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Abstract

Improvements in water quality requires the removal of nitrogen compounds from wastewater. The most promising and cost-effective methods for this purpose are biological ones based on activated sludge microorganisms such as nitrifiers, denitrifiers, and anammox bacteria. Due to the most of the nitrogen removal bacteria are uncultivable in a laboratory, the application of the molecular tools is required to investigate microorganisms involved in the nitrogen removal. In case of this study for the analysis of relative genes abundance of nitrogen removal bacteria, quantitative PCR (qPCR) based on bacterial DNA and qPCR preceded by reverse transcription (RT-qPCR) based on bacterial mRNA as a template, were used with specific bacterial functional genes ( amoA, nrxA, nirS, nirK, hzo). Samples from four anammox sequencing batch reactors (SBRs) were analyzed, while the nitrogen removal process and bacteria growth were supported by biomass immobilization and nanoparticles addition. There were statistically significant differences between results obtained in the case of mRNA and DNA (p<0.05). Statistically significant positive correlations were found between results obtained with those two approaches. In case of mRNA analysis, positive results were obtained only for hzo, amoA and partly for nirS genes, despite additional purification and removal of inhibitors from samples prior to reaction.
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Authors and Affiliations

Anna Banach-Wiśniewska
1
Filip Gamoń
1
Aleksandra Ziembińska-Buczyńska
1

  1. Silesian University of Technology, Faculty of Power and Environmental Engineering, Environmental Biotechnology Department, Gliwice, Poland
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Abstract

Leaf scald, caused by the necrotrophic fungus Monographella albescens, is one of the main threats to rice (Oryza sativa L.) around the world. This disease decreases yields in rice by up to 30% because of dead leaf tissue, damaged seeds, and sterile flowers. Currently, there is limited knowledge about the molecular mechanisms involved in rice plant resistance against this pathogen. For this purpose, six commercial cultivars of rice were primarily screened for M. albescens infection and development. Dasht and Salari were found to be the most resistant and susceptible to M. albescens infection, respectively. The plants were kept in a greenhouse at 29 ± 2°C during the day and 26 ± 2°C at night with a relative air humidity of 85 ± 5%. Forty-five days after sowing, the plants with three biological replications were inoculated by transferring a PDA disc (0.3 cm2) containing M. albescens mycelia to the middle third of the 7th, 8th, and 9th completely open leaves. The leaves were collected 24, 48, 72, 96 and 120 hai. Leaf samples were also collected from the non-inoculated plants (0 h) to serve as controls. Real-time quantitative PCR (RT-qPCR) showed rapid induction and significant accumulation of jasmonic acid (JA) and ethylene (ET) responsive genes such as lipoxygenase (LOX), allene oxide synthase 2 (Aos2), jasmonic acid carboxyl methyltransferase 1 (JMT1) and ACC synthase 1 (ACS1) in the resistant Dasht cultivar after infection with M. albescens. Furthermore, the transcripts of salicylic acid (SA) responsive phenyl alanine ammonia lyase 1 (PAL1) and nonexpressor of pathogenesis-related genes 1 (NPR1) genes were induced in the incompatible interaction. The activities of the defense enzymes superoxide dismutase (SOD), peroxidase (POX) and glutathione reductase (GR) increased strongly in Dasht in response to M. albescens infection. In addition, there was an increase in the H2O2 levels in the leaves of the Dasht cultivar during the infectious period of M. albescens associated with the enhancement of catalase (CAT) activity as well as higher levels of malondialdehyde (MDA). This is the first study on the interaction between rice and M. albescens at the molecular level. It can contribute to understanding how rice responds to pathogen infection, as well as assist with future research plans of molecular breeding regarding the tolerance to leaf scald disease.

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Authors and Affiliations

Dariush Ebadi Almas
Atefeh Rahmani Kamrodi

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