Antibacterial agents and heavy metal resistance in Gram-negative bacteria isolated from seawater, shrimp and sediment in Iskenderun Bay, Turkey

The aim of the present study was to determine the level of antibiotic resistance patterns and distribution of heavy metal resistance of bacterial isolates from seawater, sediment and shrimps, and to determine if there is a relationship between antibiotic and heavy metal resistance. We undertook studies in 2007 in the industrially polluted Iskenderun Bay, on the south coast of Turkey. The resistance of 236 Gram-negative bacterial isolates (49 from seawater, 90 from sediment and 97 from shrimp) to 16 different antibiotics, and to 5 heavy metals, was investigated by agar diffusion and agar dilution methods, respectively. A total of 31 species of bacteria were isolated: the most common strains isolated from all samples were Escherichia coli (11.4%), Aeromonas hydrophila (9.7%) and Stenotrophomonas maltophilia (9.3%). There was a high incidence of resistance to ampicillin (93.2%), streptomycin (90.2%) and cefazolin (81.3%), and a low incidence of resistance to imipenem (16.5%), meropenem (13.9%) and cefepime (8.0%). Some 56.8% of all bacteria isolated from seawater, sediment and shrimp were resistant to 7 or more antibiotics. Most isolates showed tolerance to different concentrations of heavy metals, and minimal inhibition concentrations ranged from 12.5 μg/ml to > 3200 μg/ml. The bacteria from seawater, sediment and shrimp showed high resistance to cadmium of 69.4%, 88.9%, and 81.1% respectively, and low resistance to manganese of 2%, 6.7% and 11.3% respectively. The seawater and sediment isolates which were metal resistant also showed a high resistance to three antibiotics: streptomycin, ampicillin and trimethoprim-sulphamethoxazole. In contrast, the shrimp isolates which were metal resistant were resistant to four antibiotics: cefazolin, nitrofurantoin, cefuroxime and ampicillin. Our results show that Iskenderun Bay has a significant proportion of antibiotic and heavy metal resistant Gram-negative bacteria, and these bacteria constitute a potential risk for public health.     

Incidence of heavy metal and antibiotic resistance in bacterial isolates from Blue Nile river water in Ethiopian region

Heavy metal pollution of river water has health implications. This study was aimed to test if bacteria that are heavy metal resistant can resist antibiotics. Heavy metal-resistant bacteria were isolated using Nutrient agar plates amended with 100?μg?ml^?1 of metal salts. Isolates were further screened to determine minimal inhibitory concentration against each metal. The isolates belonged to the genera of Neisseria, Bacillus, Pseudomonas, Staphylococcus, Streptococcus, Micrococcus, Corynebacterium and Proteus. Minimum inhibitory concentration of the isolates for heavy metals ranged from 200 to 2300?μg?ml^?1. Isolates of Pseudomonas, Neisseria, Bacillus, Protues, Streptococcus, Staphylococcus and Micrococcus spp. exhibited multiple antibiotic resistance. Statistically significant correlation between metal resistance and antibiotic resistance was observed among bacterial isolates. The present work suggests that some of the isolates could be hazardous to human beings as a result of natural selection of dual resistant mechanisms.     

Antibiotic resistance and tolerance to heavy metals demonstrated by environmental pathogenic bacteria isolated from the Kahwa River,Bukavu Town,Democratic Republic of the Congo

Increasing concentration of heavy metals and antibiotics within bacterial microenvironments contribute to medical problems. This research confirmed the presence of enteropathogenic bacteria in the sediment and water of Kahwa River. Vibrio cholerae, Shigella dysenteriae, Salmonella typhi and Salmonella paratyphi showed resistance to antibiotics and tolerance to heavy metals. The bacteria, particularly Vibrio, were extensively resistant to Ampicillin; less so to Chloramphenicol, Co-trimoxazole and Tetracycline with a relatively reduced resistance shown to Ciprofloxacin. Higher levels of antibiotic resistance were noticed in bacteria isolated from river sediment when compared to bacteria isolated from river water (p < 0.05). Tolerance to cadmium and lead was also more obvious in bacteria isolated from river sediment. An increasing association was observed between multiple antibiotic resistance and tolerance to heavy metals. Bacterial growth in the presence of cadmium was restricted when compared to lead.     

Wastewater treatment contributes to selective increase of antibiotic resistance among Acinetobacter spp.

The occurrence and spread of multi-drug resistant bacteria is a pressing public health problem. The emergence of bacterial resistance to antibiotics is common in areas where antibiotics are heavily used, and antibiotic-resistant bacteria also increasingly occur in aquatic environments. The purpose of the present study was to evaluate the impact of the wastewater treatment process on the prevalence of antibiotic resistance in Acinetobacter spp. in the wastewater and its receiving water. During two different events (high-temperature, high-flow, 31 °C; and low-temperature, low-flow, 8 °C), 366 strains of Acinetobacter spp. were isolated from five different sites, three in a wastewater treatment plant (raw influent, second effluent, and final effluent) and two in the receiving body (upstream and downstream of the treated wastewater discharge point). The antibiotic susceptibility phenotypes were determined by the disc-diffusion method for 8 antibiotics, amoxicillin/clavulanic acid (AMC), chloramphenicol (CHL), ciprofloxacin (CIP), colistin (CL), gentamicin (GM), rifampin (RA), sulfisoxazole (SU), and trimethoprim (TMP). The prevalence of antibiotic resistance in Acinetobacter isolates to AMC, CHL, RA, and multi-drug (three antibiotics or more) significantly increased (p < 0.01) from the raw influent samples (AMC, 8.7%; CHL, 25.2%; RA, 63.1%; multi-drug, 33.0%) to the final effluent samples (AMC, 37.9%; CHL, 69.0%; RA, 84.5%; multi-drug, 72.4%), and was significantly higher (p < 0.05) in the downstream samples (AMC, 25.8%; CHL, 48.4%; RA, 85.5%; multi-drug, 56.5%) than in the upstream samples (AMC, 9.5%; CHL, 27.0%; RA, 65.1%; multi-drug, 28.6%). These results suggest that wastewater treatment process contributes to the selective increase of antibiotic resistant bacteria and the occurrence of multi-drug resistant bacteria in aquatic environments.     

Microbial contamination of main contact surfaces of Automated Teller Machines from Metropolitan Area of Porto

ABSTRACT

Automated Teller Machines (ATM) are likely to be contaminated with various microorganisms because of their contact with the hands of many users daily. The main objective of this study was to investigate ATMs as a potential source of bacterial contamination. This study was conducted in the Metropolitan Area of Porto, in which 50 swab samples were cultured on selective media. Some isolates were identified based on colonial, morphological and biochemical characteristics. Susceptibility to several antibiotics was also evaluated for each isolated organism. Most contaminated ATMs had high numbers of Enterobacteriaceae and Enterococci. Staphylococci and Enterobacteriaceae were the most prevalent bacteria found. The recovered bacterial isolates varied in their antibiotic resistance pattern; Staphylococcus spp. demonstrated the most resistant profiles. With this pioneering study in Portugal, it was demonstrated that although ATMs in metropolitan area of Porto were not highly contaminated, some potentially pathogenic bacteria were present and resistant to some commonly used antibiotics.     

Antimicrobial resistance of heterotrophic bacteria in sewage-contaminated rivers

Sewage-contaminated rivers are ecosystems deeply disturbed by human activity due to the release of heavy metals, organic pollutants and pharmaceuticals as well as faecal and pathogenic micro-organisms, which coexist with the autochthonous microbial population. In this study, we compared the percentage of resistance in faecal and heterotrophic bacteria in rivers with different degrees of sewage pollution. As a matter of fact, no correlation was found neither between the degree of sewage pollution and the percentage of antimicrobial resistant heterotrophic bacteria nor between the number of resistant faecal bacteria and that of resistant heterotrophic bacteria. Most of the resistant isolates from the Zenne river downstream Brussels were multi-resistant and the resistance patterns were similar among the strains of each phylogenetic group. The total microbial community in this polluted river (as evaluated through a 16S rRNA gene clone library analysis) appeared to be dominated by the phyla Proteobacteria and Bacteroidetes while the phylum TM7 was the third most represented.     

Antibiotic resistance in bacteria from shrimp farming in mangrove areas

Shrimp farming is a sufficiently large and mature industry to have an effective range of antimicrobial agents for most bacterial diseases in shrimp culture. However, at present, there exists great concern over the widespread use of antibiotics in aquaculture, which may result in residue of antibiotics in water and mud, and subsequently, the development of antibiotic resistance in bacteria in the environment. There is limited understanding about the effect of antibiotic residues on bacteria resistance in shrimp farming environment. Therefore, a study was conducted to investigate bacterial resistance to Norfloxacin (NFXC), Oxolinic Acid (OXLA), Trimethoprim (TMP) and Sulfamethoxazole (SMX), which were found in four shrimp farming locations in mangrove areas in Vietnam. Findings indicate that there is a relatively high incidence of bacteria resistance to these antibiotics observed in most of the studied sites, particularly to antibiotics with concentration of 0.1 microg/ml. Yet the relation between concentration of antibiotic residues and incidence of antibiotic resistance is not clearly defined. Among individual antibiotics, the incidence of resistance to TMP and SMX was higher than the others. Identification of bacteria isolated from mud samples by DNA analyzer shows that Bacillus and Vibrio are predominant among bacteria resistant to the antibiotics. The result of the study also indicates that these antibiotics in media degraded more rapidly due to the presence of resistant bacteria.     

High diversity and abundance of antibiotic-resistant Escherichia coli isolated from humans and farm animal hosts in Jeonnam Province, South Korea

The spread of antibiotics resistance among bacteria is a threat to human health. Since South Korea uses approximately 1.5 times more antibiotics than do other OECD countries, this is likely to impact the numbers and types of antibiotic-resistant bacteria found in the environment. In this study we examined feces from domesticated animals and humans for the diversity and abundance of antibiotic-resistant Escherichia coli. Abundant antibiotic-resistant E. coli were isolated from all the tested animals and humans and were examined by horizontal, fluorophore-enhanced, rep-PCR (HFERP) DNA fingerprint analysis. A total of 793 unique, non-clonal, E. coli isolates were obtained from the 513 human and animal hosts examined. Antibiotic resistance analysis, done using 14 antibiotics, indicated that 72.3% of the isolates (573 of 793) were found resistant to more than one antibiotic. The E. coli isolated from swine were resistant to the greatest number of antibiotics. Tetracycline resistant E. coli were routinely isolated from all animal hosts (36 to 77% per host), except for dairy cattle (9.3%). Twenty nine E. coli isolates from all hosts, except for duck, were resistant to more than 10 antibiotics. Gene transfer and southern hybridization studies revealed that resistance to 13 of the antibiotics was self-transmissible, and likely mediated by plasmids and integrons. Since genetically diverse and numerically abundant antibiotic-resistant E. coli were consistently recovered from chicken, swine and other domesticated animals in South Korea, our results suggest that the use of sub-therapeutic levels of antibiotics for disease prophylaxis and growth promotion should be curtailed.     

Microbial and antibiotic resistant constituents associated with biological aerosols and poultry litter within a commercial poultry house

Poultry are known to harbor antibiotic resistant and pathogenic bacteria, and as such poultry litter and poultry house air can be contaminated with these bacteria. However, the presence of antibiotic resistant bacteria in biological aerosols and litter is largely not understood. The purpose of this study was to determine the amount of aerosolized bacteria and endotoxin, particularly fecal indicators, staphylococci, and enterococci, associated with poultry house and outdoor air. Aerosol samples were collected at multiple locations on the farm and in the house. Antibiotic resistance was investigated using the Kirby Bauer method on selected isolates using twelve different antibiotics spanning both narrow to broad spectrums of effectiveness. Overall there was a cyclical increase in bacterial concentrations as flocks progressed from pre-flock to late-flock, with > 2 orders magnitude lower concentration during pre-flock periods (no chickens), in both the litter and aerosol samples. The house environment provided for significantly concentrated bacterial and endotoxin levels. It was estimated that Staphylococcus bacteria accounted for at least 90% of cultured aerobic bacteria and culture-independent 16S rRNA analyses demonstrated that significant population changes occurred from pre- to late-flock. Rarely was an isolate resistant to more than 4 antibiotic classes; however there was a trend upwards in overall resistance of enterococci as the flock cycle progressed. It appears that although levels of antibiotic resistant bacteria were highly concentrated within the house, levels were much lower outside of the house, and very little house escape occurred.     

Multiresistant Enterobacteriaceae with class 1 and class 2 integrons in a municipal wastewater treatment plant

In this study, 1832 strains of the family Enterobacteriaceae were isolated from different stages of a municipal wastewater treatment plant, of which 221 (12.1%) were intI-positive. Among them 61.5% originated from raw sewage, 12.7% from aeration tank and 25.8% from the final effluent. All of the intI-positive strains were multiresistant, i.e. resistant to at least three unrelated antimicrobials. Although there were no significant differences in resistance range, defined as the number of antimicrobial classes to which an isolate was resistant, between strains isolated from different stages of wastewater treatment, for five β-lactams the percentage of resistant isolates was the highest in final effluent, which may reflect a selective pressure the bacteria are exposed to, and the possible route of dissemination of β-lactam resistant strains to the corresponding river. The sizes of the variable part of integrons ranged from 0.18 to 3.0 kbp and contained up to four incorporated gene cassettes. Sequence analysis identified over 30 different gene cassettes, including 24 conferring resistance to antibiotics. The highest number of different gene cassettes was found in bacteria isolated from the final effluent. The gene cassettes were arranged in 26 different resistance cassette arrays; the most often were dfrA1-aadA1, aadA1, dfrA17-aadA5 and dfrA12-orfF-aadA2. Regarding the diversity of resistance genes and the number of multiresistant bacteria in the final effluent, we concluded that municipal sewage may serve as a reservoir of integron-embedded antibiotic resistance genes.     

Bottled mineral water as a potential source of antibiotic resistant bacteria

The antibiotic resistance phenotypes of the cultivable bacteria present in nine batches of two Portuguese and one French brands of commercially available mineral waters were examined. Most of the 238 isolates recovered on R2A, Pseudomonas Isolation agar or on these culture media supplemented with amoxicillin or ciprofloxacin, were identified (based on 16S rRNA gene sequence analysis) as Proteobacteria of the divisions Beta, Gamma and Alpha. Bacteria resistant to more than three distinct classes of antibiotics were detected in all the batches of the three water brands in counts up to 10² CFU/ml. In the whole set of isolates, it was observed resistance against all the 22 antimicrobials tested (ATB, bioMérieux and disc diffusion), with most of the bacteria showing resistance to three or more classes of antibiotics. Bacteria with the highest multi-resistance indices were members of the genera Variovorax, Bosea, Ralstonia, Curvibacter, Afipia and Pedobacter. Some of these bacteria are related with confirmed or suspected nosocomial agents. Presumable acquired resistance may be suggested by the observation of bacteria taxonomically related but isolated from different brands, exhibiting distinct antibiotic resistance profiles. Bottled mineral water was confirmed as a possible source of antibiotic resistant bacteria, with the potential to be transmitted to humans.     

Bacterial community characteristics under long-term antibiotic selection pressures

To investigate bacterial community characteristics under long-term antibiotic selection pressures, water samples from the upstream and the downstream sections of two rivers individually receiving the treated penicillin G and oxytetracycline production wastewater, as well as the anaerobic and the aerobic effluent of the penicillin G production wastewater treatment plant, were taken and analyzed. Antibiotic resistance ratios of bacterial communities in water samples were estimated by culture-based analysis. The majority of bacterial colonies (approximately 55%-70%) in both downstream rivers and the aerobic effluent showed resistance to 80 μg/ml of antibiotics tested, while the resistance ratios were less than 10% and 5% respectively for both upstream rivers. Six 16S rRNA gene clone libraries were constructed with 355 sequences and 215 OTUs totally obtained representing 465 clones. The antibiotic stresses seemed not reduce the diversities of bacterial communities in antibiotic containing water samples compared to those in the two reference upstream rivers. Bacterial groups present in the two reference upstream rivers were common residents in freshwater ecosystems, with the dominant groups as the phyla Proteobacteria including Alphaproteobacteria, Betaproteobacteria and Gammaproteobacteria, as well as Actinobacteria and Bacteroidetes. The phyla Proteobacteria and Firmicutes were dominant in all antibiotic containing water samples, with the clones belonged to Deltaproteobacteria and Epsilonproteobacteria significantly abundant, as well as Gram-positive low GC bacteria in the classes Clostridia and Bacilli. It thus seemed that Deltaproteobacteria, Epsilonproteobacteria, Clostridia and Bacilli might be specifically associated with antibiotic containing environments.     

Antibiotic resistant enterococci and staphylococci isolated from flies collected near confined poultry feeding operations

Use of antibiotics as feed additives in poultry production has been linked to the presence of antibiotic resistant bacteria in farm workers, consumer poultry products and the environs of confined poultry operations. There are concerns that these resistant bacteria may be transferred to communities near these operations; however, environmental pathways of exposure are not well documented. We assessed the prevalence of antibiotic resistant enterococci and staphylococci in stored poultry litter and flies collected near broiler chicken houses. Drug resistant enterococci and staphylococci were isolated from flies caught near confined poultry feeding operations in the summer of 2006. Susceptibility testing was conducted on isolates using antibiotics selected on the basis of their importance to human medicine and use in poultry production. Resistant isolates were then screened for genetic determinants of antibiotic resistance. A total of 142 enterococcal isolates and 144 staphylococcal isolates from both fly and poultry litter samples were identified. Resistance genes erm(B), erm(A), msr(C), msr(A/B) and mobile genetic elements associated with the conjugative transposon Tn916, were found in isolates recovered from both poultry litter and flies. Erm(B) was the most common resistance gene in enterococci, while erm(A) was the most common in staphylococci. We report that flies collected near broiler poultry operations may be involved in the spread of drug resistant bacteria from these operations and may increase the potential for human exposure to drug resistant bacteria.     

Novel homologs of the multiple resistance regulator marA in antibiotic-contaminated environments

Antibiotics are commonly detected in the environment as contaminants. Exposure to antibiotics may induce antimicrobial-resistance, as well as the horizontal transfer of resistance genes in bacterial populations. We selected the resistance gene marA, mediating resistance to multiple antibiotics, and explored its distribution in sediment and water samples from surface and sewage treatment waters. Ciprofloxacin and ofloxacin (fluoroquinolones), sulphamethoxazole (sulphonamide), erythromycin, clarythromycin, and spiramycin (macrolides), lincomycin (lincosamide), and oxytetracycline (tetracycline) were measured in the same samples to determine antibiotic contamination. Bacterial populations from environmental samples were challenged with antibiotics to identify resistant isolates. The gene marA was found in almost all environmental samples and was confirmed by PCR amplification in antibiotic-resistant colonies. 16S rDNA sequencing revealed that the majority of resistant isolates belonged to the Gram-positive genus Bacillus, not previously known to possess the regulator marA. We assayed the incidence of marA in environmental bacterial populations of Escherichia coli and Bacillus by quantitative real-time PCR in correlation with the levels of antibiotics. Phylogenetic analysis indicated the possible lateral acquisition of marA by Bacillus from Gram-negative Enterobacteriaceae revealing a novel marA homolog in Bacillus. Quantitative PCR assays indicate that the frequency of this gene in antropised environments seems to be related to bacterial exposure to water-borne antibiotics.     

Antimicrobial resistance of fecal bacteria in waters of the Seine river watershed (France)

We studied the prevalences of antimicrobial resistance (AR) and multiple antimicrobial resistance (MAR) among the fecal bacteria found in the rivers of a large watershed under strong anthropogenic pressures, the Seine river watershed (France). Two groups of fecal indicator bacteria, Escherichia coli and intestinal enterococci, were tested for their susceptibility to 16 and 10 antimicrobials respectively, using the disk diffusion method. We found that 42% of the 214 E. coli river isolates were AR (resistant to at least one antimicrobial) and 35% were MAR (resistant to at least two antimicrobials). Among the 148 intestinal enterococci isolates from rivers, 83% were AR and 49% were MAR. We also investigated the sources of AR fecal bacteria found in the rivers of the watershed. A total of 715 E. coli isolates and 476 intestinal enterococci isolates were collected in point sources (municipal and hospital wastewaters) and non-point sources (surface runoff and soil leaching waters from agricultural or forest areas). For E. coli, the prevalence of AR differed widely from source to source and ranked in this order: hospital wastewaters (71%) > municipal wastewaters (44%) > agricultural non-point sources (16%) > forest non-point sources (2%). The prevalence of MAR ranked similarly, and the same trend was observed for intestinal enterococci. The AR level of fecal bacteria in the sources was related to their expected exposure level to antimicrobials before their release into the environment. A MAR index was calculated for every source and a good discrimination between them was thus obtained. At the global scale of the Seine river watershed, domestic wastewaters seemed more likely to be the predominant source of the AR fecal bacteria found in the rivers. This was corroborated by the similarity of the MAR indices from river and municipal wastewater isolates for both fecal indicators.     

Zinc-induced antibiotic resistance in activated sludge bioreactors

Increased levels of bacterial resistance to antibiotics noted in recent decades poses a significant obstacle to the effective treatment and prevention of disease. Although overuse of antibiotics in agriculture and medicine is partially responsible, environmental exposure to heavy metals may also contribute to antibiotic resistance, even in the absence of antibiotics themselves. In this study, a series of eight lab-scale activated-sludge reactors were amended with Zn and/or a suite of three antibiotics (oxytetracycline, ciprofloxacin, and tylosin), in parallel with unamended controls. Classical spread-plating methods were used to assess resistance to these compounds in culturable bacteria over 21 weeks. After seven weeks of general acclimation and development of baseline resistance levels (phase 1), 5.0 mg/L Zn was added to half of the reactors, which were then operated for an additional 7 weeks (phase 2). For the final seven weeks (phase 3), two of the Zn-amended reactors and two of the control reactors were amended with all three antibiotics, each at 0.2 mg/L. Zn amendment alone did not significantly change resistance levels at the 95% confidence level in phase 2. However, tylosin resistance increased significantly during phase 3 in the Zn-only reactors and resistance to all three antibiotics significantly increased as a consequence of combined Zn+antibiotic amendments. Ambient dissolved Zn levels in the reactors were only 12% of added levels, indicating substantial Zn removal by adsorption and/or precipitation. These results show that sub-toxic levels of Zn can cause increased antibiotic resistance in waste treatment microbial communities at comparatively low antibiotic levels, probably due to developed cross-resistance resulting from pre-exposure to Zn.     

Antibiotic resistance and antibacterial activity in heterotrophic bacteria of mineral water origin

Antibiotic resistance and antibacterial activity were determined on heterotrophic bacteria isolated from mineral waters. Of the 120 isolates Pseudomonas spp. (55.8%) was the predominant group followed by Acinetobacter spp. (14.17%), Flavobacterium spp. (10.83%), Achromobacter spp. (10%), Burkholderia cepacia (3.3%), Agrobacterium/radiobacter (2.5%), Moraxella spp. (1.7%), Aeromonas hydrophila (1.7%). Over 80% of the isolates were resistant to one or more antibiotics and the highest resistance was found for chloramphenicol, ampicillin, colistin and sulfamethizole (60%, 55%, 50% and 47.5%, respectively). Strains with multiple antibiotic resistance (MAR) represented 55% of isolates and the most resistant organism belonged to the genus Pseudomonas. Of 40 randomly selected strains, 27 (67.5%) had antibacterial activity towards one or more indicators. This activity, found in a high percentage in the genus Pseudomonas (92%), emerged mainly against closely related microorganisms. Several producers were active also against Escherichia coli, Salmonella, Listeria monocytogenes and Staphylococcus aureus. Forty-six percent of the isolates harboured 1 to 5 plasmids with molecular weights ranging from 2.1 to 41.5 MDa.     

Interactions of heavy metals with bacteria

The toxicity of heavy metals to bacteria, with particular reference to metal forms and species, has been reviewed. Factors which influence metal forms and thus their potential toxicity, such as pH, chelation and competitive interactions have been discussed. The mechanisms whereby bacteria may influence the forms of heavy metals to which they are exposed have been discussed with reference to the importance of the role of bacteria in immobilisation and environmental cycling of metals. Bacterial resistance to metal toxicity is an environmentally important phenomenon. It may occur from non-specific mechanisms, such as impermeability of the cell, or it may be due to specific resistance transfer factors. The coincidence and co-selection of resistance factors for antibiotics and heavy metals in bacterial populations and the clinical implications of this have been described.     

Microbial community functional structure in response to antibiotics in pharmaceutical wastewater treatment systems

It is widely demonstrated that antibiotics in the environment affect microbial community structure. However, direct evidence regarding the impacts of antibiotics on microbial functional structures in wastewater treatment systems is limited. Herein, a high-throughput functional gene array (GeoChip 3.0) in combination with quantitative PCR and clone libraries were used to evaluate the microbial functional structures in two biological wastewater treatment systems, which treat antibiotic production wastewater mainly containing oxytetracycline. Despite the bacteriostatic effects of antibiotics, the GeoChip detected almost all key functional gene categories, including carbon cycling, nitrogen cycling, etc., suggesting that these microbial communities were functionally diverse. Totally 749 carbon-degrading genes belonging to 40 groups (24 from bacteria and 16 from fungi) were detected. The abundance of several fungal carbon-degrading genes (e.g., glyoxal oxidase (glx), lignin peroxidase or ligninase (lip), manganese peroxidase (mnp), endochitinase, exoglucanase_genes) was significantly correlated with antibiotic concentrations (Mantel test; P < 0.05), showing that the fungal functional genes have been enhanced by the presence of antibiotics. However, from the fact that the majority of carbon-degrading genes were derived from bacteria and diverse antibiotic resistance genes were detected in bacteria, it was assumed that many bacteria could survive in the environment by acquiring antibiotic resistance and may have maintained the position as a main player in nutrient removal. Variance partitioning analysis showed that antibiotics could explain 24.4% of variations in microbial functional structure of the treatment systems. This study provides insights into the impacts of antibiotics on microbial functional structure of a unique system receiving antibiotic production wastewater, and reveals the potential importance of the cooperation between fungi and bacteria with antibiotic resistance in maintaining the stability and performance of the systems.     

Antimicrobial multiresistance in bacteria isolated from freshwater Chilean salmon farms

The intensive use of antimicrobial agents, mainly oxytetracycline, to prevent and control bacterial pathologies in Chilean salmon culture is a frequent practice. A total of 103 gram-negative oxytetracycline-resistant bacteria recovered from various sources of 4 Chilean freshwater salmon farms were identified and investigated for their susceptibility patterns to various antibacterial agents, by using an agar disk diffusion method. Antibacterial resistance patterns of isolates were not correlated with bacterial species or strain source. A high number of bacteria resistant to amoxicillin, ampicillin. erythromycin, and furazolidone, as well as an important frequency of bacterial resistance to florfenicol, chloramphenicol, cefotaxime and trimethoprim-sulfamethoxazole was found. On the contrary, the proportion of bacteria resistant to gentamicin, kanamycin, flumequine and enrofloxacin was rather low. Resistant microflora showed a high taxonomic variability and mainly consisted of non-fermenting bacteria (77.7%). These strains mainly belonged to the species Pseudomonas fluorescens (29), Aeromonas hydrophila (10), Stenotrophomonas maltophilia (6), isolated from salmon fingerlings, and Acinetobacter lwoffii (5) isolated from pelletized feed. The occurrence of simultaneous resistance to various antibacterials was frequent. We observe a high frequency of bacteria resistant to 6-10 antibacterials (74 strains), and antibiotic resistance index (ARI) values ranging from 0.38 to 0.48 for the four salmon farms studied. These results suggest that Chilean salmon farms might play a role as reservoirs of antibacterial multiresistant bacteria, thus prompting the necessity for a more restrictive attitude towards the intensive use of antibacterials in salmon farming.