Ecological Engineering 95 (2016) 180–188 Contents lists available at ScienceDirect Ecological Engineering journal homepage: www.elsevier.com/locate/ecoleng Organic load and hydraulic regime influence over the bacterial community responsible for the nitrogen cycling in bed media of vertical subsurface flow constructed wetland Catiane Pelissari a , Mayara Oliveira dos Santos a , Benny Zuse Rousso a , Alessandra Pellizzaro Bento b , Rafael Dutra de Armas c , Pablo Heleno Sezerino a,∗ a GESAD- Decentralized Sanitation Research Group, Department of Sanitary and Environmental Engineering, Federal University of Santa Catarina, Trindade, Florianópolis, Santa Catarina, Zip Code 88040-900, Brazil b Environmental Municipality Foundation of Florianópolis – FLORAM, Rua João Pio Duarte Silva, n. 535, Corrego Grande, Florianópolis, Santa Catarina, Zip Code 88037-001, Brazil c Department of Cellular Biology, Embryology and Genetics, Federal University of Santa Catarina, Trindade, Florianópolis, Santa Catarina, Zip Code 88040-900, Brazil a r t i c l e i n f o Article history: Received 25 December 2015 Received in revised form 9 June 2016 Accepted 18 June 2016 Keywords: Wastewater Vertical flow constructed wetland Microcosms Bacteria Nitrifying bacteria Denitrifying bacteria a b s t r a c t The aim of this study was to evaluate the influence of the organic loading rate (OLR) and the hydraulic regime on the nitrifying and denitrifying bacterial community present in the bed media of vertical flow constructed wetland (VFCW) employed as urban wastewater treatment solution. Two microcosms filled with sand (d 10 = 0.3 mm and uniformity coefficient = 6.2) simulated the vertical profile of VFCW and were monitored for 360 days. Microcosm 1 was operated with a constant OLR of 41 g COD m −2 d −1 and with an average hydraulic loading rate (HLR) of 72 mm d −1 . Microcosm 2 was operated with a constant HLR of 170.5 mm d −1 and with an average OLR of 104 g COD m −2 d −1 . After 180 days feeding weekly, an inter- mittent regime with 30 days of feeding followed by 30 days of rest period was adopted. In total, 7 periods were monitored (4 periods of feeding and 3 periods of rest). At the end of each period, the total, nitrify- ing and denitrifying bacterial communities were characterized using FISH technique. At the end of the experiment, genome sequencing of the bacterial community was performed at the surface and bottom of the two microcosms. The OLR and the HLR significantly affected microcosms’ performance and bacterial community structure. Rest periods did not affect Bacteria domain abundance. However, they presented an influence on nitrifying bacteria abundance. Ammonia oxidizing bacteria abundance (Nitrososmas sp and AOB -proteobacteria) was higher in microcosm 1 in feeding (15%) and rest (3%), periods when com- pared to microcosm 2 (9% and 1% for feeding and rest periods). The same dynamics were observed for nitrite oxidizing bacteria (Nitrobacter sp, Nitrospirae and Nistrospina gracilis), with relative abundance of 18% and 10% for microcosm 1, and 8% and 4% for microcosm 2, in feeding and rest periods, respectively. Differently from nitrifying bacteria dynamics, a higher abundance of denitrifying bacteria (Pseudomonas spp.) was observed in rest periods (12% for microscom 1 and 18% for microcosm 2) than in feeding periods (2% for microcosm 1 and 10% for microcosm 2). The identification of nitrifying and denitrifying bacteria in both microcosms can indicate the process of simultaneous nitrification and denitrification. © 2016 Elsevier B.V. All rights reserved. 1. Introduction Vertical flow constructed wetlands (VFCW) are an ecotechnol- ogy widely employed as treatment solution to different influent ∗ Corresponding author. E-mail addresses: catianebti@gmail.com (C. Pelissari), pablo.sezerino@ufsc.br (P.H. Sezerino). wastewater compositions. These units are designed to pro- mote advanced treatment performance, in terms of carbonaceous organic matter and suspended solids removal as well in nitrogen transformations, markedly the nitrification (Kadlec and Wallace, 2009). VFCW sizing is dependent on the hydraulic loading rates (HLR) and the organic loading rates (OLR) applied on its superficial area (Sezerino et al., 2012; Hoffmann et al., 2011; Platzer, 1999). Besides HLR and OLR, the hydraulic regime is also a fundamental param- http://dx.doi.org/10.1016/j.ecoleng.2016.06.079 0925-8574/© 2016 Elsevier B.V. All rights reserved.