groups with three replications that each replicate contained 10 fish (mean wet weight of 132.59±1.20 gfish -1 ) in an aquarium ( 60 x 150 x 60 cm 3 ) filled the water 450 L. which 12 aquaria tanks. Water quality parameters such as dissolved oxygen (6.05±0.28 mg L-1), temperature (26.98±0.19°C) and pH value (7.37±0.27) were measured every day during the experimental period. NIAMPHITHAK et al.: EFFECT OF DIETARY SUPPLEMENT OF PROBIOTIC During the trial, fish were fed ad-libitum by hand fed method twice a day (08:00 and 15:00 h) for 8 week. Feed consumption was recorded weekly and fish from each tank were weighed to measure growth every two weeks until the end of the experiment; growth performance and feed utilization were calculated.

Serum biochemistry and carcass quality
At the end of the experiment, after fish were starved for 12 hours, randomized two fish per tank that were anesthetized with tricaine methanesulfonate (MS-222). And blood samples were collected from caudal vein that kept in nonheparinized tubes.Hematocrit was determined by using microhematocrit centrifuge. The serums were separated into aliquots and analyses of serum biochemistry. Serum was analyzed for total protein (TP) and alkaline phosphatase (ALP) activity. The serum biochemical indices were determined with an automatic biochemical analyser (BS-200; Mindray,Shenzhen, China). Then the fishes were killed and liver and visceral were keeped and test investigate weighed carcass quality by Hepatic somatic index (HSI), Visceral Somatic index (VSI), Visceral Fat index ( VFI) and Muscle ratio (MR) by equation following: HSI (%) = (Liver mass(g)/ body mass (g)) × 100 VSI (%) = (total inner organ mass (g) / body mass (g)) × 100 VFI (%) = (Visceral Fat mass (g)/ body mass (g)) × 100 MR (%) = muscle mass (g)/ body mass (g)) × 100

Statistical analysis
The data were subjected to one-way analysis of variance (ANOVA) and if significant differences (p<0.05) were found, Duncan's multiple range test was used to rank the groups.

Growth performance, feed utilization and survival rate
The growth performance, feed utilization and survival rate of bocourti catfish (P.bocourti) with an initial average weight of 132.59 ± 1.20 gfish -1 fed with diet containing Lactobacillus plantarum at 0, 1x 10 7 , 1x10 8 and 1x10 9 cfug -1 diet for 8 weeks is shown in Table 2. At the end of experiment found that growth performance base on the weight gain (WG), average daily gain (ADG) and specific growth rate (SGR) and feed utilization base on feed conversion ratio (FCR) and feed efficiency ratio (FCE) were not significant different (p>0.05). But fish fed with Lactobacillus plantarum at 1x 10 7 cfug -1 diet had maximum growth performance and best feed utilization compared with fish in other group. The weight gain (WG), average daily gain (ADG), specific growth rate (SGR), feed conversion ratio (FCR) and feed efficiency ratio were ranged from 64.30 ± 9.66 to 74.30 ± 15.13 gfish-1. 1.15 ± 0.17 to 1.33±0.27 dday-1, 0.70 ± 0.09 to 0.79 ± 0.13 %day -1 , 1.91 ± .41 to 2.17 ± 0.31 and 46.76 ± 7.04 to 54.06 ± 11.01 %, respectively. And survival rate was 100 % in all groups.

DISCUSSION
This study was evaluated the effect of using Lactobacillus plantarum levels on Growth performance, Survival rate, Serum biochemical and Carcass quality of bocourti catfish (Pangasius bocourti Sauvage, 1880). From the present study, the growth performance and feed utilizations of fish were not significantly different between groups. But, the growth and feed utilization of fish fed with L. plantarum at 1x10 7 cfug -1 diet was better than other groups. Therefore, it can be concluded that giving optimal level of probiotics improved growth of fish which was consistent with study of Piccolo et al. 19 that use L. plantarum in diet of European sea bass (Dientrachus labrax) and no did not significantly different between groups. However the additions L. plantarum to Means within row followed by a common letter are not significantly different according to DMRT (P < 0.05) diet increased growth in term of weight gain, specific growth rate and protein efficiency ratio were 3.6, 5.2 and 6.9 % respectively. Lara et al. 17 reported that use of Streptococcus sp. as growth promoters on Nile tilapia (Oreochromis niloticus) increasing significantly the content of crude protein and crude lipid in the fish, also weight has increased from 0.154 g to 6.164 g. in 9 weeks of culture. Similar resulted were conducted in swordtail (Xiphophorus helleri, X. maculatus) and guppy, (Poecilia reticulate, P. sphenops), their feed was supplemented with Bacillus subtilis and Streptomyces sp. can be improved in growth and survival of swordtail and guppy after 90 and 50 days of experimental period, respectively 7,12 . These effects have been reported a similar response and positive effect on productive performance in other species. e.g. Alatic cod, Gadus morhua 13 , Juvenile Dentex, Dentex dentex L. 14 .
The carcass quality of fish in these study found that hepatosomatic index (HSI), vesicle somatic index (VSI) vesicle fat index (VFI) and muscle ratio (MR) were not significantly different between groups (p>0.05). Likely, the study of Dawood et al. 6 that used L. rhamnosus and Lactococcus lactis supplement in diet of red sea bream(Pagrus major) at 10 6 cell g -1 was found the hepatosomatic index (HSI) and somatic index (VSI) were not significantly different (p>0.05) which was consistent with study of Piccolo et al. 19 that use Lactobacillus plantarum in diet of European sea bass (Dientrachus labrax).
Hematocrit has been recognized as valuable tools for monitoring of fish health 2,18 . From the present study the hematocrit value of fish from all groups and has significantly different(p<0.01). And the trend of hematocrit increased on probictics levels in diet. The results indicated that supplement of probiotic imporved healthy of fish. The blood protein, albumin and globulin are the major proteins, which play a significant role in the immune response 22 . From this study showed not significantly different of total protein in fish serum. Because when liver is damaged, it will total protein, albumin and globulin in serum decreased 16 . The alkaline phosphatase (ALP) in these study was not significantly different. which was consistent with study of Piccolo et al. 19 that use Lactobacillus plantarum in diet of European sea bass (Dientrachus labrax). However, the trend of ALP increased on supplementary Lactobacillus level in diet of bocourti catfish. From the reported of Piccolo et al. 19 that highest values of ALP in fish resulted of increased growth rate and feeding level.

CONCLUSION
In conclusion, this present study indicated that supplementation of different concentrations Lactobacillus plantarum in diet of Bocourti catfish (P. bocourti) were not significantly different in growth performances, feed utilizations, carcass quality and serum biochemistry. The haematocrit values was increased with increased concentration of probiotics. Base on these results, use of Lactobacillus plantarum at 1x 10 7 cfug -1 supplementation in diet had maximum growth performance and best feed utilization.