Effects of probiotic supplementary bioflocs (Rhodospirillum rubrum, Bacillus subtilis, Providencia rettgeri) on growth, immunity, and water quality in cultures of Pacific white shrimp (Litopenaeus vannamei)

Abstract

Biofloc technology is a promising strategy for the super-intensive cultivation of Pacific white shrimp (Litopenaeus vannamei). However, conventional bioflocs originating from pre-existing systems have shown to be inferior to bioflocs with constant probiotic supplementation regarding the stability in yield. Hence we compared bioflocs supplemented with Rhodospirillum rubrum (RR), Bacillus subtilis (BS), and Providencia rettgeri (PR) to the clear water system (CT), in terms of growth performances, immune status, nitrogen substances, and Vibrio populations in cultures of L. vannamei together with microbial profiles in both the midgut and bioflocs. Our findings exhibited increased average daily gain (ADG) and protein efficiency ratio (PER) among shrimp reared in BS and PR while BS also reduced feed conversion ratio (FCR). Gene expression analyses of growth-related markers revealed an upregulation of amylase and peptide transporter 1 in shrimp of PR treatment, whilst the changes did not outperform the BS supplement in growth promotion. Furthermore, PR treatment was found to enhance the expression of immune-related genes toll-like receptor, prophenoloxidase (proPO), and heat shock protein 70 (hsp70). All biofloc treatments effectively reduced NO2-N level and mitigated Vibrio parahaemolyticus proliferation, particularly in the BS and PR groups. The redundant analysis illustrated shrimp growth closely associated with improved water quality instead of the expression of genes participating in digestion, absorption, or immunity. Moreover, microflora sequencing exhibited the supplemental probiotics altered microbial communities in bioflocs and gut, whereas they did not dominate in either environment. In spite of the discrepancies in microbial composition between bioflocs and midgut of shrimp, BS and PR treatments enriched Rhodobacteraceae populations in both environments. In summary, B. subtilis and P. rettgeri supplementary bioflocs promote L. vannamei growth through nutritional supplementation and water environment optimization. Supplementation of P. rettgeri will simultaneously enhance immune responses and stress resistance of L. vannamei via activating proPO cascade and upregulating hsp70. Given the apparent benefits of heterotrophic probiotic supplementary bioflocs, it is imperative to investigate more microbial species or their combinations that can stimulate bioflocs to produce more nutrients.