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Kasetsart J. (Nat. Sci.) 40 : 181 - 187 (2006) The Effect of Peptidoglycan on Immune Response in Black Tiger Shrimp (Penaeus monodon Fabricius) Watchariya Purivirojkul 1 *, Nontawith Areechon 2 and Prapansak Srisapoome 2 ABSTRACT The effect of peptidoglycan on immune response in Black Tiger Shrimp (Penaeus monodon Fabricius) was conducted under laboratory conditions. P. monodon (13±2 g) were fed for 7 days with diets containing graded levels of PG (0, 0.06, 0.12 and 0.18 g/kg feed). The immune responses were measured by phenoloxidase, superoxide anion, bactericidal activity, clearance ability and resistance against Vibrio harveyi. The results showed that shrimp which fed on diets containing PG 0.18 g/kg had these immune parameters at significantly higher (P
182 approach to increase immunity to defend shrimp against diseases. Peptidoglycan (PG), derived from the cell wall of gram positive bacteria, was reported to be used as an immunostimulant in aquaculture. It was found to increase survival rate and enhance protection against pathogens in shrimp (Soderhall and Cerenius, 1992; Boonyaratpalin and Boonyaratpalin, 1995; Sung et al., 1996; Itami et al., 1998; Vargas-Albores et al., 1998) Although peptidoglycan can enhance immunity of shrimp, shrimp farmers need to use it wisely. If shrimp farmers use excess amount of PG supplement in feed, it may waste a lot of money. This research was conducted to determine the optimum concentration and time for effective use of PG which could be recommended for shrimp aquaculture. MATERIALS AND METHODS Experimental animals The test shrimps were obtained from a farm in Chachoengsao Province, Thailand. Shrimps with approximate weight of 11-15 g per animal were used. They were acclimatized in a 45 × 60 × 45 cm 3 aerated aquarium system filled with 25 ppt chlorinated sea water and changing of water was made everyday. Twenty shrimps per replication were used. Experiment 1 Determination of optimum concentration Immune parameters Four treatments consisted of commercial shrimp pelleted feed mixed with PG at 0, 0.06, 0.12 and 0.18 g/kg feed, top-dressing with squid oil, were used in the experiment. Shrimps were fed five times daily at 3% body weight per day. The immune parameters, consisted of phenoloxidase, superoxide anion, bactericidal activity and clearance ability were measured after 7 days of the feeding trial. Kasetsart J. (Nat. Sci.) 40(1) Experiment 2 Determination of optimum time of application The results from experiment 1 were evaluated to choose only one suitable concentration for this experiment. Four treatments consisted of control, feed with PG applied 3, 5 and 7 days/week were used in the experiment. Shrimp were fed five times daily at 3% body weight per day. Shrimps were continuously fed with PG for 4 weeks, following by normal feed for 4 weeks and PG for 2 weeks, respectively. Immune parameters consisted of phenoloxidase, superoxide anion and bactericidal activity were measured every week for 10 weeks. All results were evaluated to establish the standard protocol for the application of peptidoglycan for Penaeus monodon culture including the concentration and the time of application. Immune parameters Preparation of hemolymph samples Blood sample of 0.5 ml from each sample was withdrawn from the base of 3 th walking leg of the shrimp by a syrynge containing 1.5 ml anticoagulant (K-199 + 5% L-cysteine). 1) Phenoloxidase activity assay The method was modified from Supamattaya et al. (2000). After the blood was withdrawn, the hemocytes were washed three times with shrimp saline (1,000 rpm 4 °C 10 min). Hemocyte lysate (HLS) was prepared from hemocytes in a cacodylate buffer pH 7.4 by using the sonicator at 30 amplitute for 5 second and the suspension was then centrifuged at 10,000 rpm., 4°C for 20 min. The supernatant was collected as HLS. Then 200 µl of 0.1% trypsin in cacodylate buffer was mixed to the 200 ml HLS followed by 200 µl of Ldihydroxyphenylalanine (L-DOPA) at 4 mg/ml as the substrate. Enzyme activity was measured as the absorbance of dopachrome at 490 nm.
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January - March 2006 Volume 40 Numb
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KASETSART JOURNAL NATURAL SCIENCE T
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In sacco Degradation Characteristic
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2 and management practices a sorghu
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4 the greater plant height was obta
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6 at Wolenchity (Figure 3) that led
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8 Stover and aboveground biomass Co
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10 Radford, B.J., A.J. Dry, L.N. Ro
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12 At present, new cultivars with h
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14 2000), grapevine (Lavee, 2000),
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16 repeatability of FW, FLT, FLW, S
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18 (Table 5) indicating that select
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Kasetsart J. (Nat. Sci.) 40 : 20 -
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22 measured to define fruit shape i
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24 for higher fruit number and yiel
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28 (Table 2). The numbers of flower
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30 to the color of fruit and seed c
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32 1999. Carrots and related vegeta
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34 MATERIALS AND METHODS Laboratory
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36 DISCUSSION Extensive studies hav
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38 from Gossypium hirsutum. J. Inse
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40 The natural resistance of plants
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42 4. Data collection and statistic
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44 was not significantly different
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46 showed the highest yield compare
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48 Kessmann, H., T. Staub, C. Hofma
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50 There are approximately 60 comme
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52 in PBST containing 2% ovalbumin
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54 Kasetsart J. (Nat. Sci.) 40(1) T
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56 Virus-free orchid plants show fa
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60 feed source was based on purchas
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62 intake of DM, CP, ether extract
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64 relatively slow and short. The p
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66 week 4. Milk composition was not
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68 Agricultural Experiment Station.
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70 et al., 2002). In contrast to ma
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72 level and showed marked polymorp
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76 loam (Eutric Fluvisol) soil and
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78 Eugenia caryophyllata, Echinops
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80 CONCLUSION In conclusion, out of
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82 of chromic acid titration method
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84 initials at the time of pinning
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86 vessels a point where the number
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88 Kasetsart J. (Nat. Sci.) 40(1) T
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90 CONCLUSIONS In the investigation
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92 1994; Strand et al., 1997). Now
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94 The clones were grown overnight
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96 isozymes. The copy number for nu
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98 Helianthus annuus. Theor. Appl.
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100 MATERIALS AND METHODS Plant mat
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102 The amount of Na + (%) 6 5 4 3
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104 NGE, 7-16 folds). Finally, all
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106 Anal. Biochem. 162: 156-159. Cr
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108 grown for academic purpose at N
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110 Cluster A was further separated
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112 Table 2 Similarity index of 19
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114 subjected to UPGMA and resultin
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116 Tabel 3 The similarity index of
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118 formation as seen from phylogen
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120 Genet. 101: 860-864. Lerceteau,
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122 INTRODUCTION Methylotrophic yea
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124 RESULTS AND DISCUSSION Screenin
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126 optimum temperature, insignific
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128 medium of C. boidinii (Volfova
Page 135 and 136: 130 was observed soon after 24h cul
Page 137 and 138: 132 Kasetsart J. (Nat. Sci.) 40(1)
Page 139 and 140: 134 ACKNOWLEDGEMENTS This work was
Page 141 and 142: Kasetsart J. (Nat. Sci.) 40 : 136 -
Page 143 and 144: 138 Determination of enzyme activit
Page 145 and 146: 140 Table 1 Optimum and stability o
Page 147 and 148: 142 for example recombinant E. coli
Page 149 and 150: 144 were studied for β-1,3-1,4-glu
Page 151 and 152: 146 ACKNOWLEDGEMENT This work was s
Page 155 and 156: 150 vulcanization times were calcul
Page 157 and 158: 152 RESULTS AND DISCUSSION Mechanic
Page 161 and 162: 156 blend with 0.5 phr Irganox. How
Page 165 and 166: 160 (AA-680 ShiMADZU, Atomic Absorp
Page 167 and 168: 162 warm, relatively shallow and hi
Page 169 and 170: 164 cycle in crustacean (Chen et al
Page 171 and 172: 166 Salaenoi, J. 2004. Changes of e
Page 173 and 174: 168 MATERIALS AND METHODS This rese
Page 175 and 176: 170 DISCUSSION The average total am
Page 179 and 180: 174 repacked and scanned three time
Page 181 and 182: 176 found the most in the shrimp fe
Page 183 and 184: 178 PLS model The comparison betwee
Page 185: 180 brix value and dry matter of ma
Page 189 and 190: 184 higher (P
Page 191 and 192: 186 levels were significantly eleva
Page 195 and 196: 190 currents. Zone 4: River outlet
Page 197 and 198: 192 standing waters, thus the occur
Page 199 and 200: 194 and moving to 59%TL in juvenile
Page 203 and 204: 198 thin membrane (Figure 1A, B). T
Page 205 and 206: 200 were clearly seen. The presence
Page 207 and 208: 202 could change to female at 6-12
Page 211 and 212: 206 water until the water became cl
Page 213 and 214: 208 reported by Thu and Preston (19
Page 215 and 216: 210 cavalcade hay. The crude protei
Page 217 and 218: 212 CONCLUSIONS Ruminal disappeared
Page 219 and 220: 214 ∅rskov, E.R. and I. McDonald.
Page 221 and 222: 216 is constrained mainly due to fa
Page 223 and 224: 218 SNF and fat composition in raw
Page 225 and 226: 220 in terms of both manual samplin
Page 227 and 228: 222 Harding, F. 1995. Compositional
Page 229 and 230: 224 proteins described by Barbut an
Page 231 and 232: 226 starch/flour, shaken vigorously
Page 233 and 234: 228 % Stress relaxation 54 49 44 39
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234 categories including puffed sna
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236 Cohesiveness of mass Crispness
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238 Cohesiveness of mass Sweetness
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242 3. Physicochemical properties 3
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244 Kasetsart J. (Nat. Sci.) 40(1)
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246 ACKNOWLEDGEMENTS This research
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248 are now becoming more and more
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250 downstream control. Lam Chae re
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254 Inflow (mcm.) Inflow (mcm.) Inf
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256 were above 0.70. The testing re
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258 1999-2000 could be used for tra
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262 we assume that r ≥ 2 and d r
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266 Figure 1 Prototype domains. num
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268 DISCUSSION Consequences indicat
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272 George, B. K. 1997. The GIS Boo
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discussed; and appropriateness of t
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LITERATURE CITED FORMAT Manuscripts
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PRE-SUBMISSION CHECKLIST (see “In
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Send application materials to Payme
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