Antibiotic Resistance and Transferable R Plasmids in Coliforms from a Polluted Estuary

The aim of this work was to find out how far the Nervion Estuary serves as a reservoir of R plasmids and/or for their dissemination. The fecal pollution bacterial indicator population was estimated along the estuary and 167 coliform isolates were assayed for resistance to 16 antibiotics. About sixty-nine per cent of the coliform isolates were antibiotic-resistant and forty-two per cent of these were simultaneously resistant to three or more of 16 antibiotics tested. Furthermore, 31.5% of enterobacterial strains resistant to any of the antibiotics tested could transfer their resistance determinants in conjugal matings. Most of the transferred plasmids confer resistance to ampicillin or to other β-lactamic antibiotics and several codify resistance to tetracyclin, chloramphenicol of fosfomycin. Taking account of the coliforms density found and the ratio of these isolates able to transfer R plasmids, an average of 5 × 10³ coliform strains able to transfer R plasmids in 100 ml of water was obtained. This amount must indicate a significant potential public health risk because these polluted waters get into an area with intensive fishing and recreational use.     

Antibiotic resistant bacteria in fish from the Concepción Bay, Chile.

Antibiotic resistant bacteria from commercial demersal and pelagic fish captured in the Concepción Bay, Chile were investigated. Viable counts of antibiotic resistant bacteria isolated from gill and intestinal content samples showed high frequencies of resistance to ampicillin, streptomycin and tetracycline, while the proportion of chloramphenicol resistance was rather low. A high incidence of resistance to ampicillin, streptomycin, tetracycline and nitrofurantoin, as well as almost an absence of resistance to gentamicin, amikacin and cotrimoxazole was found among selected isolates which represented the resistant bacterial population. These strains mainly belonged to Vibrionaceae and Enterobacteriaceae and were predominantly resistant to 3 and 4 antibacterials. Isolates from demersal fish exhibited resistance to as many as 8-10 compounds, whereas those from pelagic fish were resistant to seven or fewer antibiotics. These results suggest that Chilean commercial fishes residing in waters near the disposals of urban sewage might play a role as carriers of antibiotic resistant bacteria prompting a health risk to public health for fish consumers.     

Isolates of Staphylococcus aureus and saprophyticus resistant to antimicrobials isolated from the Lebanese aquatic environment

The indiscriminate use of antimicrobials especially in developing countries has evoked serious bacterial resistance and led to the emergence of new and highly resistant strains of bacteria to commonly used antimicrobials. In Lebanon, pollution levels and bacterial infections are increasing at a high rate as a result of inadequate control measures to limit untreated effluent discharges into the sea or freshwater resources. The aim of this study was to isolate and molecularly characterize various Staphylococcus strains isolated from sea water, fresh water, sediments, and crab samples collected from representative communities along the coast of Lebanon. The results on the antimicrobial resistance indicated that the level of resistance of Staphylococcus aureus varied with various antimicrobials tested. The resistance patterns ranged between 45% in freshwater isolates and 54.8% in seawater ones. Fifty one percent of the tested isolates have shown resistance to at least one of the five tested antimicrobials; with seawater isolates exhibiting the highest rates of antimicrobial resistance.     

Gilthead seabream (Sparus aurata) carrying antibiotic resistant enterococci. A potential bioindicator of marine contamination?

Antibiotic resistance in bacteria is a growing problem that is not only restricted to the clinical setting but also to other environments such as marine species that harbor antibiotic resistant bacteria and therefore may serve as reservoirs for antibiotic-resistance genetic determinants. The aim of this study was to evaluate antibiotic resistance phenotypes in enterococci isolated from fecal samples of gilthead seabream and the associated mechanisms of resistance. A collection of 118 samples were analyzed and 73 enterococci were recovered. The strains showed high percentages of resistance to erythromycin and tetracycline (58.9% and 17.8%, respectively). Lower level of resistance (<13%) was detected for quinupristin-dalfopristin, ampicillin, high-level-gentamicin, high-level-streptomycin, high-level-kanamycin, ciprofloxacin and chloramphenicol. The erm(B), tet(L) or tet(M), aac(6′)-aph(2″) and aph(3′)-IIIa genes were shown in isolates resistant to erythromycin, tetracycline, high-level gentamicin and high-level kanamycin, respectively. Antibiotic resistance in natural microbiota is becoming a concern of human and environmental health.     

Antibiotic resistant pathogens versus human impacts: a study from three eco-regions of the Chennai coast, southern India

We assessed the occurrence of pollution indicators and antibiotic resistant bacterial isolates from water and sediment samples of three different eco-regions of the Chennai coast between March - May of 2010. Total of 960 bacterial strains belonging to four genera were isolated which show the highest frequencies of resistance to vancomycin (53.6%) and penicillin (52.6%) (except Enterococcus sp., which is highly resistant to erythromycin) and lowest frequencies of resistance to chloramphenicol (3.43%), ciprofloxacin (3.95%), gentamicin (4.68%), and tetracycline (6.97%). The E. coli, Vibrio sp., Salmonella sp. and Enterococcus sp. show high frequency of resistance to 2-5 antibacterials of 60.4%, 45.83%, 69.16% and 46.6%, respectively. High pollution indices (PI - 6.66-14.06) and antibiotic resistance indices (ARI - 0.29-0.343) indicate that the coastal environment is highly exposed to antibiotic sources that suggesting to avoid direct contact.     

Molecular characterizations of oxytetracycline resistant bacteria and their resistance genes from mariculture waters of China

Oxytetracycline-resistant bacteria were isolated from a mariculture farm in China, and accounted for 32.23% and 5.63% of the total culturable microbes of the sea cucumber and the sea urchin rearing waters respectively. Marine vibrios, especially strains related to Vibrio splendidus or V. tasmaniensis, were the most abundant resistant isolates. For oxytetracycline resistance, tet(A), tet(B) and tet(D) genes were detected in both sea cucumber and sea urchin rearing ponds. The dominant resistance type for V. tasmaniensis-like strains was the combination of both tet(A) and tet(B) genes, while the major resistance type for V. splendidus-like strains was a single tet(D) gene. Most of the sea cucumber tet-positive isolates harbored a chloramphenicol-resistance gene, either cat IV or cat II, while only a few sea urchin tet-positive isolates harbored a cat gene, actually cat IV. The coexistence of tet and cat genes in the strains isolated from the mariculture farm studied was helpful in explaining some of the multi-resistance mechanisms.     

Chromium Removal through Biosorption and Bioaccumulation by Bacteria from Tannery Effluents Contaminated Soil

Four bacterial isolates (two resistant and two sensitive to chromium) were isolated from soil contaminated with tannery effluents at Jajmau (Kanpur), India, and were identified by 16S rDNA gene sequencing as Stenotrophomonas maltophilia, Exiguobacterium sp., Pantoea sp., and Aeromonas sp. Biosorption of chromium by dried and living biomasses was determined in the resistant and sensitive isolates. The effect of pH, initial metal concentration, and contact time on biosorption was studied. At pH 2.5 the living biomass of chromium resistant isolate Exiguobacterium sp. ZM‐2 biosorbed maximum amount of Cr⁶⁺ (29.8 mg/g) whereas the dried biomass of this isolate biosorbed 20.1 mg/g at an initial concentration of 100 mg/L. In case of chromate sensitive isolates, much difference was not observed in biosorption capacities between their dried and living biomasses. The maximum biosorption of Cr³⁺ was observed at pH 4.5. However, biosorption was identical in resistant and sensitive isolates. The data on chromium biosorption were analyzed using Langmuir and Freundlich isotherm model. The biosorption data of Cr⁶⁺ and Cr³⁺ from aqueous solution were better fitted in Langmuir isotherm model compared to Freundlich isotherm model. Metal recovery through desorption was observed better with dried biomasses compared to the living biomasses for both types of chromium ions. Bioaccumulation of chromate was found higher in chromate resistant isolates compared to the chromate sensitive isolates. Transmission electron microscopy confirmed the accumulation of chromium in cytoplasm in the resistant isolates.     

Bacterial flora and antibiotic resistance from eggs of green turtles Chelonia mydas: An indication of polluted effluents

Sea turtles migrate to various habitats where they can be exposed to different pollutants. Bacteria were collected from turtle eggs and their resistance to antibiotics was used as pollutant bio-indicators of contaminated effluents. Eggs were collected randomly from turtles when they were laying their eggs. A total of 90 eggs were collected and placed into sterile plastic bags (3 eggs/turtle) during June-December of 2003. The bacteria located in the eggshell, albumen and yolk were examined, and 42% of the eggs were contaminated with 10 genera of bacteria. Pseudomonas spp. were the most frequent isolates. The albumen was found to be the part of the egg to be the least contaminated by bacterial infection. Bacterial isolates tested with 14 antibiotics showed variations in resistance. Resistance to ampicillin was the highest. The presence of antibiotic resistant bacteria in eggs indicates that the green turtle populations were subjected to polluted effluents during some of their migratory routes and feeding habitats. Scanning electron microscopy revealed that Salmonella typhimurium penetrated all eggshell layers.     

Antimicrobial-resistant coliforms in New York Bight

A preliminary study of sediments and overlying water samples obtained at the approximate centre of the sewage sludge bed in the New York Bight, revealed significant numbers of coliforms resistant to antibiotics and heavy metals. Some of the isolates were shown to transfer antibiotic resistance during conjugation with a recipient strain of Salmonella gallinarum. The selection of bacterial populations resistant to antimicrobials may be significant in the ecology of this impoverished area, and requires further study.     

成都府河水体细菌动态变化及耐药性

将府河从都江堰宝瓶口段至望江公园段分设9个监测点,在秋冬、春夏两季分别取水,首次建立由细菌总数、总大肠菌群数及粪大肠菌群数三个微生物学指标组成的体系,对府河两季水质进行对比监测分析,以综合评价水质、分析季节影响因素并反映出污染物来源.以都江堰宝瓶口大肠杆菌耐药率作为背景值,对整个河段的9个采样点分离纯化鉴定得到102株大肠杆菌,用K-B法进行对比药敏试验.结果表明,府河从都江堰段起,穿过成都市区后,至望江公园段细菌总数、大肠菌群数及粪大肠菌群数虽有波动,但细菌总数总体呈增加趋势,春夏季府河营养富集,水质有所降低.府河中大肠杆菌耐药性较小,β-内酰胺类耐药率在0.96％～33.33％之间,氨基糖苷类1.39％～7.84％,氟喹诺酮类11.43％～11.76％,磺胺类复方新诺明为52.78％,其中β-内酰胺类耐药性、氟喹诺酮类耐药性与源头水质有关,其余与医疗机构排放的废水及生活污水有关,此研究为控制成都府河水体中的污染防控提供了理论依据.     

Antibiotic Resistance of Gram Negatives isolates from loggerhead sea turtles (Caretta caretta) in the central Mediterranean Sea

Previous studies on fish and marine mammals support the hypothesis that marine species harbor antibiotic resistance and therefore may serve as reservoirs for antibiotic-resistance genetic determinants. The aim of this study was to assess the resistance to antimicrobial agents of Gram negative strains isolated from loggerhead sea turtles (Caretta caretta). Oral and cloacal swabs from 19 live-stranded loggerhead sea turtles, with hooks fixed into the gut, were analyzed. The antimicrobial resistance of the isolates to 31 antibiotics was assessed using the disk-diffusion method. Conventional biochemical tests identified Citrobacter spp., Proteus spp., Enterobacter spp., Escherichia spp., Providencia spp., Morganella spp., Pantoea spp., Pseudomonas spp. and Shewanella spp. Highest prevalences of resistance was detected to carbenicillin (100%), cephalothin (92.6%), oxytetracycline (81.3%) and amoxicillin (77.8%). The isolates showing resistance to the widest range of antibiotics were identified as Citrobacter freundii, Proteus vulgaris, Providencia rettgeri and Pseudomonas aeruginosa. In this study, antibiotic resistant bacteria reflect marine contamination by polluted effluents and C. caretta is considered a bioindicator which can be used as a monitor for pollution.     

Mechanism of cadmium resistance and adsorption of a yeast strain Rhodotorula sp. Y11

The mechanism of cadmium resistance of a yeast strain Rhodotorula sp. Y11 isolated from mine soil was investigated. We found that the yeast cells treated with different methods showed different cadmium-adsorption models. Grown in medium supplied with 100 mg/L of cadmium, 3.29% of the cell-absorbed cadmium was accounted in the cytoplasm. However, only 1% was taken into the cytoplasm and 99% was bound to the cell wall using the lyophilized biomass to adsorb cadmium in double distilled water. Treatments with alkali, ethanol-chloroform and proteinase showed different influences on the biosorption of whole cells and isolated cell walls. FT-IR analysis showed that acetyl of chitin was the active compound in the cells to absorb cadmium. The production of Metallothioneins, proteins related to the resistance to heavy metal in yeast, was evidently induced by cadmium, achieving 638.8 μg/g wet weight, which was about 85 folds higher than that in the uninduced biomass and was also much higher than that reported previously. The molecular weight of Metallothioneins was 6500 Da estimated by SDS-PAGE.     

A Comparison of Growth Biomarkers for Assessing Sublethal Effects of Cadmium on a Marine Gastropod, Nassarius festivus

The effect of cadmium on the growth of an intertidal marine gastropod, Nassarius festivus, was investigated using a number of biomarkers namely conventional measurements in terms of increase in shell length or body weight, scope for growth (SfG) and RNA/DNA ratio. After eight days of cadmium exposure, the no observable effect concentration (NOEC) and lowest observable effect concentration (LOEC) for the rate of increase in wet weight were 1.00 and 1.38 mg/l, respectively, while those for shell length did not show any significant difference from the control until day 16. After 16 days of cadmium exposure, the LOEC for both shell length and wet weight measurements was 0.22 mg/l. The 8-day SfG test was sensitive to cadmium stress, showing a significant decrease at 0.16 mg/l cadmium when compared to the control. The NOEC and LOEC of RNA/DNA ratio determination were 0.52 and 0.93 mg/l, respectively. An assessment based on sensitivity and technical complexity suggests that Scope for Growth is the most sensitive growth biomarker, followed by the RNA/DNA ratio, and then the conventional growth measurement based on shell size and body weight.     

Antibiotic Resistance and Plasmid Profiles of Vibrio Isolates from Cultured Silver Sea Bream, Sparus sarba

A total of 51 potential pathogenic vibrios were isolated from moribund silver sea bream (Sparus sarba) collected from fish farms in Hong Kong. Using the API 20E system and the scheme of Alsina and Blanch (1994), 7 species were identified from all isolates. These species were Vibrio alginolyticus (24 strains), Vibrio vulnificus (12 strains), Vibrio parahaemolyticus (7 strains), Vibrio logei (4 strains), Vibrio pelagius II (2 strains), Vibrio fluvialis (1 strain) and Vibrio meditterranei (1 strain). The three dominant species (V. alginolyticus, V. vulnificus and V. parahaemolyticus) were confirmed to be virulent to sea bream by experimental challenge.

All isolates were screened for plasmid DNA by agarose gel electrophoresis and tested for susceptibility to 16 antibiotics by the agar dilution method. Of the 51 isolates examined, all strains were sensitive to ceftriaxone, streptomycin, nalidixic acid and rifampicin, and almost all were sensitive to ceftazidime, netilimicin, chloramphenicol and sulfamethoxazole. Most isolates were resistant to ampicillin (60.8%), cefuroxime (66.7%), amikacin (55%), kanamycin (58.8%) and trimethoprim (76.5%). Fifteen of the 51 isolates harboured 1–4 plasmids, with sizes ranging from 9 to 123 kb. Both the plasmids and the associated antibiotic resistance (ampicillin, cefuroxime and trimethoprim) of 9 isolates could be transferred to a recipient by single-step conjugation. However, the frequencies were very low, ranging from 10⁻¹¹ to 10⁻⁹. The present results indicate that resistance to these antibiotics is chromosomal.     

Molecular characterizations of chloramphenicol- and oxytetracycline-resistant bacteria and resistance genes in mariculture waters of China

In order to gain an understanding of the diversity and distribution of antimicrobial-resistant bacteria and their resistance genes in maricultural environments, multidrug-resistant bacteria were screened for the rearing waters from a mariculture farm of China. Both abalone Haliotis discushannai and turbot Scophthalmus maximus rearing waters were populated with abundant chloramphenicol-resistant bacteria. These bacteria were also multidrug resistant, with Vibriosplendidus and Vibriotasmaniensis being the most predominant species. The chloramphenicol-resistance gene cat II, cat IV or floR could be detected in most of the multidrug-resistant isolates, and the oxytetracycline-resistance gene tet(B), tet(D), tet(E) or tet(M) could also be detected for most of the isolates. Coexistence of chloramphenicol- and oxytetracycline-resistance genes partially explains the molecular mechanism of multidrug resistance in the studied maricultural environments. Comparative studies with different antimicrobial agents as the starting isolation reagents may help detect a wider diversity of the antimicrobial-resistant bacteria and their resistance genes.     

Echinoderms from Azores islands: An unexpected source of antibiotic resistant Enterococcus spp. and Escherichia coli isolates

The prevalence of antibiotic resistance and the implicated mechanisms of resistance were evaluated in Enterococcus spp. and Escherichia coli, isolated from a total of 250 faecal samples of echinoderms collected from Azorean waters (Portugal). A total of 144 enterococci (120 Enterococcus faecium, 14 E. hirae, 8 E. faecalis, 2 E. gallinarum) and 10 E. coli were recovered. High percentages of resistance in enterococci were found for erythromycin, ampicillin, tetracyclin and ciprofloxacin. The erm(A) or erm(B), tet(M) and/or tet(L), vat(D), aac(6′)-aph(2″) and aph(3′)-IIIa genes were found in isolates resistant to erythromycin, tetracycline, quinupristin/dalfopristin, high-level gentamicin and high-level kanamycin, respectively. Resistance in E. coli isolates was detected for streptomycin, amikacin, tetracycline and tobramycin. The aadA gene was found in streptomycin-resistant isolates and tet(A) + tet(B) genes in tetracycline-resistant isolates. The data recovered are essential to improve knowledge about the dissemination of resistant strains through marine ecosystems and the possible implications involved in transferring these resistances either to other animals or to humans.     

Bioremoval of Cadmium by Lemna minor in Different Aquatic Conditions

This study was undertaken to determine the cadmium removal efficiency of Lemna minor when it was used for treatment of wastewater having different characteristics, i. e., pH, temperature and cadmium concentration. Plants were cultivated in different pH solutions (4.5–8.0) and temperatures (15–35°C) in the presence of cadmium (0.1–10.0 mg/L) for 168 h. The amount of biomass obtained in the study period, the concentrations of cadmium in the tissues and in the media and net uptake of cadmium by Lemna have been determined for each condition. The percentages of cadmium uptake (PMU) and bioconcentration factors (BCF) were also calculated. The highest accumulation was obtained for the highest cadmium concentration of 10.0 mg Cd/L as 11.668 mg Cd/g at pH 6.0, and as 38.650 mg Cd/g at 35°C and pH 5.0. The cadmium accumulation gradually increased with initial concentration of the medium, but the opposite trend was observed for the PMU. However, the maximum PMU was obtained as 52.2% in the solution with the lowest concentration of 0.1 mg Cd/L. A mathematical model was used to describe the cadmium uptake and the equation obtained was seen to fit the experimental data very well.     

Role of shellfish hatchery as a reservoir of antimicrobial resistant bacteria

The main aim of this study was to determine the occurrence of resistant bacteria in florfenicol-treated and untreated scallop larval cultures from a commercial hatchery and to characterize some selected florfenicol-resistant strains. Larval cultures from untreated and treated rearing tanks exhibited percentages of copiotrophic bacteria resistant to florfenicol ranging from 0.03% to 10.67% and 0.49–18.34%, respectively, whereas florfenicol resistance among oligotrophic bacteria varied from 1.44% to 35.50% and 3.62–95.71%, from untreated and treated larvae, respectively. Florfenicol resistant microbiota from reared scallop larvae mainly belonged to the Pseudomonas and Pseudoalteromonas genus and were mainly resistant to florfenicol, chloramphenicol, streptomycin and co-trimoxazole. This is the first study reporting antimicrobial resistant bacteria associated to a shellfish hatchery and the results suggest that a continuous surveillance of antimicrobial resistance even in absence of antibacterial therapy is urgently required to evaluate potential undesirable consequences on the surrounding environments.     

Reuse of Drinking Water Treatment Waste for Remediation of Heavy Metal Contaminated Groundwater

Lime softening produces an estimated 10,000 metric tons of dry drinking water treatment wastes (DWTW) per year, costing an estimated one billion dollars annually for disposal worldwide. Lime softening wastes have been investigated for reuse as internal curing agents or supplementary cementitious materials in concrete as well as a high-capacity sorbent for heavy metal removal. Lead, cadmium, and zinc are common heavy metals in groundwater contaminated by mine tailings. Cement-based filter media (CBFM) are a novel material-class for heavy metal remediation in groundwater. This study investigated the incorporation of DWTW as a recycled, low-cost additive to CBFM for the removal of lead, cadmium, and zinc. Jar testing at three different metal concentrations and breakthrough column testing using synthetic groundwater were performed to measure removal capacity and reaction kinetics. Jar testing results show as DWTW content increases at low concentrations, removal approaches 100% but at high metal concentrations removal decreases due to saturation or exhaustion of the removal mechanisms. Removal occurs through the formation of metal carbonate precipitates, surface sorption, and ion exchange with calcium according to the preferential series Pb⁺² > Zn⁺² > Cd²⁺. Removal kinetics were also measured through column testing and exceeded estimated calculations derived from batch jar testing isotherms due to the large formation of oolitic metal carbonates. Lead, cadmium, and zinc was concentrated in the column precipitates from 0.29, 0.23, and 20.0 μg/g in the influent solution to approximately 200, 130, 14,000 μg/g in the reacted DWTW-CBFM. The control and DWTW-CBFM columns had statically similar removal for zinc and lead. In the DWTW-CBFM, cadmium had decreased removal of approximately 25% due to proportionately decreased hydroxide content from cement replacement with 25% DWTW. This study shows the potential for DWTW as an enhancement to CBFM, thereby valorizing an otherwise waste material. Furthermore, the concentrative abilities of CBFM through precipitate and oolitic mineral formation could provide a minable waste product and close the waste-product cycle for DWTW.     

Drug-resistance transfer in Escherichia coli in New York Bight sediment

Sediment from the sewage sludge dump site area in the New York Bight contains bacteria resistant to antibiotics and heavy metals. The purpose of this study was to determine if antibiotic resistance could be transferred from donor to recipient Escherichia coli strains inoculated into glass vessels containing sediment and seawater obtained near the dump site. Temperature was maintained at 10°C, the mean winter temperature of benthic water at the dump site. Transconjugants (recipients which inherit donor genes for tetracycline resistance) were isolated from the seawater and sediment within one hour after inoculation and were found in all subsequent sediment samples for one month. Donor and recipient E. coli remained viable in the sediment for at least one month. Our results indicate that sewage sludge polluted sediment may serve as an environment conducive to conjugal transfer of antibiotic resistance genes.