Patent
Indigenous probiotic bacteria for application in pacific white shrimp culture
Description:Registration Date:November 10, 2013<br><br>With collaboration: Babak Ghaednia
Text:

Shrimp culture and rearing industry in particular white western shrimp species (Litopenaeusvannamei) is one of the major activities in the world, including Iran. The outbreaks of diseases are the main factors limiting production. Due to the adverse effects of abusing antibiotics and chemical disinfectants, disease control and prevention requires novel methods that are affordable, effective and safe for the environment and humans. For this purpose during the five months, sampling was done from three main sites of shrimp culture in Bushehr province and 150 pieces cultured white western shrimp, 135 water and sediment samples were collected from the ponds, input and output channels by standard methods. Physicochemical parameters of water and biometry of shrimps were done and recorded. Probiotic isolation was performed by culturing samples in Tryptic soy agar and TCBS after incubation in 30º C for 24-48 h. The antimicrobial effects of culture extract of isolates were evaluated againstV. harveyi with the well diffusion method and the best isolates were selected. The molecular identification of selected bacteria was performed by 16S ribosomal DNA gene sequence analysis technique. The growth kinetics of selected bacteria and effect of environmental factors on antimicrobial compound production and stability of them in salinity (1.5-5.5%) and (0-50ppt), pH (5-9) and temperature (30-40º C) and (35-100º C) were studied respectively.

After the determination of antibiotic resistance profiles of them, the In vivo test on theL. vannamei post larvae was performed and effect of these bacteria on growth performance, prevention of vibriosis and water quality include (ammonia, nitrate nitrite, turbidity, total bacteria andVibrionaceae load were examined. For extraction and identification of bioactive material with anti-bacterial properties produced by selected bacteria, dialysis, SDS-PAGE electrophoresis, ultraviolet spectroscopy and gas chromatography mass was performed.

According to the results of this dissertation the most frequency of facultative aerobic and anaerobic heterotrophic bacteria was seen in shrimp digestion tract (3.04 ± 0.75) ×105CFU g-1 in September.GenusVibrio spp. (37.88%) andBacillus spp. (27.27%) had the most frequency respectively.

Of the 198 bacterial isolates, two bacterial strains from the digestive tract and sediment which had the highest inhibition and a stability antagonistic effect onV. harveyi were selected. Based on 16S ribosomal DNA gene sequence analysis, they were identified asBacillus subtilis subsp. inaquosorum strain IS02 (GenBank: JN856456.1) andBacillus vallismotis IS03 (GenBank: JQ085958.1) and recorded in the National Center for Biotechnology Information (NCBI). After exposure ofL. vannamei post larvae (mean weight 0.3 ± 0.15 g) by selected bacteria, the mean values of final weight, DWG, RGR and SGR in all groups treated with probiotics were significantly higher than those of the Control (P<0.05). Accumulative mortality percent in treatments were significantly lower than controls (p< 0.05). The results indicated selected bacteria were effective in vibriosis prevention, reducing ammonia, turbidity, total bacteria andVibrionaceae family load in the water.

Overall, according to results both of the selected bacteria were adapted to the ecological conditions of shrimp culture and could be used as appropriate probiotics in this industry. It is hoped that the results of this research is to produce a step towards the realization of national production of indigenous probiotic in Iran.

Author:Maryam Mibakhsh
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