Characterization of Phragmites cummunis rhizosphere bacterial communities and metabolic products during the two stage sequential treatment of post methanated distillery effluent by bacteria and wetland plants

This study deals with the characterization of rhizosphere bacterial communities and metabolic products produced during the two stage sequential treatment of post methanated distillery effluent by bacteria and constructed wetland plants. Results showed that bacterial treatment followed by wetland plants (Phragmites cummunis) resulted 94.5% and 96.0% reduction in BOD and COD values, respectively. The PCR-RFLP analysis showed the presence of Stenotrophomonas, Enterobacter, Pantoea, Acinetobacter and Klebsiella sp., as dominant rhizosphere bacterial communities which play an important role in degradation and decolorization of PMDE in wetland treatment system. Further, the LC-MS-MS and other spectrophotometric analysis have shown that most of the pollutants detected in untreated PMDE were diminished from bacteria and wetland plant treated PMDE indicating that bacteria and wetland plant rhizosphere microbes utilized them as carbon, nitrogen and energy source. While, methylbenzene, furfuryl alcohol, and 4-vinyl-2-methoxyphenol were detected as metabolites in bacteria and hexadecanol in wetland plant rhizosphere treated PMDE.     

Endophytic bacteria enhance remediation of tannery effluent in constructed wetlands vegetated with Leptochloa fusca

The use of constructed wetlands (CWs) is a promising approach for the remediation of wastewater. The present study aims to develop a plant–bacteria system within CWs for the efficient remediation of tannery effluent. In a vertical-flow CW vegetated with Leptochloa fusca (Kallar grass), a consortium of three different endophytic bacteria, Pantoea stewartii ASI11, Microbacterium arborescens HU33, and Enterobacter sp. HU38, was used for bioaugmentation. CWs vegetated with only L. fusca had the potential to remediate tannery effluent, but augmentation with endophytic bacteria enhanced the growth of L. fusca while aiding in the removal of both organic and inorganic pollutants from the tannery effluent. Moreover, the bacterial augmentation decreased toxicity in the effluent as well. A higher number of chromium (Cr)-resistant bacteria were isolated from the rhizosphere and endosphere of L. fusca inoculated with the endophytes than from uninoculated plants. Due to promising bioremediation and detoxification potential of L. fusca, it is reported for the first time as a potential candidate to develop effective CWs for the remediation of polluted effluents in combination with pollutant-degrading endophytic bacteria.     

Microbial diversity in hypersaline wastewater: the example of tanneries

In contrast to conventional wastewater treatment plants and saline environments, little is known regarding the microbial diversity of hypersaline wastewater. In this study, the microbial communities of a hypersaline tannery effluent, and those of three treatment systems operating with the tannery effluent, were investigated using 16S rDNA phylogenetic markers. The comparative analysis of 377 bacterial sequences revealed the high diversity of this type of hypersaline environment, clustering within 193 phylotypes (≥ 97% similarity) and covering 14 of the 52 divisions of the bacterial domain, i.e. Proteobacteria, Bacteroidetes, Firmicutes, Actinobacteria, Chlorobi, Planctomycetes, Spirochaetes, Synergistes, Chloroflexi, Thermotogae, Verrucomicrobia, OP3, OP11 and TM7. Most of the phylotypes were related to halophilic and pollutant-degrading bacteria. Using statistical analysis, the diversity of this type of environment was compared to that of other environmental samples selected on the basis of their salinity, oxygen content and organic load.     

Bacterial decolorization and detoxification of black liquor from rayon grade pulp manufacturing paper industry and detection of their metabolic products

This study deals with the decolorization of black liquor (BL) by isolated potential bacterial consortium comprising Serratia marcescens (GU193982), Citrobacter sp. (HQ873619) and Klebsiella pneumoniae (GU193983). The decolorization of BL was studied by using the different nutritional as well as environmental parameters. In this study, result revealed that the ligninolytic activities were found to be growth associated and the developed bacterial consortium was efficient for the reduction of COD, BOD and color up to 83%, 74% and 85%, respectively. The HPLC analysis of degraded samples of BL has shown the reduction in peak area compared to control. Further, the GC-MS analysis showed that, most of the compounds detected in control were diminished after bacterial treatment while, formic acid hydrazide, 4-cyclohexane-1,2-dicarboxylic acid, carbamic acid, 1,2-benzenedicarboxylic acid and erythropentanoic acid were found as new metabolites. Further, the seed germination test using Phaseolus aureus has supported the detoxification of bacterial decolorized BL.     

A simple strategy for investigating the diversity and hydrocarbon degradation abilities of cultivable bacteria from contaminated soil

The use of indigenous bacterial strains is a valuable bioremediation strategy for cleaning the environment from hydrocarbon pollutants. The isolation and selection of hydrocarbon-degrading bacteria is therefore crucial for obtaining the most promising strains for site decontamination. Two different media, a minimal medium supplemented with a mixture of polycyclic aromatic hydrocarbons and a MS medium supplemented with triphenyltetrazolium chloride, were used for the isolation of bacterial strains from two hydrocarbon contaminated soils and from their enrichment phases. The hydrocarbon degradation abilities of these bacterial isolates were easily and rapidly assessed using the 2,6-dichlorophenol indophenol assay. The diversity of the bacterial communities isolated from these two soil samples and from their enrichment phases was evaluated by the combination of a bacterial clustering method, fluorescence ITS-PCR, and bacterial identification by 16S rRNA sequencing. Different PCR-based assays were performed in order to detect the genes responsible for hydrocarbon degradation. The best hydrocarbon-degrading bacteria, including Arthrobacter sp., Enterobacter sp., Sphingomonas sp., Pseudomonas koreensis, Pseudomonas putida and Pseudomonas plecoglossicida, were isolated directly from the soil samples on minimal medium. The nahAc gene was detected only in 13 Gram-negative isolates and the sequences of nahAc-like genes were obtained from Enterobacter, Stenotrophomonas, Pseudomonas brenneri, Pseudomonas entomophila and P. koreensis strains. The combination of isolation on minimal medium with the 2,6-dichlorophenol indophenol assay was effective in selecting different hydrocarbon-degrading strains from 353 isolates.     

Biodegradation of naphthalene-2-sulfonic acid present in tannery wastewater by bacterial isolates <Emphasis Type="Italic">Arthrobacter</Emphasis> sp. 2AC and <Emphasis Type="Italic">Comamonas</Emphasis> sp. 4BC

Two bacterial strains, 2AC and 4BC, both capable of utilizing naphthalene-2-sulfonic acid (2-NSA) as a sole source of carbon, were isolated from activated sludges previously exposed to tannery wastewater. Enrichments were carried out in mineral salt medium (MSM) with 2-NSA as the sole carbon source. 16S rDNA sequencing analysis indicated that 2AC is an Arthrobacter sp. and 4BC is a Comamonas sp. Within 33 h, both isolates degraded 100% of 2-NSA in MSM and also 2-NSA in non-sterile tannery wastewater. The yield coefficient was 0.33 g biomass dry weight per gram of 2-NSA. A conceptual model, which describes the aerobic transformation of organic matter, was used for interpreting the biodegradation kinetics of 2-NSA. The half-lives for 2-NSA, at initial concentrations of 100 and 500 mg/l in MSM, ranged from 20 h (2AC) to 26 h (4BC) with lag-phases of 8 h (2AC) and 12 h (4BC). The carbon balance indicates that 75–90% of the initial TOC (total organic carbon) was mineralized, 5–20% remained as DOC (dissolved organic carbon) and 3–10% was biomass carbon. The principal metabolite of 2-NSA biodegradation (in both MSM and tannery wastewater) produced by Comamonas sp. 4BC had a MW of 174 and accounted for the residual DOC (7.0–19.0% of the initial TOC and 66% of the remaining TOC). Three to ten percent of the initial TOC (33% of the remaining TOC) was associated with biomass. The metabolite was not detected when Arthrobacter sp. 2AC was used, and a lower residual DOC and biomass carbon were recorded. This suggests that the two strains may use different catabolic pathways for 2-NSA degradation. The rapid biodegradation of 2-NSA (100 mg/l) added to non-sterile tannery wastewater (total 2-NSA, 105 mg/l) when inoculated with eitherArthrobacter 2AC or Comamonas 4BC showed that both strains were able to compete with the indigenous microorganisms and degrade 2-NSA even in the presence of alternate carbon sources (DOC in tannery wastewater = 91 mg/l). The results provide information useful for the rational design of bioreactors for tannery wastewater treatment.     

Microbial community analysis in crab ponds by denaturing gradient gel electrophoresis

In order to investigate the role of microbial community in aquatic ecology and biogeochemical cycles, the bacterial community in crab ponds was investigated and the effects of aeration and season on the bacterial community were also assessed. Total DNAs from the water samples were amplified with universal primers and the amplicons were then resolved by denaturing gradient gel electrophoresis. Bands from the resulting profiles were excised and sequenced. Cluster analysis of the resulting profiles showed that the microbial community was affected by aeration and season. The microbial community between the surface and bottom of the water was very similar. A total of fifteen bands were obtained in this study. Three of them were 91–99% similar to uncultured bacterium clones. Three were 95–99% similar to uncultured Verrucomicrobia bacteria. Three were 97–100% similar to Actinobacterium sp.. Two were similar to Candidatus Limnoluna rubra with similarity 96 and 99%, respectively. Four were 99% similar to Rhodococcus sp., 100% similar to Sporosarcina sp., 100% similar to Stenotrophomonas sp., and 98% similar to Hydrogenophaga sp., respectively. The concentrations of dissolved oxygen, total nitrogen, total phosphorous, nitrite, and ammonia and pH values were significantly affected by season while only the pH value and the concentrations of dissolved oxygen and total nitrogen were significantly affected by aeration.     

16S Ribosomal DNA-Based Analysis of Bacterial Diversity in Purified Water Used in Pharmaceutical Manufacturing Processes by PCR and Denaturing Gradient Gel Electrophoresis

The bacterial community in partially purified water, which is prepared by ion exchange from tap water and is used in pharmaceutical manufacturing processes, was analyzed by denaturing gradient gel electrophoresis (DGGE). 16S ribosomal DNA fragments, including V6, -7, and -8 regions, were amplified with universal primers and analyzed by DGGE. The bacterial diversity in purified water determined by PCR-DGGE banding patterns was significantly lower than that of other aquatic environments. The bacterial populations with esterase activity sorted by flow cytometry and isolated on soybean casein digest (SCD) and R2A media were also analyzed by DGGE. The dominant bacterium in purified water possessed esterase activity but could not be detected on SCD or R2A media. DNA sequence analysis of the main bands on the DGGE gel revealed that culturable bacteria on these media were Bradyrhizobium sp., Xanthomonas sp., and Stenotrophomonas sp., while the dominant bacterium was not closely related to previously characterized bacteria. These data suggest the importance of culture-independent methods of quality control for pharmaceutical water.     

Efficacy of Acinetobacter sp. B9 for simultaneous removal of phenol and hexavalent chromium from co-contaminated system

The present study shows the feasibility of a newly isolated strain Acinetobacter sp. B9 for concurrent removal of phenol and Cr (VI) from wastewater. The experiments were conducted in a batch reactor under aerobic conditions. Initially, when mineral salt solution was used as the culture medium, the strain was found to utilize phenol as sole carbon and energy source while no Cr (VI) removal was observed. However, the addition of glucose as co-carbon source resulted in the removal of both toxicants. This co-removal efficiency of the strain was further improved with nutrient-rich media (NB). Optimum co-removal was determined at 188 mg L^?1 of phenol and 3.5 mg L^?1 of Cr (VI) concentrations at pH 7.0. Strain B9 followed the orthometabolic pathway for phenol degradation. Transmission electron microscopy (TEM) and Fourier transform infrared spectroscopy (FT-IR) studies showed sorption of chromium as one of the major mechanisms for Cr (VI) removal by B9 cells. Acinetobacter sp. B9 was later on checked for bioremediation of real tannery wastewater. After 96 h of batch treatment of tannery effluent containing an initial 47 mg L^?1 phenol and 16 mg L^?1 Cr (VI), complete removal of phenol and 87 % reduction of Cr (VI) were attained, showing high efficiency of the bacterial strain for potential application in industrial pollution control.     

Occurrence and removal of parasites, enteric bacteria and faecal contamination indicators in wastewater natural reclamation systems in Tenerife-Canary Islands, Spain

Two wastewater natural reclamation systems (WWNRS) have been compared regarding their efficiencies on faecal bacteria removal and the persistence of enteric pathogens. These WWNRS are constituted of a combination of anaerobic treatment, small sub-surface flow constructed wetland refilled of volcanic ashes and a final pond as water reservoir. Faecal coliforms, enterococci, Escherichia coli, Clostridium perfringens, somatic coliphages, Salmonella sp., Campylobacter sp., Cryptosporidium sp., Giardia sp. and helminth eggs were analyzed in constructed wetlands inlet and outlet and storage pond effluent. Low numbers of protozoan positive samples (4.54% in Albergue de Bolico for both protozoa, and 19.05% in Carrizal Alto for Giardia sp.) and absence of helminth eggs were found. Both systems demonstrated efficient reduction of faecal contamination indicators in the wastewaters (removal rates values of 2 log₁₀). The natural systems for wastewater treatment used to be efficient in Salmonella abatement, this fact was confirmed in the reported systems, since enterobacteriaceae were found in only one of the effluents. Campylobacter species associated with the access of animals to storage ponds were detected in the reclaimed water.     

Restoration of the ability to settle bulking sludge by bacterial seeding in wastewater treatment

An attempt to restore the settling ability of denatured bulking sludge was under-taken by changing the microflora in waste treatment tanks. A cell suspension of mixed cultures of ten strains of bacteria, which were isolated from normal activated sludge from night soil plants, and a type culture Zoogloea ramigera IAM1236 was seeded into a laboratory-scale aeration tank containing bulking sludge collected from municipal night soil or a food processing (bean curd production) waste treatment plant. The tank was fed with synthetic wastewater or industrial waste and aerated for 22 days. After 5 days, the microflora in the sludge changed remarkably with the seeding of the bacterial culture; filamentous organisms disappeared and active protozoa (Vorticella sp., Epistylis sp., and Lecane sp.) appeared. The sludge became compact and settled rapidly. The SV₃₀ of the sludge temporarily increased, but in the end decreased from 97 to 20%. The CODcr value decreased from 300 to 20 ppm. In the tank without seeding, the sludge contained almost filamentous organisms only which floated and finally decomposed. The effects were confirmed by applied tests in 700-ton scale and 100-ton scale aeration tanks of municipal night soil and a food processing waste treatment plant, respectively.     

Comparison of different pilot scale bioreactors for the treatment of a real wastewater from the textile industry

Wastewater from textile industry usually undergoes activated sludge biotreatment ahead of refining treatments, final discharge or reuse. To identify the most effective bioreactor typology for the secondary treatment of a wastewater resulting from a textile industry of the Biella district (Italy), four pilot units characterized by a different configuration and fluid dynamics (i.e., Bioflotation^®, Fixed Bed Biofilm Reactor (FBBR), flow-jet aeration and standard aerobic sludge reactors) were operated in parallel, inoculated with the same microbial consortium and fed with identical streams of wastewater discharged from wet textile processes of the industy. COD, TC and non-ionic surfactants were monitored in effluents of the compared bioreactors working under continuous mode and the cultivable heterotrophic microorganisms prevailing in each of them were isolated and characterized as the end of the study. The results demonstrated that the air supply system greatly influenced the treatment efficiency which reached the highest value in the case of Bioflotation^® and FBBR technology. A highly specialized bacterial biomass mostly composed by strains of the Pseudomonas, Stenotrophomonas and Ochrobactrum genera was isolated in such reactors, thus suggesting that a direct correlation between reactor configuration, decontamination performances and microbial biomass composition exist.     

Abundance and Diversity of Bacterial Nitrifiers and Denitrifiers and Their Functional Genes in Tannery Wastewater Treatment Plants Revealed by High-Throughput Sequencing

Biological nitrification/denitrification is frequently used to remove nitrogen from tannery wastewater containing high concentrations of ammonia. However, information is limited about the bacterial nitrifiers and denitrifiers and their functional genes in tannery wastewater treatment plants (WWTPs) due to the low-throughput of the previously used methods. In this study, 454 pyrosequencing and Illumina high-throughput sequencing, combined with molecular methods, were used to comprehensively characterize structures and functions of nitrification and denitrification bacterial communities in aerobic and anaerobic sludge of two full-scale tannery WWTPs. Pyrosequencing of 16S rRNA genes showed that Proteobacteria and Synergistetes dominated in the aerobic and anaerobic sludge, respectively. Ammonia-oxidizing bacteria (AOB) amoA gene cloning revealed that Nitrosomonas europaea dominated the ammonia-oxidizing community in the WWTPs. Metagenomic analysis showed that the denitrifiers mainly included the genera of Thauera, Paracoccus, Hyphomicrobium, Comamonas and Azoarcus, which may greatly contribute to the nitrogen removal in the two WWTPs. It is interesting that AOB and ammonia-oxidizing archaea had low abundance although both WWTPs demonstrated high ammonium removal efficiency. Good correlation between the qPCR and metagenomic analysis is observed for the quantification of functional genes amoA, nirK, nirS and nosZ, indicating that the metagenomic approach may be a promising method used to comprehensively investigate the abundance of functional genes of nitrifiers and denitrifiers in the environment.     

Phycoremediation of Tannery Wastewater Using Microalgae Scenedesmus Species

A number of microalgae species are efficient in removing toxicants from wastewater. Many of these potential species are a promising, eco-friendly, and sustainable option for tertiary wastewater treatment with a possible advantage of improving the economics of microalgae cultivation for biofuel production. The present study deals with the phycoremediation of tannery wastewater (TWW) using Scenedesmus sp. isolated from a local habitat. The test species was grown in TWW under laboratory conditions and harvested on the 12th day. The results revealed that the algal biomass during the growth period not only reduced the pollution load of heavy metals (Cr-81.2–96%, Cu-73.2–98%, Pb-75–98% and Zn-65–98%) but also the nutrients (NO₃ >44.3% and PO₄ >95%). Fourier Transform Infrared (FTIR) spectrums of Scenedesmus sp. biomass revealed the involvement of hydroxyl amino, carboxylic and carbonyl groups. The scanning electron micrograph (SEM) and Energy Dispersive X-ray Spectroscopic analysis (EDS) revealed the surface texture, morphology and element distribution of the biosorbent. Furthermore, the wastewater generated during wet-blue tanning process can support dense population of Scenedesmus sp., making it a potential growth medium for biomass production of the test alga for phycoremediation of toxicants in tannery wastewaters.     

Initial Stages in the Biodegradation of the Surfactant Sodium Dodecyltriethoxy Sulfate by Pseudomonas sp. Strain DES1

The biodegradation of the surfactant sodium dodecyltriethoxy sulfate by Pseudomonas sp., strain DES1 (isolated from activated sludge plant effluent) has been studied. Growth of the organism when the ³⁵S-labeled surfactant was present as the sole source of carbon and energy led to the appearance in the culture fluid of five ³⁵S-labeled organic metabolites. These have been identified as mono-, di-, and triethylene glycol monosulfates (major metabolites) and acetic acid 2-(ethoxy sulfate) and acetic acid 2-(diethoxy sulfate), authentic samples of which have been prepared and characterized. Evidence is presented that the major metabolites were produced by rupture of one or another of the three ether linkages present in the surfactant molecule, probably via the agency of a single etherase enzyme. Acetic acid 2-(ethoxy sulfate) and acetic acid 2-(diethoxy sulfate) were formed by the oxidation of the free alcohol groups of di- and triethylene glycol monosulfates, respectively, and increased in amount during the stationary phase of growth. Inorganic ³⁵S-sulfate also appeared in significant quantities in culture fluids and arose from the parent surfactant (presumably via the action of an alkylsulfatase) and not from any of the five metabolites. The appearance of sulfated organic metabolites during the exponential phase of growth and their quantitative relationship remained remarkably constant, even when additional carbon and energy sources (succinate or yeast extract) were also present in the growth media.     

Retention in Treated Wastewater Affects Survival and Deposition of Staphylococcus aureus and Escherichia coli in Sand Columns

The fate and transport of pathogenic bacteria from wastewater treatment facilities in the Earth''s subsurface have attracted extensive concern over recent decades, while the impact of treated-wastewater chemistry on bacterial viability and transport behavior remains unclear. The influence of retention time in effluent from a full-scale municipal wastewater treatment plant on the survival and deposition of Staphylococcus aureus and Escherichia coli strains in sand columns was investigated in this paper. In comparison to the bacteria cultivated in nutrient-rich growth media, retention in treated wastewater significantly reduced the viability of all strains. Bacterial surface properties, e.g., zeta potential, hydrophobicity, and surface charges, varied dramatically in treated wastewater, though no universal trend was found for different strains. Retention in treated wastewater effluent resulted in changes in bacterial deposition in sand columns. Longer retention periods in treated wastewater decreased bacterial deposition rates for the strains evaluated and elevated the transport potential in sand columns. We suggest that the wastewater quality should be taken into account in estimating the fate of pathogenic bacteria discharged from wastewater treatment facilities and the risks they pose in the aquatic environment.     

Bioreduction of Cr(VI) by alkaliphilic Bacillus subtilis and interaction of the membrane groups

Detoxification of Cr(VI) under alkaline pH requires attention due to the alkaline nature of many effluents. An alkaliphilic gram-positive Bacillus subtilis isolated from tannery effluent contaminated soil was found to grow and reduce Cr(VI) up to 100% at an alkaline pH 9. Decrease in pH to acidic range with growth of the bacterium signified the role played by metabolites (organic acids) in chromium resistance and reduction mechanism. The XPS and FT-IR spectra confirmed the reduction of Cr(VI) by bacteria into +3 oxidation state. Chromate reductase assay indicated that the reduction was mediated by constitutive membrane bound enzymes. The kinetics of Cr(VI) reduction activity derived using the monod equation proved (K_s = 0.00032) high affinity of the organism to the metal. This study thus helped to localize the reduction activity at subcellular level in a chromium resistant alkaliphilic Bacillus sp.     

Bacterial communities in a crude oil gathering and transferring system (China)

Bacterial communities in crude oil and oil field production water samples from an oil gathering and transferring system in Changqing Oil field in China were investigated by 16S rRNA denaturing gradient gel electrophoresis (DGGE) analysis followed by gene cloning and sequencing. DGGE profiles showed that bacterial communities are far more rich in the water samples than that in the crude oil samples, and that bacteria related to Ochrobactrum sp. and Stenotrophomonas sp. were detected in all crude oil and oil field water samples. Bacteria related to Burkholderia sp., Brevundimonas sp., and Propionibacterium sp. were detected in the crude oil samples but not in water samples. Bacteria related to Hippea sp., Acidovorax sp., Arcobacter sp., Pseudomonas sp., Thiomicrospira sp., Brevibacterium sp., Tissierella sp. and Peptostreptococcus sp. were detected in the water samples but not in crude oil samples. Using an archaea-specific primer set, methanogens related to Methanomicrobials and Methanosarcinales were found in water samples but not in crude oil samples. The comparability of the microbial communities in the water and crude oil phase during the period of oil gathering and transferring process was 83.3% and 88.2%, respectively, indicating a stable structure of the microbial communities.     

Characterization of Bacterial Communities in Sediments Receiving Various Wastewater Effluents with High-Throughput Sequencing Analysis

454 Pyrosequencing was applied to examine bacterial communities in sediment samples collected from a river receiving effluent discharge from rural domestic sewage (RDS) and various factories, including a tannery (TNS), clothing plant (CTS), and button factory (BTS), respectively. For each sample, 4,510 effective sequences were selected and utilized to do the bacterial diversity and abundance analysis, respectively. In total, 1,288, 2,036, 1,800, and 2,150 operational taxonomic units were obtained at 3 % distance cutoff in TNS, CTS, BTS, and RDS, respectively. Bacterial phylotype richness in RDS was higher than the other samples, and TNS had the least richness. The most predominant class in the TNS, CTS, and BTS samples is Betaproteobacteria. Cyanobacteria (no_rank) is the most predominant one in the RDS sample. Circa 31 % sequences in TNS were affiliated with the Rhodocyclales order. In the four samples, Aeromonas, Arcobacter, Clostridium, Legionella, Leptospira, Mycobacterium, Pseudomonas, and Treponema genera containing pathogenic bacteria were detected. Characterization of bacterial communities in sediments from various downstream branches indicated that distinct wastewater effluents have similar potential to reduce the natural variability in river ecosystems and contribute to the river biotic homogenization.     

Wastewater Irrigation Increases the Abundance of Potentially Harmful Gammaproteobacteria in Soils in Mezquital Valley,Mexico

Wastewater contains large amounts of pharmaceuticals, pathogens, and antimicrobial resistance determinants. Only a little is known about the dissemination of resistance determinants and changes in soil microbial communities affected by wastewater irrigation. Community DNAs from Mezquital Valley soils under irrigation with untreated wastewater for 0 to 100 years were analyzed by quantitative real-time PCR for the presence of sul genes, encoding resistance to sulfonamides. Amplicon sequencing of bacterial 16S rRNA genes from community DNAs from soils irrigated for 0, 8, 10, 85, and 100 years was performed and revealed a 14% increase of the relative abundance of Proteobacteria in rainy season soils and a 26.7% increase in dry season soils for soils irrigated for 100 years with wastewater. In particular, Gammaproteobacteria, including potential pathogens, such as Pseudomonas, Stenotrophomonas, and Acinetobacter spp., were found in wastewater-irrigated fields. 16S rRNA gene sequencing of 96 isolates from soils irrigated with wastewater for 100 years (48 from dry and 48 from rainy season soils) revealed that 46% were affiliated with the Gammaproteobacteria (mainly potentially pathogenic Stenotrophomonas strains) and 50% with the Bacilli, whereas all 96 isolates from rain-fed soils (48 from dry and 48 from rainy season soils) were affiliated with the Bacilli. Up to six types of antibiotic resistance were found in isolates from wastewater-irrigated soils; sulfamethoxazole resistance was the most abundant (33.3% of the isolates), followed by oxacillin resistance (21.9% of the isolates). In summary, we detected an increase of potentially harmful bacteria and a larger incidence of resistance determinants in wastewater-irrigated soils, which might result in health risks for farm workers and consumers of wastewater-irrigated crops.