Bacterial analysis in the early developmental stages of the black tiger shrimp (Penaeus monodon)

Abstract

Microbial colonization is an essential process in the early life of animal hosts-a crucial phase that could help influence and determine their health status at the later stages. The establishment of bacterial community in a host has been comprehensively studied in many animal models; however, knowledge on bacterial community associated with the early life stages of Penaeus monodon (the black tiger shrimp) is still limited. Here, we examined the bacterial community structures in four life stages (nauplius, zoea, mysis and postlarva) of two black tiger shrimp families using 16S rRNA amplicon sequencing by a next-generation sequencing. Although the bacterial profiles exhibited different patterns in each developmental stage, Bacteroidetes, Proteobacteria, Actinobacteria and Planctomycetes were identified as common bacterial phyla associated with shrimp. Interestingly, the bacterial diversity became relatively stable once shrimp developed to postlarvae (5-day-old and 15-day-old postlarval stages), suggesting an establishment of the bacterial community in matured shrimp. To our knowledge, this is the first report on bacteria establishment and assembly in early developmental stages of P. monodon. Our findings showed that the bacterial compositions could be shaped by different host developmental stages where the interplay of various host-associated factors, such as physiology, immune status and required diets, could have a strong influence.

Figure 1

Schematic diagram of collection of early developmental stages in black tiger shrimp. Shrimp and rearing water (W) were collected at the stages of nauplius (N), zoea (Z), mysis (M), 5-day-old postlarva (PL5) and 15-day-old postlarva (PL15) from different family (Family A and Family B) for microbiota analysis.

Figure 2

Diversity analysis. Chao1 index (a) and Shannon index (b) were used to estimate bacterial diversity of shrimp and rearing water at stages of nauplius (N), zoea (Z), mysis (M), 5-day-old postlarva (PL5) and 15-day-old postlarva (PL15) from different family (Family A and Family B) as shown with gray and white plot graph, respectively). The box delimits the 25^th and 75^th percentile, the line in each box indicates the median, and the whiskers indicate the lowest and highest values. Different letters show significant difference between groups by ANOVA (p value < 0.05) (n = 3).

Figure 3

Dominant bacterial communities associated with shrimp and rearing water. Shrimp samples and rearing water were collected at nauplius (N), zoea (Z), mysis (M), 5-day-old postlarval (PL5) and 15-day-old postlarval (PL15) stages from two families (Family A and Family B). Distribution of bacterial phyla associated to shrimp and their rearing water with their relative abundance greater 0.2% were shown and those with their abundance less than 0.2% were grouped as other phyla. (a) Stacked bar plots represent the top 5 dominant bacterial genera associated with shrimp from each family at each life stage from each phylum, Proteobacteria (b), Bacteroidetes (c), Planctomycetes (d) and Actinobacteria. (e) Non-top five genera were shown under other genera.

Figure 4

Principal coordinates analysis (PCoA) of bacterial communities associated in shrimp at different developmental stages (filled symbols) and rearing water (unfilled symbols) from Family A and Family B under different growth stages (a) and dendrogram of cluster analysis showing similarities in percent of OTUs in shrimp at different developmental stages. (b) Bacterial profiles were analyzed base on Bray-Curtis Dissimilarity method. Plot colors, blue, orange, yellow, green and pink represent nauplius (N), zoea (Z), mysis (M), 5-day-old postlarval (PL5) and 15-day-old postlarval (PL15) stages, respectively.

Figure 5

LEfSe analysis show differentially abundance OTUs of bacterial community-associated with early shrimp stages, which were divided to nauplius (N), zoea (Z), mysis (M), 5-day-old postlarval (PL5) and 15-day-old postlarval (PL15) from Family A and Family B. The threshold of the logarithmic linear discriminant analysis (LDA) score was 4.0. Circle, square and triangle, represent OTUs belong to Proteobacteria, Planctomycetes, and Bacteroidetes, respectively.

Figure 6

Venn diagram shows the numbers of OTUs that were unique or shared in shrimp at each life stage (a). The bubble plot represents relative abundance of unique or shared bacterial taxa at OTU level in shrimp at each life stages (nauplius, zoea, mysis, and 5-day-old postlarval (PL5) and 15-day-old postlarval (PL15) from Family A and B) and their rearing water.