title: Application of hierarchical oligonucleotide primer extension (HOPE) to assess relative abundances of ammonia- and nitrite-oxidizing bacteria.
abstract: Establishing an optimal proportion of nitrifying microbial populations, including ammonia-oxidizing bacteria (AOB), nitrite-oxidizing bacteria (NOB), complete nitrite oxidizers (comammox) and ammonia-oxidizing archaea (AOA), is important for ensuring the efficiency of nitrification in water treatment systems. Hierarchical oligonucleotide primer extension (HOPE), previously developed to rapidly quantify relative abundances of specific microbial groups of interest, was applied in this study to track the abundances of the important nitrifying bacterial populations.
  
  
  
  The method was tested against biomass obtained from a laboratory-scale biofilm-based trickling reactor, and the findings were validated against those obtained by 16S rRNA gene-based amplicon sequencing. Our findings indicated a good correlation between the relative abundance of nitrifying bacterial populations obtained using both HOPE and amplicon sequencing. HOPE showed a significant increase in the relative abundance of AOB, specifically Nitrosomonas, with increasing ammonium content and shock loading (p < 0.001). In contrast, Nitrosospira remained stable in its relative abundance against the total community throughout the operational phases. There was a corresponding significant decrease in the relative abundance of NOB, specifically Nitrospira and those affiliated to comammox, during the shock loading. Based on the relative abundance of AOB and NOB (including commamox) obtained from HOPE, it was determined that the optimal ratio of AOB against NOB ranged from 0.2 to 2.5 during stable reactor performance.
  
  
  
  Overall, the HOPE method was developed and validated against 16S rRNA gene-based amplicon sequencing for the purpose of performing simultaneous monitoring of relative abundance of nitrifying populations. Quantitative measurements of these nitrifying populations obtained via HOPE would be indicative of reactor performance and nitrification functionality.
authors.affiliation: Biological and Environmental Science & Engineering Division (BESE), King Abdullah University of Science and Technology (KAUST), Water Desalination and Reuse Center (WDRC), Thuwal, 23955-6900, Saudi Arabia.
authors.forename: Giantommaso
authors.initials: G
authors.lastname: Scarascia
authors.affiliation: Biological and Environmental Science & Engineering Division (BESE), King Abdullah University of Science and Technology (KAUST), Water Desalination and Reuse Center (WDRC), Thuwal, 23955-6900, Saudi Arabia.
authors.forename: Hong
authors.initials: H
authors.lastname: Cheng
authors.affiliation: Biological and Environmental Science & Engineering Division (BESE), King Abdullah University of Science and Technology (KAUST), Water Desalination and Reuse Center (WDRC), Thuwal, 23955-6900, Saudi Arabia.
authors.forename: Moustapha
authors.initials: M
authors.lastname: Harb
authors.affiliation: Biological and Environmental Science & Engineering Division (BESE), King Abdullah University of Science and Technology (KAUST), Water Desalination and Reuse Center (WDRC), Thuwal, 23955-6900, Saudi Arabia. peiying.hong@kaust.edu.sa.
authors.forename: Pei-Ying
authors.initials: PY
authors.lastname: Hong
mesh_terms: D000641:Ammonia / Q000378:metabolism*; D001419:Bacteria / Q000145:classification / Q000235:genetics* / Q000378:metabolism*; D018441:Biofilms; D019149:Bioreactors / Q000382:microbiology; D017931:DNA Primers; D004269:DNA, Bacterial; D059013:Microbial Consortia / Q000235:genetics; D015999:Multivariate Analysis; D058465:Nitrification; D009573:Nitrites / Q000378:metabolism*; D009606:Nitrosomonas / Q000235:genetics / Q000378:metabolism; D010084:Oxidation-Reduction; D010802:Phylogeny; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D062065:Wastewater / Q000382:microbiology; D014867:Water / Q000737:chemistry; D018508:Water Purification
chemical_list: D000641:Ammonia; D017931:DNA Primers; D004269:DNA, Bacterial; D009573:Nitrites; D012336:RNA, Ribosomal, 16S; D062065:Wastewater; D014867:Water
keywords: 16S rRNA gene-based amplicon sequencing; AOB/NOB ratio; Quantitative monitoring; Shock loading event; Single nucleotide primer extension