Diversity of marine and brackish water nitrite-oxidizing consortia developed for activating nitrifying bioreactors in aquaculture

  • P. G. Preena
  • V. J. R. Kumar
  • C. Achuthan
  • R. George
  • R. Boobal
  • R. R. Nair
  • I. S. B. SinghEmail author
Original Paper


Nitrite is a well-known toxicant in aquaculture, produced as intermediate in nitrification. Two nitrite-oxidizing bacterial consortia, one from marine environment and the other from brackish water, were developed by enrichment technique at National Centre for Aquatic Animal Health, for removal of nitrite from recirculating aquaculture systems. In the present study, bacterial diversity of the consortia was assessed based on 16S ribosomal RNA and the functional gene analysis. Clone libraries of 16S ribosomal RNA gene and nitrite oxidoreductase A gene were constructed, and amplified ribosomal DNA restriction analysis was carried out to cluster the clones. Dendrograms generated through molecular characterization showed 29 and 27 clusters in marine and brackish water consortia, respectively. Phylogenetic analyses of representative clones from each cluster depicted profound diversity in the consortia consisting autotrophic nitrifiers belonging to Proteobacteria, anaerobic ammonia oxidizers, Actinobacteria, Bacteroidetes and heterotrophic denitrifiers. Functional gene analysis corroborated with the presence of specific nitrite oxidizers. Quantitative polymerase chain reaction showed the abundance of nitrite oxidizers in the order of 1.51 ± 0.38 × 109/g and 4.88 ± 0.42 × 107/g in marine and brackish water consortia, respectively. Diversity indices and pattern of distribution of organisms within the consortia were analyzed using Geneious, VITCOMIC, Mega 5 and Primer software. The marine nitrite-oxidizing consortium showed higher Shannon–Wiener diversity and mean population diversity than brackish water consortium, suggesting that the former was having more diverse flora and higher potential to be used as startup cultures for activating nitrifying bioreactors subsequent to acclimatization to the required salinity.


Dendrogram Geneious software Mean population Phylogenetic analyses Shannon–Wiener index 



The first author acknowledges Department of Science and Technology (DST) under INSPIRE Scheme with Grant Number IF10193 for the Fellowship and Cochin University of Science and Technology for the support.

Supplementary material

13762_2017_1580_MOESM1_ESM.tif (29 kb)
Fig. 1 Dendrogram of marine nitrite-oxidizing consortium (NIOPCU-1) on the basis of ARDRA (TIFF 29 kb)
13762_2017_1580_MOESM2_ESM.tif (29 kb)
Fig. 2 Dendrogram of brackish water nitrite-oxidizing consortium (NIONPCU-1) on the basis of ARDRA (TIFF 28 kb)
13762_2017_1580_MOESM3_ESM.tif (11.9 mb)
Fig. 3 Phylogenetic analysis of the 16S rRNA gene sequences from marine nitrite-oxidizing consortium (NIOPCU-1) (indicated using Δ) with the matched sequences from GenBank database by using Mega 5.0 software (TIFF 12188 kb)
13762_2017_1580_MOESM4_ESM.tif (11.2 mb)
Fig. 4 Phylogenetic analysis of the 16S rRNA gene sequences from brackish water nitrite-oxidizing consortium (NIONPCU-1) (indicated using Δ) with the matched sequences from GenBank database by using Mega 5.0 software (TIFF 11518 kb)
13762_2017_1580_MOESM5_ESM.xlsx (11 kb)
Fig. 5 Mean diversity in entire population of marine (NIOPCU-1) and brackish (NIONPCU-1) water nitrite-oxidizing consortia (XLSX 11 kb)
13762_2017_1580_MOESM6_ESM.xlsx (11 kb)
Fig. 6 Species richness evenness and dominance observed in marine nitrite-oxidizing consortium (NIOPCU-1) (XLSX 10 kb)
13762_2017_1580_MOESM7_ESM.xlsx (11 kb)
Fig. 7 Species richness evenness and dominance observed in brackish water nitrite-oxidizing consortium (NIONPCU-1) (XLSX 10 kb)
13762_2017_1580_MOESM8_ESM.xlsx (11 kb)
Fig. 8 Shannon–Wiener diversity index observed in marine (NIOPCU-1) and brackish (NIONPCU-1) water nitrite-oxidizing consortia (XLSX 10 kb)
13762_2017_1580_MOESM9_ESM.xlsx (11 kb)
Fig. 9 Copy number of nor A gene present in marine (NIOPCU-1) and brackish (NIONPCU-1) water nitrite-oxidizing consortia determined using quantitative PCR (XLSX 11 kb)


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Copyright information

© Islamic Azad University (IAU) 2017

Authors and Affiliations

  • P. G. Preena
    • 1
  • V. J. R. Kumar
    • 1
    • 2
  • C. Achuthan
    • 1
  • R. George
    • 1
  • R. Boobal
    • 1
  • R. R. Nair
    • 1
  • I. S. B. Singh
    • 1
    Email author
  1. 1.National Centre for Aquatic Animal HealthCochin University of Science and TechnologyCochinIndia
  2. 2.Department of AquacultureKerala University of Fisheries and Ocean StudiesPanangad, KochiIndia

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