Fecal coliforms of wastewater treatment plants: antibiotic resistance, survival on surfaces and inhibition by sodium chloride and ascorbic acid.

Three hundred and sixteen fecal coliform strains isolated from raw sewage and final effluents of two representative wastewater treatment plants were examined for antibiotic resistance. For plant A (which was disinfecting its effluents before discharge), 55% and 41% of the strains isolated from the influent and the effluent, respectively were resistant to one or more antibiotics. For plant B (which was not disinfecting its effluents before discharge), 33% and 58% of the strains isolated from the influent and the effluent, respectively were resistant to one or more antibiotics. A considerable proportion of these bacteria were resistant to three or more antibiotics. NaCl (3%) and ascorbic acid (0.1%) reduced the growth rate of both sensitive and resistant strains, and more inhibitory effects on the sensitive strains than on the resistant strains were observed. All resistant strains tested survived on various types of surfaces (e.g. glass, stainless steel, agar, cabbage, parsley and sand) significantly better than sensitive strains.     

Antimicrobial susceptibility of non-O1 Vibrio cholerae isolated from wastewater stabilization ponds in Marrakesh,Morocco

The 365 strains of Vibrio cholerae, isolated in Marrakesh from raw sewage and stabilization pond effluent, were all identified as non-O1 Vibrio cholerae. When tested for their susceptibilities to ampicillin, amoxicillin, cephalothin, streptomycin, novobiocin, chloramphenicol, nalidixic acid and trimethoprim-sulphamethoxazole, 13% of the strains from raw sewage and 20% of those from stabilization pond effluent were found to be resistant to one or more of the antibiotics. There were no significant differences, in terms of drug resistance, between isolates from the new sewage and those from the ponds' effluent.The authors are with the Université Cadi Ayyad, Faculté des Sciences Semialia, Département de Biologie, Laboratoire de Microbiologie, BP S/15, Marrakesh, Morocco     

Effect of wastewater stabilization ponds on antimicrobial susceptibility and haemolysin occurrence among motile Aeromonas strains

Aeromonas strains (total=953) isolated from raw wastewater, stabilization pond effluent and sediments were evaluated for their susceptibilities to 17 antibiotics and for their ability to produce haemolysins. Stabilization ponds did not seem to select highly resistant strains of aeromonads. There were no differences in the resistance patterns of isolates from raw sewage, stabilization pond effluent and sediments. All strains were found to possess multiple resistance, most commonly to ampicillin, amoxicillin and novobiocin. Almost 90% of the strains of A. hydrophila and A. caviae were resistant to cephalothin, whereas more than 80% of A. sobria isolates were found to be susceptible to this antibiotic. Resistance to trimethoprim, oxytetracycline, nalidixic acid, chloramphenicol, tetracycline, trimethoprim-sulphamethoxazol, polymyxin B, kanamycin or erythromycin among all isolates did not exceed 10%. Moreover, no strain was found to be resistant to gentamycin and only 9 of the 953 isolates exhibited resistance to cefotaxim. The percentage of haemolytic strains was significantly higher in the stabilization pond effluent than in raw sewage. This high incidence of haemolytic activity was connected with a high proportion of A. sobria whereas, in samples from the raw sewage or stabilization pond sediments a high proportion of A. caviae decreased the total amount of haemolytic aeromonads. The high incidence of haemolytic activity (+) was associated particularly with A. sobria (93.3%) and A. hydrophila (88.7%) whereas A. caviae was found to be the lowest haemolytic species (16.3%).B. Imziln and Y.M.O. Lafdal are with Cadi Ayyad University, Faculty of Sciences Semialia, Department of Biology, Laboratory of Microbiology, BP S/15 Marrakech, Morocco. M. Jana is with the Hôpital Millitaire Avicenne Marrakech, Morocco.     

Phylogenetic groups,virulence genes and quinolone resistance of integron-bearing <Emphasis Type="Italic">Escherichia coli</Emphasis> strains isolated from a wastewater treatment plant

We investigated phylogenetic affiliation, occurrence of virulence genes and quinolone resistance in 109 integron-containing strains of Escherichia coli isolated from a wastewater treatment plant. Selection for integron-bearing strains caused a shift toward phylogroup D, which was most numerous, followed by A, B1 and B2. Phylogroups D and B2, both of which are reported to include virulent extraintestinal pathotypes, made up 50.5% of all isolates and were present in every stage of wastewater treatment, including final effluent. Diarrheagenic pathotypes made up 21% of the strains. The average virulence factor genes score was low (1.40) and the range was from 0 to 5. Quinolone and fluoroquinolone resistance was observed in 56.0% and 50.4% of the strains, respectively; however, it was not associated with virulence factor score. Although the average virulence factor score was low, 17.4% of strains had three and more virulence genes. They were isolated mostly from raw sewage, but 30% of them were cultured from final effluent. Release of multiresistant integron-bearing E. coli strains with virulence traits into the environment may create potential threat and be of public health concern.     

Antibiotic resistance patterns of fecal coliforms isolated from domestic sewage before and after treatment in an aerobic lagoon.

The resistance of 260 strains of fecal coliforms, isolated from raw domestic sewage and aerobic lagoon effluent, to ampicillin, aureomycin, chloromycetin, gentamicin, streptomycin, sulfadiazine, and tetracycline, was determined. Aerobic lagoon treatment produced a 20-fold reduction in the fecal coliform numbers. No statistically significant difference in antibiotic resistance was observed between the fecal coliforms found in raw sewage and in lagoon effluent despite a trend towards the loss of resistance in the latter. Antibiotic resistance, either single or multiple, did not contribute to, or detract from, bacterial survival in the aerobic lagoon. Of the isolates, 15% showed resistance to two or more antiobiotics.     

Microbial Ecology of Activated Sludge: I. Dominant Bacteria

Over 300 bacterial strains were isolated from seven samples of activated sludge by plating on sewage agar. Gram-negative bacteria of the genera Zoogloea and Comamonas predominated. Many isolates (51%) showed sudanophilic inclusions of poly-β-hydroxybutyric acid, whereas 34% accumulated iodophilic material on media containing starch. A large number required either vitamins or amino acids, or both, for growth. None of the isolates tested for their ability to bring about changes in autoclaved sewage produced an effluent comparable in quality to the activated sludge control, although the Zoogloea did produce activated sludgelike flocs. A study of 150 bacterial strains isolated from raw sewage revealed that they differed from the sludge isolates in several respects. Coliforms, which constitute nearly a quarter of the sewage isolates, were rarely encountered in sludge.     

Multiple antibiotic resistances of <Emphasis Type="Italic">Enterococcus</Emphasis> isolates from raw or sand-filtered sewage

Fifty antibiotic-resistant Enterococcus strains were isolated from raw sewage of a wastewater treatment plant and from the same sewage after trickling through a 25-cm sand column, which retained >99% of the initial population. All 50 Enterococcus isolates were resistant against triple sulfa and trimethoprim/sulfamethoxazole and none were resistant against vancomycin. Most of the isolates from raw sewage were resistant to more antibiotics than the isolates from sand column effluent. One Enterococcus isolate from raw sewage (no. 61) and one Enterococcus isolate from sand column effluent (no. 95) had ten antibiotic resistances each. Isolate no. 95 maintained its resistances in the absence of antibiotics during the whole study. It was compared with isolate no. 70, which was one of the isolates, being resistant only against the two sulfonamides. Phenotypically and biochemically, the two organisms were strains of Enterococcus faecalis. Sequence analysis of partical 16S rDNA allowed alignment of isolate no. 95 as a strain of Enterococcus faecium and of isolate no. 70 as a strain of E. faecalis. E. faecium strain no. 95 carried at least six different plasmids, whereas for E. faecalis strain no. 70, no discrete plasmid band was seen on the gels.     

Antibiotic-resistance profile in environmental bacteria isolated from penicillin production wastewater treatment plant and the receiving river

The antibiotic‐resistance characteristics of bacterial strains in antibiotic production wastewater treatment plants (WWTP) that contain high concentrations of antibiotics are unknown, as are the environmental effects of the discharge of wastewater from such facilities. In this study, 417 strains were individually isolated from the effluent of a WWTP that treated penicillin G production wastewater, as well as from downstream and upstream areas of the receiving river. The minimum inhibition concentrations (MICs) of 18 antibiotics representing seven classes were then determined for each of these strains. Relatively high similarity in the bacterial composition existed between the wastewater and downstream river samples when compared with the upstream sample. High resistance ratios and MIC values were observed for almost all antibiotics in wastewater isolates, followed by strains from downstream river, of which the resistance ratios and levels were still significantly higher than those of upstream strains. The resistance ratios and levels also significantly differed among strains belonged to different species in the penicillin production wastewater effluent and downstream river. In both samples, the resistances to β‐lactam antibiotics were more frequent, with much higher levels, than the other class antibiotics. Then five clinically important resistant genes mainly coding for extended‐spectrum β‐lactamases (ESBLs) were determined for all strains, only bla_TEM‐1 which did not belong to ESBL was detected in 17.3% and 11.0% of strains isolated from wastewater and downstream river respectively. Class I integrons were detected in 14% of wastewater isolates and 9.1% of downstream isolates, and primarily contained gene cassettes conferring resistance to aminoglycoside antibiotics. The unexpectedly high levels of multiple antibiotic resistance in strains from wastewater and downstream river were speculated to be mainly due to multidrug efflux systems.     

Paper Mill Effluent Decolorization by Fifty Streptomyces Strains

Fifty actinomycete strains isolated from lignocellulosic substrates were examined for the ability to remove the color from a paper mill effluent obtained after semichemical alkaline pulping of wheat straw. Streptomyces sp. strains UAH 15, UAH 23, UAH 30, and UAH 51 were selected for their ability to decolorize the effluent in a liquid medium containing 1% (wt/vol) glycerol, 0.2% (wt/vol) ammonium sulfate, and 80% (vol/vol) effluent. The highest levels of decolorization achieved after the strains grew were 60 to 65%. Strains UAH 30 and UAH 51 were selected for further study because of their different patterns of effluent decolorization during growth. Fractionation of the decolorized effluent by gel permeation chromatography demonstrated that there were reductions in the levels of absorbance of the high- and medium-molecular-weight compounds. These fractions were mainly responsible for the color of the effluent, while the last fractions, the low-molecular-weight compounds, could have been responsible for the residual color of the decolorized effluent. Thin-layer chromatography revealed significant differences among the patterns of bands corresponding to the acidified supernatants obtained after precipitation of alkali-lignin from the effluent samples decolorized by different Streptomyces strains.     

Molecular characterization of Acidithiobacillus ferrooxidans and A. thiooxidans strains isolated from mine wastes in Brazil

Nineteen strains of Acidithiobacillus ferrooxidans and Acidithiobacillus thiooxidans, including 12 strains isolated from coal, copper, gold and uranium mines in Brazil, strains isolated from similar sources in other countries and the type strains of the two species were characterized together with the type strain of A. caldus by using a combination of molecular systematic methods, namely ribotyping, BOX- and ERIC-PCR and DNA-DNA hybridization assays. Data derived from the molecular fingerprinting analyses showed that the tested strains encompassed a high degree of genetic variability. Two of the Brazilian A. ferrooxidans organisms (strains SSP and PCE) isolated from acid coal mine waste and uranium mine effluent, respectively, and A. thiooxidans strain DAMS, isolated from uranium mine effluent, were the most genetically divergent organisms. The DNA-DNA hybridization data did not support the allocation of Acidithiobacillus strain SSP to the A. ferrooxidans genomic species, as it shared only just over 40% DNA relatedness with the type strain of the species. Acidithiobacillus strain SSP was not clearly related to A. ferrooxidans in the 16S rDNA tree.     

One-year survey of enteroviruses, adenoviruses, and reoviruses isolated from effluent at an activated-sludge purification plant.

Samples of raw sewage, primary effluent, and secondary effluent from a large activated-sludge purification plant near Melbourne (Victoria, Australia) were collected every second week for 1 year. Viruses were detected in all secondary effluent samples and in six of seven samples obtained after final chlorination. Adenoviruses (85% reduction) and reoviruses (28% reduction) were removed less efficiently by this treatment process than were enteroviruses (93% reduction). In addition, 57 of 171 samples of effluent tested were positive for either adenoviruses or reoviruses, or both, when enteroviruses were not isolated. This clearly shows that the use of enteroviruses as sole indicators of viruses in water may miss up to one-third of instances of viral contamination. Enteroviruses and adenoviruses were isolated most frequently in HeLa-R cell cultures, whereas reoviruses were most often isolated in primary monkey kidney cells.     

Growth of Escherichia coli in a pulp and cardboard mill

The coliform flora of a pulp and cardboard mill that uses birch as the raw material and ammonium sulphate as the process chemical was studied. Escherichia coli was observed to multiply in the mill. It persisted as the dominant thermotolerant coliform in the effluent. Klebsiellae were encountered among total coliforms only. The E. coli strains isolated had the biochemical characteristics and maximum growth temperatures typical to the species. However, serotyping and hemolysin test differentiated these strains from pathogenic and fecal E. coli.     

Exploring the application of biostimulation strategy for bacteria in the bioremediation of industrial effluent

The purpose of this study was to isolate and characterise toxic element-resistant bacteria from acid mine drainage water and to apply them in the bioremediation of industrial effluent, as well as to identify optimal effluent:nutrient concentration for onsite biostimulation strategy. Wastewater samples were collected from acid mine drainage and industry. Inductively coupled plasma optical emission spectroscopy (ICP-OES) was employed for elemental composition analysis. Isolated bacterial strains were characterised using molecular methods. Bioremediation assays were employed to determine the extent of bacterial tolerance and removal of toxic elements using a biostimulation strategy employing minimal salt medium (MSM) at varied concentrations and positive and negative controls of only MSM and industrial effluent, respectively. Two bacterial strains demonstrated resistance to toxic elements, Bacillus sp. MGI101 and Lysinibacillus sp. MGI102 both isolated from the AMD sites. However, no observable growth of toxic metal-resistant bacteria was obtained from the industrial effluents. Bacterial strains MGI101 and MGI102 demonstrated high resistance to target toxic elements during the screening and tolerance assays. Remarkably, Bacillus sp. MGI101 demonstrated greater ability to remove toxic elements including arsenic, chromium, zinc, copper and aluminium in undiluted solutions of the industrial effluent, with its highest removal capacity observed at > 60% for arsenic and aluminium. Both Bacillus sp.MGI101 and Lysinibacillus sp. MGI102 demonstrated varied abilities for the removal of toxic elements from dilution concentration of effluent mixed with MSM. However, the optimal dilution ratio observed in this experiment was 5:15 (effluent:MSM). Overall results demonstrated that isolated bacterial strains have the potential to be employed in bioremediation programmes of acid mine drainages and multi-element contaminated water.

     

Fungal treatment of a delignification effluent from a nitrocellulose industry

Twelve strains of filamentous fungi, most of them belonging to the Deuteromycetes class, were isolated from activated sludge adapted to the delignification effluent from a nitrocellulose industry and screened to be used in the treatment of the effluent. The screening experiment was carried out using the effluent without co-substrate, treated for 120 h and pH 5. Aspergillus 2BNL1, Aspergillus 1AAL1 and Lentinus edodes UEC 2019 showed the highest effluent color reduction rates between 83% and 95%. The white-rot fungus L. edodes UEC 2019 was used as the control for the decolorization. In addition to color reduction, total phenol was also reduced in 56% and 79% by Aspergillus 2BNL1 and L. edodes UEC 2019, respectively. A kinetic experiment showed that Aspergillus 2BNL1 and Aspergillus 1AAL1 reduced the effluent color in the range of 81-95% at the first 24 h while L. edodes required 72 h to achieve a similar result. UV/Visible spectra revealed that all fungi treatments were able to decrease the chromophore compounds present in the effluent, except Aspergillus 1AAL1 that increased the UV absorptions. The molar weight distribution analysis showed that the three fungi were able to change the pattern of the effluent chromatogram, probably by degradation of the high molecular weight compounds.     

Toxigenic Clostridium difficile PCR ribotypes from wastewater treatment plants in southern Switzerland

The occurrence of Clostridium difficile in nine wastewater treatment plants in the Ticino Canton (southern Switzerland) was investigated. The samples were collected from raw sewage influents and from treated effluents. Forty-seven out of 55 characterized C. difficile strains belonged to 13 different reference PCR ribotypes (009, 010, 014, 015, 039, 052, 053, 066, 070, 078, 101, 106, and 117), whereas 8 strains did not match any of those available in our libraries. The most frequently isolated ribotype (40%) was 078, isolated from six wastewater treatment plants, whereas ribotype 066, a toxigenic emerging ribotype isolated from patients admitted to hospitals in Europe and Switzerland, was isolated from the outgoing effluent of one plant. The majority of the isolates (85%) were toxigenic. Forty-nine percent of them produced toxin A, toxin B, and the binary toxin (toxigenic profile A(+) B(+) CDT(+)), whereas 51% showed the profile A(+) B(+) CDT(-). Interestingly, eight ribotypes (010, 014, 015, 039, 066, 078, 101, and 106) were among the riboprofiles isolated from symptomatic patients admitted to the hospitals of the Ticino Canton in 2010. Despite the limitation of sampling, this study highlights that toxigenic ribotypes of C. difficile involved in human infections may occur in both incoming and outgoing biological wastewater treatment plants. Such a finding raises concern about the possible contamination of water bodies that receive wastewater treatment plant effluents and about the safe reuse of treated wastewater.     

Drug resistant coagulase-positive staphylococci strains in food

From the 1572 food samples, examined in Microbiology Department of Frozen Food Industry Research Laboratory in Lód?, 79 (5.0%) coagulase-positive staphylococci strains were isolated. All the strains were sensitive to vancomycin, gentamycin and chloramphenicol. Only individual staphylococci strains were resistant to erythromycin (1.3%), lincomycin (2.5%) and ciprofloxacin (2.5%). 20.3% strains, isolated mainly from raw meat, were resistant to doxycycline and 6.3% to oxacillin. 38.0% of coagulase-positive staphylococci strains had positive results of cefinase test. One strain isolated from minced meat was resistant to methicillin and at the same time it was producing beta-lactamases.     

The microbiology of a pharmaceutical effluent and its public health implications

The effluent of a pharmaceutical company was examined microbiologically. Its bacterial count was 2.15 × 10⁵ c.f.u./ml and there was evidence of faecal contamination with MPN of > 1800. The organisms encountered included Staphylococcus aureus, Escherichia coli, Proteus vulgaris, Serratia marcescens and Pseudomonas aeruginosa. The resistances of the 25 bacterial strains isolated from the effluent to the commonly used antibiotics were studied. About 80% of the isolates were resistant to Amoxycillin, 76% to Nitrofurantoin, 64% to Cotrimoxazole and Augmentin, 60% were resistant to Nalidixic acid, 52% were resistant to Tetracycline and Ofloxacin, while resistance of 12% was obtained for Gentamicin. Among the eight antibiotics tested, seven patterns of drug resistance were obtained and all of them were multiple-drug resistance with the number of antibiotics ranging from 2–8. All the strains of E. coli and S. aureus had high MIC values for Cloxacillin and Amoxycillin. In all, 13 strains of the bacterial isolates had evidence for the production of -lactamases. The potential of the effluent in spreading drug resistance and the public health implications are discussed.     

Removal of bacterial indices of pollution during sewage treatment via rotating biological contactor

Summary Two rotating biological contactors (RBC), one bench scale and the other pilot-plant scale, were assessed by using bacterial indices of pollution. The removal efficiency for the pilot RBC was much better than that obtained under laboratory conditions. However, the residual bacterial densities were high enough to represent a potential health hazard. In addition, five different serotypes of enteropathogenic Escherichia coli were isolated from influent and three from effluent samples and 45% of the tested strains were multiple antibiotic resistant.     

The potential of microalgal biomass production for biotechnological purposes using wastewater resources

The utilization of microalgae for wastewater treatment represents an attractive opportunity for wastewater valorization through the use of the produced biomass. Five strains of microalgae were isolated from municipal wastewater and grown in autoclaved and non-autoclaved effluent at 30 °C and 150 μmol photons m^?2 s^?1 to study biomass production, nutrient removal, and the biochemical composition of the biomass. All strains reached high biomass productivity (35.6 to 54.2 mg dry weight L^?1 day^?1) within 4 days of batch culturing. In this period, ammonium-N and phosphate were reduced by more than 60 and 90 %, respectively. The high growth rate (0.57 to 1.06 day^?1) ensured a rapid removal of nutrients and thereby a short retention time. By the fourth day of cultivation, the algal biomass contained 32 % protein, but only 11 % lipids and 18 % carbohydrates. It was found that the biomass was a suitable raw material for biogas production by anaerobic digestion. Biodigestion of obtained biomass was simulated by employing the Aspen HYSYS modeling software, resulting in methane yields comparable to those found in the literature. The elemental analysis of the algal biomass showed very low concentrations of pollutants, demonstrating the potential of use of the digestate from biodigestion as a bio-fertilizer.     

Use of the molecular typing methods to evaluate the control ofListeria monocytogenes contamination in a raw milk and dairy products

Nineteen serogroup 1/2a Listeria monocytogenes strains isolated from raw milk, dairy products and salt water in one dairy were analyzed. Pulsed field gel electrophoresis (PFGE) and ribotyping were used to determine whether these strains isolated over a 8-month period are epidemiologically related. The samples of raw milk were contaminated by different L. monocytogenes clones. The clones isolated from dairy products (with the exception of one sample) and salt water were identical. Comparative genetic analysis of the clones isolated from raw milk, salt water and dairy products revealed the source of contamination and identified the L. monocytogenes strain involved in this process.