Probiotic Applications in Crustaceans - Aquaculture Nutrition: Gut Health, Probiotics and Prebiotics - page 298

intestinal microbial communities, or larval associated microbiota, of crustaceans according
to numerous studies using conventional culture based or molecular culture-independent
techniques (Oxley et al. 2002; Payne et al. 2006; Johnson et al. 2008; Daniels et al. 2010;
Liu et al. 2011a). Autochthonous probiotic strains are often locally isolated, produced and
used, most of the time based on an empirical and field led approach. Although this practice
is widely applied, only a limited number of published reports exist. An interesting example
is the development of probiotic products achieved in South American shrimp hatcheries
and initiated by farmers with the aim of improving the levels of beneficial Vibrio spp.,
the sucrose fermenting bacteria, in their systems (Garriques and Wyban 1993). The first
successful application was reported in Ecuadorian hatcheries by Griffith (1995). Further,
Garriques and Arevalo (1995) isolated a V. alginolyticus strain from seawater and used
it to treat larval mortality when challenged with V. parahaemolyticus . Other trials by the
same authors confirmed the improvement in survival and also showed a growth promoting
effect, though statistical analysis of their results carried out later by Gomez-Gil et al. (2000)
indicated no statistical difference in survival and weight gain between treated and control
groups. Nevertheless, since 1995 broad applications of probiotic V. alginolyticus (Ili strain)
have taken place in commercial penaeid hatcheries in Ecuador. The main application of this
probiotic was to control larval diseases and the applications therefore resulted in a large
reduction in antimicrobial products being used in hatcheries. The work reported by these
authors suggested that V. alginolyticus may confer some degree of protection against disease
even if a lack of scientific explanation was evident. According to Rodríguez et al. (2007),
the Ili strain of V. alginolyticus isolated from healthy shrimp larviculture is currently used
in CENAIM's larvae production protocols. The strain is administered at the nauplii 5 stage
and successfully mitigates what is commonly referred to as 'bolitas' syndrome or Zoea 2
syndrome (Vandenberghe et al. 1999).
In contrast to the broad application of Gram-negative bacterial probiotics in the field, rela-
tively few in-depth studies have been published on this subject. Alavandi et al. (2004) isolated
a Pseudomonas sp. and V. fluviatis strains from the gut of farmed sub-adult P. monodon . Selec-
tion was based on several properties such as: predominance in the gut microenvironment,
ability to secrete extracellular macromolecules and digestive enzymes, and production of iron
sequestering compounds like siderophores. However, the authors concluded that the selection
criteria used were not satisfactory as they did not measure any effect on the immune indica-
tors (such as haemocyte counts, PO activity and antibacterial activity in the haemolymph) of
P. monodon shrimp. Other attempts were more successful, as demonstrated by the works of
Chythanya et al. (2002), Gullian (2004), Balcázar (2003) and Balcázar et al. (2006; 2007).
The first of these selected a Pseudomonas strain I-2 (tentatively identified as Ps. aeruginosa ),
isolated from seawater, and demonstrated its inhibitory activity towards several pathogenic
Vibrionaceae such as V. parahaemolyticus , V. harveyi , V. fluvialis, , V. vulnificus and Photobac-
terium damselae . They concluded that Pseudomonas I-2 strain could be used as a biocontrol
agent in shrimp hatcheries and farms. More recently, Hai et al. (2007) also Pseudomonas
synxantha and Ps. aeruginosa strains for use in the culture of western king prawns ( Penaeus
latisulcatus ). The selection was based on the results of five inhibition tests conducted with
various pathogens. Interestingly these authors looked at the in vivo effect of these probiotics
at different levels (Hai et al. 2009; Hai and Fotedar 2009). They first report that the probiotics
were able to improve the health (determined by assessing haemocyte numbers and bacterial
load in the haemolymph) of the shrimp, and that their combination was more efficient than the