Bacterial Population in Intestines of the Black Tiger Shrimp (Penaeus monodon) under Different Growth Stages

“The sequence data was processed using Mothur software to remove low quality sequences [69].”

“When sequences were classified into taxonomic ranks using the Ribosomal Database Project (RDP) Bayesian classifier with a confidence threshold of 80% [40], phylum distribution of bacteria in intestines at four growth stages varied from 3 to 6 phyla.”

“PCR-DGGE profiles of intestine samples from four shrimp stages were performed according as previously described with some modifications [20,75,76].”

“To provide comprehensive coverage of bacterial communities, a highthroughput pyrosequencing method has been employed to obtain a higher number of sequence-reads [23].”

“The method has recently gained popularity for an in-depth characterization of various microbial communities such as those in the human [24,25,26], fermented food [27], soil [28,29,30,31], extreme environments [32], the deep marine biosphere [33,34], wastewater treatment bioreactors, [35] and plant biospheres [36,37].”

“The method has recently gained popularity for an in-depth characterization of various microbial communities such as those in the human [24,25,26], fermented food [27], soil [28,29,30,31], extreme environments [32], the deep marine biosphere [33,34], wastewater treatment bioreactors, [35] and plant biospheres [36,37].”

“The coverage index was calculated by 1– (n/N), where n is the number of phylotypes and N is the total number of sequences [41].”

“in less harsh condition for bacteria to reside within [60,62], resulting in different intestinal bacterial diversity from the harsher conditions in the juveniles.”

“The method has recently gained popularity for an in-depth characterization of various microbial communities such as those in the human [24,25,26], fermented food [27], soil [28,29,30,31], extreme environments [32], the deep marine biosphere [33,34], wastewater treatment bioreactors, [35] and plant biospheres [36,37].”

“The intrinsic developmental changes in transition to later growth stage may exert selective pressure for those highly adapted bacteria to host gut habitat [64,65].”

“0) [43,44], and mouse (5.”

“The method has recently gained popularity for an in-depth characterization of various microbial communities such as those in the human [24,25,26], fermented food [27], soil [28,29,30,31], extreme environments [32], the deep marine biosphere [33,34], wastewater treatment bioreactors, [35] and plant biospheres [36,37].”

“The intestinal tract is a complex ecosystem that harbors a diverse bacterial community and this bacterial population has been shown to have profound influence on immunity, nutrient processing and protective processes in the host animal [1,2,3,4,5,6].”

“Detection of bacterial taxa by real-time PCR assay and primer specificity was previously developed [73,74].”

“Operational taxonomy units (OTUs) were clustered by Mothur using the average neighbor algorithm at 0.03 dissimilarity level to classify to OTUs [71].”

“This family of bacteria are widely distributed in marine environment [50] and has previously been reported to be found in high relative abundance in intestine of various aquatic marine organisms including the black tiger shrimp [21,22,48,49,51].”

“The method has recently gained popularity for an in-depth characterization of various microbial communities such as those in the human [24,25,26], fermented food [27], soil [28,29,30,31], extreme environments [32], the deep marine biosphere [33,34], wastewater treatment bioreactors, [35] and plant biospheres [36,37].”

“The intestinal tract is a complex ecosystem that harbors a diverse bacterial community and this bacterial population has been shown to have profound influence on immunity, nutrient processing and protective processes in the host animal [1,2,3,4,5,6].”

“For instance, rarefraction curve analysis of bacterial communities in zebrafish intestines has not been saturated, even when a range of 5,000 to 10,000 sequence reads were obtained [47].”

“0) [43,44], and mouse (5.”

“Bacterial diversity was estimated by Shannon index, in which a higher value means a greater bacterial diversity [42].”

“The microbiota and their influences on immune response, disease resistance, and nutrition for the animal hosts have been intensively studied in many vertebrate model organisms [3,38,39].”

“PCR-DGGE profiles of intestine samples from four shrimp stages were performed according as previously described with some modifications [20,75,76].”

“While some Vibrio species are reported as shrimp pathogens [17,67], there is some evidence suggesting beneficial roles of Vibrio species to increase disease resistance in aquatic animal hosts [21].”

“This factor, in addition to the difference in shrimp diets between different developmental stages might contribute to a higher bacterial diversity post-larval shrimp [5,63].”

“monodon) were hatched at the Shrimp Genetic Improvement Center (Surat Thani province, Thailand) and routinely checked to ensure that they are free from specificpathogens: Taura Syndrome Virus, White Spot Syndrome Virus, Yellow Head Virus, Monodon Baculovirus, and Infectious Hypodermal and Hematopoietic Necrosis Virus…”

“In the zebrafish model, the gut microbial community plays a significant role in digestive tract development [7].”

“Bacteria in intestines of other aquatic animals have also been shown to contribute to host’s immune system development, and the potential application of beneficial bacteria to aquaculture feeds, especially in fish and shellfish larviculture, has been increasingly investigated [8].”

“Many studies in crustaceans, including the black tiger shrimp, focused on the development of probiotic applications to enhance disease resistance and growth [12,13,14].”

“Bacteria associated with animal digestive tract might be beneficial to the host such as natural barriers against pathogen colonization through competition for binding sites [66].”

“This family of bacteria are widely distributed in marine environment [50] and has previously been reported to be found in high relative abundance in intestine of various aquatic marine organisms including the black tiger shrimp [21,22,48,49,51].”

“The intestinal tract is a complex ecosystem that harbors a diverse bacterial community and this bacterial population has been shown to have profound influence on immunity, nutrient processing and protective processes in the host animal [1,2,3,4,5,6].”

“Furthermore, to enable an analysis of multiple samples in parallel within a single run, a barcode tag with a short unique nucleotide sequence was added to distinguish each sample [31].”

“The method has recently gained popularity for an in-depth characterization of various microbial communities such as those in the human [24,25,26], fermented food [27], soil [28,29,30,31], extreme environments [32], the deep marine biosphere [33,34], wastewater treatment bioreactors, [35] and plant biospheres [36,37].”

“Although our DGGE analysis revealed only bacteria genera with high abundance and may not provide an in-depth analysis of bacterial population, this method is proven to be useful for comparison of bacteria community profiles in food and soil samples [27,58,59].”

“The method has recently gained popularity for an in-depth characterization of various microbial communities such as those in the human [24,25,26], fermented food [27], soil [28,29,30,31], extreme environments [32], the deep marine biosphere [33,34], wastewater treatment bioreactors, [35] and plant biospheres [36,37].”

“The intestinal tract is a complex ecosystem that harbors a diverse bacterial community and this bacterial population has been shown to have profound influence on immunity, nutrient processing and protective processes in the host animal [1,2,3,4,5,6].”

“The gut layer in Penaeid shrimp is ectodermal origin, which regularly shreds off during the molting period [54].”

“Shrimp from the post-larval through juvenile stages have been reported to have higher bacterial disease susceptibility than those at the later stages [15,16,17,18].”

“Although our DGGE analysis revealed only bacteria genera with high abundance and may not provide an in-depth analysis of bacterial population, this method is proven to be useful for comparison of bacteria community profiles in food and soil samples [27,58,59].”

“This family of bacteria are widely distributed in marine environment [50] and has previously been reported to be found in high relative abundance in intestine of various aquatic marine organisms including the black tiger shrimp [21,22,48,49,51].”

“However, the immune system of post-larval shrimp is not as developed as in juvenile shrimp [19,62].”

“Traditionally, bacterial populations in the intestines have been characterized by both culture-dependent and culture-independent techniques based on 16S rRNA sequences [20,21,22].”

“While some Vibrio species are reported as shrimp pathogens [17,67], there is some evidence suggesting beneficial roles of Vibrio species to increase disease resistance in aquatic animal hosts [21].”

“Many studies in crustaceans, including the black tiger shrimp, focused on the development of probiotic applications to enhance disease resistance and growth [12,13,14].”

“Although our DGGE analysis revealed only bacteria genera with high abundance and may not provide an in-depth analysis of bacterial population, this method is proven to be useful for comparison of bacteria community profiles in food and soil samples [27,58,59].”

“Shrimp from the post-larval through juvenile stages have been reported to have higher bacterial disease susceptibility than those at the later stages [15,16,17,18].”

“in less harsh condition for bacteria to reside within [60,62], resulting in different intestinal bacterial diversity from the harsher conditions in the juveniles.”

“The method has recently gained popularity for an in-depth characterization of various microbial communities such as those in the human [24,25,26], fermented food [27], soil [28,29,30,31], extreme environments [32], the deep marine biosphere [33,34], wastewater treatment bioreactors, [35] and plant biospheres [36,37].”

“The intrinsic developmental changes in transition to later growth stage may exert selective pressure for those highly adapted bacteria to host gut habitat [64,65].”

“5) [45].”

“Moreover, several studies have shown that Artemia nauplii are one of major vectors to introduce bacteria including pathogens to hatchery system [56,57].”

“This family of bacteria are widely distributed in marine environment [50] and has previously been reported to be found in high relative abundance in intestine of various aquatic marine organisms including the black tiger shrimp [21,22,48,49,51].”

“Moreover, several studies have shown that Artemia nauplii are one of major vectors to introduce bacteria including pathogens to hatchery system [56,57].”

“The microbiota and their influences on immune response, disease resistance, and nutrition for the animal hosts have been intensively studied in many vertebrate model organisms [3,38,39].”

“Once shrimp reached juvenile stages, their diet is solely based on commercial pellets [52,53].”

“The method has recently gained popularity for an in-depth characterization of various microbial communities such as those in the human [24,25,26], fermented food [27], soil [28,29,30,31], extreme environments [32], the deep marine biosphere [33,34], wastewater treatment bioreactors, [35] and plant biospheres [36,37].”

“The shifting from the live feed in post-larval shrimp to commercial pellets in juvenile shrimp might explain the difference in diversity of bacterial composition in shrimp rearing ponds because bacteria from Proteobacteria, Bacteroidetes, Firmicutes, Actinobacteria have also been found associated to Artemia nauplii [5…”

“Once shrimp reached juvenile stages, their diet is solely based on commercial pellets [52,53].”

“black tiger shrimp has been in a major decline due to higher incidents of viral and bacterial disease outbreaks [10,11].”

“The method has recently gained popularity for an in-depth characterization of various microbial communities such as those in the human [24,25,26], fermented food [27], soil [28,29,30,31], extreme environments [32], the deep marine biosphere [33,34], wastewater treatment bioreactors, [35] and plant biospheres [36,37].”

“Proteobacteria are widespread in aquatic invertebrate gut microbiotas and are often a dominant component of this community in other Crustacea (Hakim, 2015; Holt et al., 2020; Huang, 2014; Meziti, 2010; Rungrassamee, 2013, 2014; Zhang, 2014).”

“DGGE was performed as previously described [29] with slight modifications.”