Wastewater irrigation increases abundance of 1 potentially harmful Gammaproteobacteria in 2 soils from Mezquital Valley , Mexico

21 Wastewater contains a large amount of pharmaceuticals, pathogens, and antimicrobial 22 resistance determinants. Only little is known about the dissemination of resistance 23 determinants and changes in soil microbial communities affected by wastewater irrigation. 24 Community DNA from Mezquital Valley soils under irrigation with untreated wastewater for 25 0 to 100 years was analyzed by quantitative real time PCR for the presence of sul genes, 26 encoding resistance to sulfonamides. Amplicon sequencing of bacterial 16S rRNA genes from 27 community DNA from soils irrigated for 0, 8, 10, 85, and 100 years was performed revealing 28 a 14% increase of the relative abundance of Proteobacteria in rainy season and a 26.7% 29 increase in dry season soils irrigated for 100 years with wastewater. In particular, 30 Gammaproteobacteria, including potential pathogens like Pseudomonas, Stenotrophomonas 31 and Acinetobacter spp. were found in wastewater-irrigated fields. 16S rRNA gene sequencing 32 of 96 isolates from soils irrigated with wastewater for 100 years (48 from dry and 48 from 33 rainy season) revealed that 46% affiliated with Gammaproteobacteria (mainly potentially 34 pathogenic Stenotrophomonas strains) and 50% with Bacilli, whereas all 96 isolates from 35 rain-fed soils (48 from dry and 48 from rainy season) affiliated with Bacilli. Up to six 36 antibiotic resistances were found in isolates from wastewater-irrigated soils, sulfamethoxazole 37 resistance was the most abundant (33.3% of the isolates), followed by oxacillin resistance 38 (21.9% of the isolates). In summary, we detected an increase of potentially harmful bacteria 39 and larger incidence of resistance determinants in wastewater-irrigated soils which might 40 result in health risks for farmworkers and consumers of wastewater-irrigated crops. 41 42 on S etem er 3, 2017 by gest ht://aem .sm .rg/ D ow nladed fom