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The main objective of this study is the evaluation of RPPW anaerobic digestion feasibility at laboratory scale under Mesophilic condition. The experiment is conducted using a two-stage biofilm digester of 5 L capacity with mobile support material. Anaerobic treatment of wastewater from recycled pulp and paper industry in Morocco was tested using a laboratory-scale anaerobic biofilm digester that operated under mesophilic conditions over a 70-day. Chemical oxygen demand (COD) efficiency, volatile and total solid (VS, TS) elimination of the substrate during the process were: 78%, 52% and 48% respectively. The system was stable throughout its operating cycle with an optimum pH (7.24), alkalinity (1750 mg CaCO3/L) and a volatile fatty acid value (760 mg/L). The experimental daily biogas production measured reaches a value of 5 L/day with a composition of 71% methane, 27.6% carbon dioxide, 0.2 oxygen and 7713 ppm of the H2S. The study results show that the anaerobic biofilm reactor is a suitable technique for recycled pulp and paper wastewater (RPPW) treatment. The reactor shows high performances in terms of process stability, removal efficiency (> 70%) and biogas production. Anaerobic digestion is an efficient waste treatment technology that uses natural anaerobic decomposition to reduce the volume of waste while producing biogas. However, research is needed to strengthen microbial metabolism, biochemistry and the functioning of the rector to improve biogas production. The RPPW AD experiment with biofilm digester technology was stable throughout the operation period. The digester knows an overloaded in the last phase of the experiment which leads to an inhibition of biogas production.

Nowadays, thanks to nanotechnological progress, which itself guides us more and more closely toward not only the efficient design of innovative nanomaterials or nanostructures, but to the improvement of their functionality, we benefit from an important asset in the battle against pathogenic illnesses. Herein, we report a versatile biocompatible plasmonic nanoplatform based on a Whatman paper incorporating positively-charged gold nanospherical particles via the immersion approach. The morphological characterization of the as-engineered-plasmonic paper was examined by SEM (scanning electron microscopy) and HRTEM (high-resolution transmission electron microscopy) investigations, while its surface chemical modification with a synthetic polypeptide, specifically RRWHRWWRR-NH2 (P2), was proved by monitoring the plasmonic response of loaded gold nanospheres and the emission signal of P2 via fluorescence spectroscopy. The as-functionalized plasmonic paper is non-cytotoxic towards BJ fibroblast human cells at bactericidal concentrations. Finally, the antimicrobial activity of the P2-functionalized plasmonic paper on both planktonic bacteria and biofilms was tested against two reference strains: Gram-positive Bacteria, i.e., Staphylococcus aureus and the Gram-negative Bacteria, i.e., Escherichia coli, determining microbial inhibition of up to 100% for planktonic bacteria. In line with the above presented nanoplatform&rsquo, s proper design, followed by their functionalization with active antimicrobial peptides, new roads can be open for determining antibiotic-free treatments against different relevant pathogens.

Secondary effluent from paper mill was characterized by poor biodegradability and containing recalcitrant compounds. In this study, four bioreactors, including a sequencing batch biofilm reactor (SBBR), a stirred-tank reactor (STR) and two submerged aeration reactors (SAR) were used to treat secondary effluent from a recycled paper mill respectively. The results indicated that chemical oxygen demand (COD) was increased by SAR2 treatment and COD removal efficiency for SBBR, SAR1 and STR was 39.7%, 15.7% and 30.9% respectively. It is suggested that recalcitrant compounds were removed by SBBR, SAR1 and STR respectively. Total nitrogen (TN) and total phosphorus (TP) of wastewater were increased by treatments of each bioreactor, which suggested that endogenous respiration of biomass occurred during the treatment. Microbial analysis of sludge from different bioreactors suggested that the removal of recalcitrant compounds in SBBR and STR might be related to the presence of unique microorganisms in each reactor.

The goal of this work is to pursue analytical approaches that elucidate electron and proton diffusion inside the Shewanella oneidensis biofilm and bulk liquid, which will inevitably promote the translation of Microbial Fuel Cell (MFC) technology for renewable, "green energy" solutions that are in demand to sustain the world's ever-increasing energy demands and to mitigate the depletion of current resources. This study provides a novel strategy for monitoring electron/proton fluxes in 3-D multi-laminate structures of paper as a scaffold to support S. oneidensis biofilms and bulk media liquid. Multiple layers of paper containing bacterial cells and/or media are stacked to form a layered 3-D model of the overall biofilm/bulk liquid construct. Mass transport of electrons and protons into this 3-D system can be quantified along with the exploration of microbial energy production. Assembly of a 3D paper stack can be modular and allows us to control the thickness of the overall biofilm/bulk liquid construct with the different diffusion distances of the electrons/protons through the stack. By measuring the current generated from the 3-D stack, the electron and proton diffusivity through biofilms were quantitatively investigated. We found that (i) the diffusion length of the electrons/protons in the S. oneidensis biofilm/bulk liquid is a determinant factor for the MFC performance, (ii) the electron transfer through the endogenous mediators of S. oneidensis can be a more critical factor to limit the current/power generation of the MFCs than the proton transfer in the MFC system and (iii) the thicker biofilm allows higher and longer current generation but requires more time to reach a peak current value and increases the total energy loss of the MFC system.

Biofilm formation is a major cause of reduced paper quality and increased down time during paper manufacturing. This study uses Illumina next‐generation sequencing to identify the microbial populations causing quality issues due to their presence in biofilms and slimes. The paper defects investigated contained traces of the films and/or slime of mainly two genera, Tepidimonas and Chryseobacterium. The Tepidimonas spp. found contributed on average 68% to the total bacterial population. Both genera have been described previously to be associated with biofilms in paper mills. There was indication that Tepidimonas spp. were present as compact biofilm in the head box of one paper machine and was filtered out by the paper web during production. On the other hand Tepidimonas spp. were also present to a large extent in the press and white waters of two nonproblematic paper machines. Therefore, the mere presence of a known biofilm producer alone is not sufficient to cause slimes and therefore paper defects and other critical factors are additionally at play. For instance, we identified Acidovorax sp., which is an early colonizer of paper machines, exhibiting the ability to form extracellular DNA matrices for attachment and biofilm formation.

Sugar-rich diets and poor dental hygiene promote the formation of a biofilm (plaque) that strongly adheres to the dental enamel surface and fosters the evolution of aciduric bacteria. The acid contributes to demineralization of the exterior tooth enamel, which accelerates after the pH drops below a critical value (∼5.5) for extended time periods resulting in the need for restorative procedures. Preventative techniques to alert the dentist and caries-susceptible patients regarding vulnerability to dental decay require a clinical measure of plaque activity. Therefore, there is a need to evaluate the acid production capability of plaque deposits in the pits and fissures of occlusal and interproximal regions. A ratiometric fluorescence pH-sensing device has been developed using an FDA-approved dye and LED excitation. Fluorescein spectral profiles were collected using a spectrometer and analyzed with a spectral unmixing algorithm for calibration over the pH range of 4.5 to 7. An in vivo pilot study on human subjects was performed using a sucrose rinse to accelerate bacterial metabolism and to measure the time-dependent drop in pH. The optical system is relatively immune to confounding factors such as photobleaching, dye concentration, and variation in excitation intensity associated with earlier dye-based pH measurement techniques.

The present clinical study aimed to investigate if there are differences in microbiological outcomes dependent on the subgingival biofilm collection method. Subgingival biofilm samples were collected from the four deepest pockets (>5 mm) of 17 patients with aggressive periodontitis (AgP) and 33 patients with chronic periodontitis (CP), first by paper point and thereafter by curette. Samples obtained with the same method were pooled together from each patient and forwarded for molecular microbiological analysis by a commercially available assay (IAI Pado Test 4.5) that estimates total bacterial load and levels of Porphyromonas gingivalis, Tannerella forsythia, Treponema denticola and Aggregatibacter actinomycetemcomitans. Data analysis included frequency of detection, quantification and correlation of detection levels between the two sampling methods. P. gingivalis, T. forsythia and T. denticola were detected in >90% of the samples, and their detection levels exhibited a strong correlation between sampling methods. The detection consistency of A. actinomycetemcomitans was 56% between the two sampling methods. A. actinomycetemcomitans was more readily detected by paper point compared with curette collection with a stronger correlation between the two methods in AgP. Subgingival biofilm sampling by curette or paper point does not yield differences in the detection of the three 'red complex' species. However, A. actinomycetemcomitans was more consistently detected by means of paper point collection, which can be crucial in the decision to administer antibiotics as an adjunctive periodontal treatment.

A pilot scale granular activated carbon-sequencing batch biofilm reactor with a capacity of 2.2 m3 was operated for over three months to evaluate its performance treating real recycled paper industry wastewater under different operational conditions. In this study, dissolved air floatation (DAF) and clarifier effluents were used as influent sources of the pilot plant. During the course of the study, the reactor was able to biodegrade the contaminants in the incoming recycled paper mill wastewater in terms of chemical oxygen demand (COD), adsorbable organic halides (AOX; specifically 2,4-dichlorophenol (2,4-DCP)) and ammoniacal nitrogen (NH3-N) removal efficiencies at varying hydraulic retention times (HRTs) of 1-3 days, aeration rates (ARs) of 2.1-3.4 m3/min and influent feed concentration of 40-950 mg COD/l. Percentages of COD, 2,4-DCP and NH3-N removals increased with increasing HRT, resulting in more than 90% COD, 2,4-DCP and NH3-N removals at HRT values above two days. Degradation of COD, 2,4-DCP and NH3-N were seriously affected by variation of ARs, which resulted in significant decrease of COD, 2,4-DCP and NH3-N removals by decreasing ARs from 3.4 m3/min to 2.1 m3/min, varying in the ranges of 24-80%, 6-96% and 5-42%, respectively. In comparison to the clarifier effluent, the treatment performance of DAF effluent, containing high COD concentration, resulted in a higher COD removal of 82%. The use of diluted DAF effluent did not improve significantly the COD removal. Higher NH3-N removal efficiency of almost 100% was observed during operation after maintenance shutdown compared to normal operation, even at the same HRT of one day due to the higher dissolved oxygen concentrations (1-7 mg/l), while no significant difference in COD removal efficiency was observed.

Biofilms cause runnability problems in paper machines and are therefore controlled with biocides. Peracetic acid is usually effective in preventing bulky biofilms. This study investigated the microbiological status of a paper machine where low concentrations (≤ 15 ppm active ingredient) of peracetic acid had been used for several years. The paper machine contained a low amount of biofilms. Biofilm-forming bacteria from this environment were isolated and characterized by 16S rRNA gene sequencing, whole-cell fatty acid analysis, biochemical tests, and DNA fingerprinting. Seventy-five percent of the isolates were identified as members of the subclades Sphingomonas trueperi and S. aquatilis, and the others as species of the genera Burkholderia (B. cepacia complex), Methylobacterium, and Rhizobium. Although the isolation media were suitable for the common paper machine biofoulers Deinococcus, Meiothermus, and Pseudoxanthomonas, none of these were found, indicating that peracetic acid had prevented their growth. Spontaneous, irreversible loss of the ability to form biofilm was observed during subculturing of certain isolates of the subclade S. trueperi. The Sphingomonas isolates formed monoculture biofilms that tolerated peracetic acid at concentrations (10 ppm active ingredient) used for antifouling in paper machines. High pH and low conductivity of the process waters favored the peracetic acid tolerance of Sphingomonas sp. biofilms. This appears to be the first report on sphingomonads as biofilm formers in warm water using industries.

Biofilm samples collected from inside and outside the press and former sections of paper machines in a Northwestern Ontario paper mill for a period of 2 years were characterized microbiologically and electrochemically. Bacterial community profiling was done using polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) and selected bacterial isolates were identified using 16S rDNA analysis. The bacterial community showed the presence of Proteobacteria, Firmicutes, and Actinobacteria. Sphingomonas sp. was found to be the most common bacterial species, which showed the highest production of extracellular polymeric substances. Bacteria isolated from biofilms showed better adhesion properties than those from water samples. Cyclic voltammetry and electrochemical impedance spectroscopy studies showed that bacteria isolated from biofilms and feed water collected from inside the machine were more easily oxidized than those from outside, suggesting the need for a more rigorous biofilm abatement strategy for inside paper machines.

Methanol-oxidizing bacteria may play an important role in the development and use of biological treatment systems for the removal of methanol from industrial effluents. Optimization of methanol degradation potential in such systems is contingent on availability of nutrients, such as nitrogen, in the most favorable form and concentration. To that end, this study examined the variation in growth, methanol degradation, and bacterial diversity of two mixed methylotrophic cultures that were provided nitrogen either as ammonium or nitrate and in three different concentrations. Methanol-degrading cultures were enriched from biofilms sampled at a pulp and paper mill and grown in liquid batch culture with methanol as the only carbon source and either ammonium or nitrate as the only added nitrogen source. Results indicate that growth and methanol removal of the mixed cultures increase directly with increased nitrogen, added in either form. However, methanol removal and bacterial diversity, as observed by polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) methods, were higher when using nitrate as the nitrogen source for enrichment and growth, rather than ammonium. Based on results described here, nitrate may potentially be a better nitrogen source when enriching or working with mixed methylotrophic cultures, and possibly more effective when used as a nutrient addition to biofilters.

Formation of biofilms causes severe problems in paper machines, and hence financial costs. It would be preferable to prevent attachment of the primary-colonizing bacteria than to control the growth of secondary communities, which are sheltered by exopolysaccharide slime layers. We have therefore investigated the early succession of paper-machine biofilms by incubating stainless-steel test coupons in the process water-flow lines in two paper machines operating in slightly alkaline conditions in temperatures (45 and 49 degrees C) supporting thermophilic microbes. Microbial succession was profiled using length heterogeneity analysis of PCR-amplified 16S rRNA genes (LH-PCR) and linking the sequence data of the created 16S rRNA gene libraries to the dominant LH-PCR peaks. Although the bacterial fingerprints obtained from the attached surface communities varied slightly in different samples, the biomarker signals of the dominating primary-colonizing bacterial groups remained high over time in each paper machine. Most of the 16S rRNA gene copies in the early biofilms were assigned to the genera Rhodobacter, Tepidimonas, and Cloacibacterium. The dominance of these sequence types decreased in the developing biofilms. Finally, as phylogenetically identical primary-colonizers were detected in the two different paper mills, the machines evidently had similar environmental conditions for bacterial growth and potentially a common source of contamination.

Deinococcus geothermalis has frequently been isolated from pink colored deposits of paper industry processes. Laboratory studies have shown that D. geothermalis is capable of forming on nonliving surfaces patchy biofilms that are resistant to adverse agents such as extreme pH, desiccation, solubilising detergents and biocides. This study was done to quantitatively assess the role of D. geothermalis as a biofouler in paper industry. Colored deposits were collected from 24 European and North American paper and board machines and the densities of the bacterial 16S rRNA genes and those of the red slime producers D. geothermalis and Meiothermus spp. were measured by QPCR (quantitative real time PCR). D. geothermalis was found at nine machines, usually from splash area deposits, but its contribution was minor, 0.001-1%, to the total bacterial burden of 8.3 to log 10.5 log units per gram wet-weight of the deposits. When D. geothermalis was found in a measurable quantity, Meiothermus spp. also was found, often in bulk quantity (7-100% of the total bacteria). The data are in line with the properties of D. geothermalis known from laboratory biofilm studies, indicating this species is a pioneer coloniser of machine surfaces and may help other bacteria to adhere and grown into biofilms, rather than competing with them.

To provide the first step towards a microbial ecology-based understanding of irreversible membrane biofouling, four laboratory-scale membrane bioreactors (MBRs) were operated to investigate the identity of bacterial populations highly correlated with irreversible membrane biofouling. The conventional MBR was divided into two separate experimental units. Unit one consisted of four suspended-growth, activated sludge, sequencing batch bioreactors treating a synthetic paper mill wastewater. Unit two consisted of a microfiltration membrane cell. Amplified ribosomal deoxyribonucleic acid restriction analysis (ARDRA) was used to compare the predominant bacterial populations in samples of mixed liquor and irreversibly bound to the membrane surface. The results of ARDRA showed a significant difference between the planktonic and sessile bacterial communities suggesting that irreversible biofouling of microfiltration membranes may be more highly correlated to specific bacterial populations rather than the total, bulk concentration of biomass. A custom-built mini-flow cell and light microscopy were used to visualise the early formation of biofilms by two pure cultures (Escherichia coli and Acinetobacter calcoaceticus) on membrane surfaces. The results confirmed that A. calcoaceticus was able to enhance the initiation of biofilm formation on microfiltration membranes.

The fate of nutrients in the modern effluent treatment plant depends on several factors, for example type of treatment plant, availability of nutrients in the specific effluent, dosing of nutrients and sludge age/production. New technologies with the aim to increase the efficiency and stability of the conventional activated sludge process have strongly affected the possibilities to control discharge of nutrients in pulp and paper effluents. A paradox is that a reduction of organic material may often lead to an increase of nutrient discharges. It is of the utmost importance that the operators have good knowledge of the factors affecting nutrient uptake and release in order to minimise nutrient discharge and obtain optimal plant performance. Dosing of nitrogen and phosphorus is one key factor in the sensitive balance in most pulp and paper effluent treatment plants. Correct dosing is crucial as high or low doses might lead not only to increased discharge of nutrients but also to severe operational problems with poor sludge quality, which in turn affects the plant performance for longer periods.

This study reports on a multivariate analysis of the moving bed biofilm reactor (MBBR) wastewater treatment system at a Canadian pulp mill. The modelling approach involved a data overview by principal component analysis (PCA) followed by partial least squares (PLS) modelling with the objective of explaining and predicting changes in the BOD output of the reactor. Over two years of data with 87 process measurements were used to build the models. Variables were collected from the MBBR control scheme as well as upstream in the bleach plant and in digestion. To account for process dynamics, a variable lagging approach was used for variables with significant temporal correlations. It was found that wood type pulped at the mill was a significant variable governing reactor performance. Other important variables included flow parameters, faults in the temperature or pH control of the reactor, and some potential indirect indicators of biomass activity (residual nitrogen and pH out). The most predictive model was found to have an RMSEP value of 606 kgBOD/d, representing a 14.5% average error. This was a good fit, given the measurement error of the BOD test. Overall, the statistical approach was effective in describing and predicting MBBR treatment performance.

A stainless steel paper-embedded biofilm reactor (PEBR) was developed for Candida spp. growth, permitting confluent distribution of nutrients by capillary diffusion through ordinary laboratory filter paper. Antibiogram disks were distributed along the filter paper rim, and the PEBR received 0.1 or 0.01 % crystal violet (CV) at 200 μL min(-1) and at 37 °C, for 48 h. CV was recovered from the disks and measured at 540 nm. Candida albicans SC5314 cells were applied onto antibiogram disks. The bioreactor was assembled, and YEPD broth was admitted (200 μL min(-1)) at 37 °C, for 72 h. Biofilm growth was estimated via the MTT reduction test. Controls were disks that received the same treatments, except for the fungus. The PEBR was considered high-throughput table, low-cost, and feasible to grow C. albicans biofilms.

This work was partially supported by the Portuguese Science Foundation (FCT-MEC) through the projects EXCL/CTM-NAN/0201/2012, Pest-C/CTM/LA0025/2013-14, Pest-C/EQB/LA0006/2013-14, PTDC/BBB-BEP/0753/2012 and the ERC Advanced Grant (INVISIBLE, contract number 228144). Geobacter sulfurreducens cells were kindly provided by Prof. Derek Lovley from University of Massachusetts (USA). We would like to thank to Dr. Daniela Nunes for the SEM images. Electrochemically active bacteria (EAB) have the capability to transfer electrons to cell exterior, a feature that is currently explored for important applications in bioremediation and biotechnology fields. However, the number of isolated and characterized EAB species is still very limited regarding their abundance in nature. Colorimetric detection has emerged recently as an attractive mean for fast identification and characterization of analytes based on the use of electrochromic materials. In this work, WO 3 nanoparticles were synthesized by microwave assisted hydrothermal synthesis and used to impregnate non-treated regular office paper substrates. This allowed the production of a paper-based colorimetric sensor able to detect EAB in a simple, rapid, reliable, inexpensive and eco-friendly method. The developed platform was then tested with Geobacter sulfurreducens, as a proof of concept. G. sulfurreducens cells were detected at latent phase with an RGB ratio of 1.10 ± 0.04, and a response time of two hours. publishersversion published

Parallel laboratory investigations were conducted to examine aspects of two distinct but related bioprocess strategies for low sludge production in the treatment of the same TCF kraft pulp mill effluent. The purpose of this article has been to compare the performance results from these two bench-scale trials with respect to nutrient demands, nutrient discharge, COD removal, and waste sludge characteristics. The LSP (Low Sludge Production) process can be used to significantly reduce sludge yield with excellent sludge characteristics. These sludge characteristics seemed to be related to elevated protozoan grazing pressures. The BAS (Biofilm-Activated Sludge) process achieves similar reduced sludge yields and sludge characteristics while at the same time significantly reducing the nutrient demands and discharge levels. For both LSP and BAS process optimization, the selector nutrient loading is critical to the overall process performance. Selector nutrient requirements are distinct from the overall process nutrient requirements.

Biofilms cause several problems in papermaking. This report describes a microbiological survey of colored biofilms in six paper and board machines, including two case studies of outbreaks of colored slimes in which the causative bacteria were found. A total of 95 pink-, red-, orange- or yellow-pigmented strains were isolated. Nearly all (99%) of the strains grew at 52 degrees C, 72% grew at 56 degrees C, but only 30% grew at 28 degrees C, indicating that most of the strains were moderately thermophilic. Biofilm formation potential and biocide susceptibility of the strains were analyzed with a microtiter plate assay. In the presence of 5 ppm of methylene bisthiocyanate or 2,2-dibromo-3-nitrilopropionamide in paper-machine water, 55 strains formed biofims. Moreover, 39 strains increased biofilm production by 5-753% in the presence of biocide, suggesting that biocide concentrations inhibitory to planktonic but not to surface-attached cells may actually promote biofouling. The cells may have inactivated a portion of the biocides, as the cell density in this assay was high, corresponding to the highest cell densities occurring in the circulating waters. Four groups of colored bacteria that were isolated from several mills were identified. Pink-pigmented Deinococcus geothermalis and red-pigmented Meiothermus silvanus occurred as common primary biofilm-formers in paper machines. This report is the first description of the involvement of Meiothermus species in red-slime formation in the paper industry. The third group of bacteria (putative new species related to Roseomonas) contained strains that were not biofilm formers, but which were commonly found in slimes of neutral or alkaline machines. The fourth group contained red-pigmented biofilm-forming strains representing a novel genus of alpha- Proteobacteria related to Rhodobacter. Many colored paper-machine bacteria are species previously known from microbial mats of hot springs. Some characteristics of the bacterial groups are described here in order to facilitate their recognition in future cases of colored-slime outbreaks in the paper industry.

Experiments were designed (1) to investigate the bioadhesion, biofilm formation, foxing, and micropitting in documentary collections, (2) to assess the risk of biodeterioration, (3) to investigate the environmental microbial concentration, and (4) to study the influence of environmental factors in biodeterioration of documentary heritage in three archives. The importance of this work in the field of biodeterioration of documentary heritage was verified by bioadhesion and biofilm formation by microorganisms isolated from the collections under study. Bacillus sp. and Scopulariopsis sp. isolated from paper books showed considerable evidence of attacking the paper structure and of pigment production, constituting a hazard to the loss of documentary heritage. Fil: Lavin, Paola. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico la Plata. Instituto de Investigaciones Fisicoquímicas Teóricas y Aplicadas; Argentina. Universidad Nacional de La Plata; Argentina Fil: Gómez de Saravia, Sandra Gabriela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico la Plata. Instituto de Investigaciones Fisicoquímicas Teóricas y Aplicadas; Argentina. Universidad Nacional de La Plata; Argentina Fil: Guiamet, Patricia Sandra. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico la Plata. Instituto de Investigaciones Fisicoquímicas Teóricas y Aplicadas; Argentina. Universidad Nacional de La Plata; Argentina

The microbial content of printing paper machines, running at a temperature of 45-50 degrees C and at pH 4.5-5, was studied. Bacteria were prevalent colonizers of the machine wet end and the raw materials. A total of 390 strains of aerobic bacteria were isolated and 86% of these were identified to genus and species by biochemical, chemotaxonomic and phylogenetic methods. The most common bacteria found at the machine wet end were Bacillus coagulans and other Bacillus species, Burkholderia cepacia, Ralstonia pickettii, and in pink slimes, accumulating in the wire area and press section, species of Deinococcus, aureobacterium and Brevibacterium. Paper-making chemicals also contained species of Aureobacterium, B. cereus, B. licheniformis, B. sphaericus, Bordetella, Hydrogenophaga, Klebsiella pneumoniae, Pantoea agglomerans, Pseudomonas stutzeri, Staphylococcus and sometimes other enteric bacteria, but these did not colonize the process water. Yeasts and moulds were not present in significant numbers. A total of 131 strains were tested for their potential to degrade paper-making raw materials; 91 strains were found to have degradative activity, mainly species of Burkholderia and Ralstonia, Sphingomonas and Bacillus, and enterobacteria produced enzymes which degraded paper-making chemicals. Stainless steel adhering strains occurred in slimes and wire water and were identified as Burkholderia cepacia, B. coagulans and Deinococcus geothermalis. Coloured slimes were formed on the machine by species of Deinococcus, Acinetobacter and Methylobacterium (pink), Aureobacterium, Pantoea and Ralstonia (yellowish) and Microbulbifer-related strains (brown). The impact of the strains and species found in the printing paper machine community on the technical quality of paper, machine operation, and as a potential biohazard (Hazard Group 2 bacteria), is discussed.

Paper mills are open systems, which provide favorable conditions for microbial growth. Microbial contamination can cause substantial economic losses, including the deterioration of raw materials, interference with production processes by breakdowns and lowering product quality, and eventually, problems in wastewater treatment. Damage is caused by acidification, attack on raw materials, the formation of odorous products, discoloration of pigments, and the formation of methane and hydrogen, thereby producing potentially explosive conditions. Population analyses have revealed that a wide variety of microorganisms are involved, but there appear to be no typical strains associated with paper mills. Current trends in process engineering, such as chlorine-free bleaching, processing at neutral pH, closed cycles, and the use of recycled paper also favor microbial growth and biofilm (slime) formation. A fundamental problem associated with slimes is the extensive matrix of extracellular polymeric substances, which is composed of a large variety of highly hydrated polysaccharides, proteins, nucleic acids, and lipids. No 'silver bullet' against biofouling can be expected, and effective countermeasures have to be based on holistic approaches.

Qi Wang,1 Thomas J Webster21Bioengineering Program, 2Department of Chemical Engineering, College of Engineering, Northeastern University, Boston, MA, USAAbstract: Bacterial infections are commonly found on paper towels and other paper products, leading to the potential spread of bacteria and consequent health concerns. The objective of this in vitro study was to introduce antibacterial properties to standard paper towel surfaces by coating them with selenium nanoparticles. Scanning electron microscopy was used to measure the size and distribution of the selenium coatings on the paper towels. Atomic force microscopy was used to measure the surface roughness of paper towels before and after they were coated with selenium nanoparticles. The amount of selenium precipitated on the paper towels was measured by atomic absorption spectroscopy. In vitro bacterial studies with Staphylococcus aureus were conducted to assess the effectiveness of the selenium coating at inhibiting bacterial growth. Results showed that the selenium nanoparticles coated on the paper towel surface were well distributed with semispherical geometries about 50 nm in diameter. Most importantly, the selenium nanoparticle-coated paper towels inhibited S. aureus growth by 90% after 24 and 72 hours compared with the uncoated paper towels. Thus, the study showed that nanoparticle selenium-coated paper towels may lead to an increased eradication of bacteria in a wider range of clinical environments and in the food industry, thus improving human health.Keywords: selenium nanoparticles, paper towel, antibacterial, Staphylococcus aureus

Phosphate is an essential nutrient for heterotrophic bacteria, affecting bacterioplankton in aquatic ecosystems and bacteria in biofilms. However, the influence of phosphate limitation on bacterial competition and biofilm development in multispecies populations has received limited attention in existing studies. To address this issue, we isolated 13 adhesive bacteria from paper machine aggregates. Intergeneric inhibition of Pseudomonas aeruginosa WW5 by Serratia marcescens WW4 was identified under phosphate-limited conditions, but not in Luria–Bertani medium or M9 minimal medium. The viable numbers of the pure S. marcescens WW4 culture decreased over 3 days in the phosphate-limited medium; however, the mortality of S. marcescens WW4 was significantly reduced when it was co-cultured with P. aeruginosa WW5, which appeared to sustain the S. marcescens WW4 biofilm. In contrast, viable P. aeruginosa WW5 cells immediately declined in the phosphate-limited co-culture. To identify the genetic/inhibitory element(s) involved in this process, we inserted a mini-Tn5 mutant of S. marcescens WW4 that lacked inhibitory effect. The results showed that an endonuclease bacteriocin was involved in this intergeneric inhibition by S. marcescens WW4 under phosphate limitation. In conclusion, this study highlights the importance of nutrient limitation in bacterial interactions and provides a strong candidate gene for future functional characterisation.

Polyhydroxyalkanoate (PHA) biosynthesis in paper mill wastewater treated by a Moving Bed Biofilm Reactor (MBBR) was evaluated. A MBBR was operated during 300 d. The increasing effect of the Organic Load Rate (OLR) from 0.13 kg BOD(5)/m(3)·d to 2.99 kg BOD(5)/m(3)·d and the influence of two relationship of BOD(5:) N: P (100: 5: 1 and 100: 1: 0.3) on the PHA biosynthesis were evaluated. With an OLR of 0.13 kg BOD(5)/m(3)·d, the maximum organic matter removal measure as Biochemical Oxygen Demand (BOD(5)) was 98.7% for a BOD(5:) N: P relationship of 100: 5: 1. Meanwhile for BOD(5): N: P relationship of 100: 1: 0.3, the maximum efficiency was 87.2% (OLR: 2.99 kg BOD(5)/m(3)·d). The behaviour of the Chemical Oxygen Demand (COD) and total phenolic compound removal efficiencies were below 65.0% and 41.0%, respectively. PHA biosynthesis was measured as a percentage of cells that accumulate PHA, where the maximum percentage was 85.1% and 78.7% when MBBR was operated under a BOD(5): N: P relationship of 100: 5: 1 and 100: 1: 0.3, respectively. Finally, the PHA yields in this study were estimated to range between 0.11 to 0.72 mg PHA/mg VSS and 0.06 to 0.15 mg PHA/mg COD.

In this study, the potential of a pilot-scale granular activated carbon sequencing batch biofilm reactor (GAC-SBBR) for removing chemical oxygen demand (COD), ammoniacal nitrogen (NH3-N) and 2,4-dichlorophenol (2,4-DCP) from recycled paper wastewater was assessed. For this purpose, the response surface methodology (RSM) was employed, using a central composite face-centred design (CCFD), to optimise three of the most important operating variables, i.e., hydraulic retention time (HRT), aeration rate (AR) and influent feed concentration (IFC), in the pilot-scale GAC-SBBR process for recycled paper wastewater treatment. Quadratic models were developed for the response variables, i.e., COD, NH3-N and 2,4-DCP removal, based on the high value (>0.9) of the coefficient of determination (R(2)) obtained from the analysis of variance (ANOVA). The optimal conditions were established at 750 mg COD/L IFC, 3.2 m(3)/min AR and 1 day HRT, corresponding to predicted COD, NH3-N and 2,4-DCP removal percentages of 94.8, 100 and 80.9%, respectively.

International audience; This study aimed to characterize biofilms from the paper industry and evaluate the effectiveness of enzymatic treatments in reducing them. The extracellular polymeric substances (EPS) extracted from six industrial biofilms were studied. EPS were mainly proteins, the protein to polysaccharide ratio ranging from 1.3 to 8.6 depending on where the sampling point was situated in the paper making process. Eight hydrolytic enzymes were screened on a 24-h multi-species biofilm. The enzymes were tested at various concentrations and contact durations. Glycosidases and lipases were inefficient or only slightly efficient for biofilm reduction, while proteases were more efficient: after treatment for 24 h with pepsin, Alcalase® or Savinase®, the removal exceeded 80%. Savinase® appeared to be the most adequate for industrial conditions and was tested on an industrial biofilm sample. This enzyme led to a significant release of proteins from the EPS matrix, indicating its potential efficiency on an industrial scale.

The use of enzymes has a high potential in the pulp and paper industry to improve the economics of the paper production process and to achieve, at the same time, a reduced environmental impact. Specific enzymes contribute to reduce the amount of chemicals and energy required for the modification of fibers and helps to prevent the formation or development of biofilms. This review is aimed at presenting the latest progresses made in the application of enzymes as refining aids and biofilm control agents.

A paper mill producing 500,000 ton of graphic paper annually has an onsite wastewater treatment plant that treats 7,240,000 m3 of wastewater per year, mechanically first, then biologically and at last by ozonation. Increased paper production capacity led to higher COD load in the mill effluent while production of higher proportions of brighter products gave worse biodegradability. Therefore the biological capacity of the WWTP needed to be increased and extra measures were necessary to enhance the efficiency of COD reduction. The full scale implementation of one MBBR with a volume of 1,230 m3 was accomplished in 2000 followed by another MBBR of 2,475 m3 in 2002. An ozonation step with a capacity of 75 kg O3/h was added in 2004 to meet higher COD reduction demands during the production of brighter products and thus keeping the given outflow limits. Adding a moving bed biofilm reactor prior to the existing activated sludge step gives: (i) cost advantages when increasing biological capacity as higher COD volume loads of MBBRs allow smaller reactors than usual for activated sludge plants; (ii) a relief of strain from the activated sludge step by biological degradation in the MBBR; (iii) equalizing of peaks in the COD load and toxic effects before affecting the activated sludge step; (iv) a stable volume sludge index below 100 ml/g in combination with an optimization of the activated sludge step allows good sludge separation—an important condition for further treatment with ozone. Ozonation and subsequent bio-filtration pre-treated waste water provide: (i) reduction of hard COD unobtainable by conventional treatment; (ii) controllable COD reduction in a very wide range and therefore elimination of COD-peaks; (iii) reduction of treatment costs by combination of ozonation and subsequent bio-filtration; (iv) decrease of the color in the ozonated wastewater. The MBBR step proved very simple to operate as part of the biological treatment. Excellent control of the COD-removal rate in the ozone step allowed for economical usage and therefore acceptable operation costs in relation to the paper production.

The antifouling potential of electric polarization combined and not combined with biocides was studied in nonsaline warm water with high organic content. Deinococcus geothermalis is a bacterium known for forming colored biofilms in paper machines and for its persistence against cleaning and chemical treatments. When D. geothermalis biofilms grown for 24 h in simulated paper machine water were exposed to cathodic or cathodically weighted pulsed polarization at least 60% (P < 0.05) of the biofilms were removed from stainless steel (AISI 316L). Biofilm removal by 25 ppm (effective substances 5-25 ppm) of oxidizing biocides (bromochloro-5,5-dimethylhydantoin, 2,2-dibromo-2-cyanoacetamide, peracetic acid) increased to 70% when combined with cathodically weighted pulsed polarization. Using a novel instrument that allows real-time detection of reactive oxygen species (ROS) we showed that the polarization program effective in antifouling generated ROS in a pulsed manner on the steel surface. We thus suggest that the observed added value of oxidative biocides combined with polarization depended on ROS. This suggestion was supported by the finding that a reductive biocide, methylene bisthiocyanate, counteracted the antifouling effect of polarization.

The aim of this study was to determine the effect of varying nutrient conditions on biofilm formation of a Pseudomonas putida strain isolated from a paper machine under controlled conditions. Biofilm accumulation, was investigated using a laminar flow cell reactor in a defined mineral medium. Our results indicate that increasing nutrient concentration (from 0.1 to 0.5 g l−1 glucose, C/N=40, C/P=100) or phosphate concentration (from C/P=200 to C/P=100) increased the rate and extent of biofilm accumulation, however, higher nutrient (1 g l−1 glucose, C/N=40, C/P=100) or phosphate (C/P=50) concentration reduced biofilm accumulation rate because of a higher detachment. The rate and extent of biofilm accumulation increased with nitrogen concentration (from C/N=90 to C/N=20). Detachment is a key parameter that influences biofilm accumulation since the early stage (2 h) of colonisation and strongly depends on nutrient conditions. In practice, controlling nutrient levels may be interesting to reduce biofilm formation in the paper industry.

This paper reviews nutrient issues within the pulp and paper industry summarising: nitrogen and phosphorus cycles within treatment systems; sources of nutrients within pulping and papermaking processes; minimising nutrient discharge; new approaches to nutrient minimisation; and the impact of nutrients in the environment. Pulp and paper industry wastewaters generally contain insufficient nitrogen and phosphorus to satisfy bacterial growth requirements. Nutrient limitation has been linked to operational problems such as sludge bulking and poor solids separation. Nutrients have been added in conventional wastewater treatment processes to ensure optimum treatment performance. Minimising the discharge of total nitrogen and phosphorus from a nutrient limited wastewater requires both optimised nutrient supplementation and effective removal of suspended solids from the treated wastewater. In an efficiently operated wastewater treatment system, the majority of the discharged nutrients are contained within the biomass. Effective solids separation then becomes the controlling step, and optimisation of secondary clarification is crucial. Conventional practice is being challenged by the regulatory requirement to reduce nitrogen and phosphorus discharge. Two recent developments in pulp and paper wastewater treatment technologies can produce discharges low in nitrogen and phosphorus whilst operating under conventionally nutrient limited conditions: i) the nutrient limited BAS process (Biofilm-Activated Sludge) which combines biofilm and activated sludge technologies under nutrient limited conditions and ii) an activated sludge process based on the use of nitrogen-fixing bacteria. Aerated stabilisation basins often operate without nutrient addition, relying on settled biomass in the benthal zone feeding back soluble nutrients, or the fixation of atmospheric nitrogen. Thus effective nutrient minimisation strategies require a more detailed understanding of nutrient cycling and utilisation. Where it is not possible to meet discharge constraints with biological treatment alone, a tertiary treatment step may be required. In setting nutrient control guidelines, consideration should be given to the nutrient limitations of the receiving environment, including other cumulative nutrient impacts on that environment. Whether an ecosystem is N or P limited should be integrated with wastewater treatment considerations in the further design and development of treatment technology and regulatory guidelines. End-of-pipe legislation alone cannot predict environmental effects related to nutrients and must be supplemented by an effects-based approach.

Thermophilic aerobic treatment of settled pulp and paper mill effluent was studied under mill premises with two comparative pilot processes; suspended carrier biofilm process (SCBP) and activated sludge process (ASP). Full-scale mesophilic activated sludge process was a reference treatment. During the runs (61 days) hydraulic retention times (HRTs) were kept 13+/-5 h and 16+/-6 h for SCBP and ASP, respectively. Corresponding volumetric loadings rates (VLR) were 2.7+/-0.9 and 2.2+/-1.0 kg CODfilt m(-3)d(-1). Temperatures varied between 46 to 60 degrees C in both processes. Mesophilic ASP was operated with HRT of 36 h, corresponding VLR of 0.7 kg CODfilt m(-3)d(-1). Both SCBP and ASP achieved CODfilt (GF/A filtered) removals up to 85%, while the mesophilic ASP removal was 89+/-2%. NTU values were markedly higher (100-300) in thermophilic effluents than in mesophilic effluent (30). Effluent turbidity was highly dependent on temperature; in batch experiment mesophilic effluent sample had NTU values of 30 and 60 at 35 degrees C and 55 degrees C, respectively. As a conclusion, both thermophilic treatments gave high CODfilt removals, which were close to mesophilic process removal and were achieved with less than half of HRT.

A combination of the suspended carrier biofilm process and the activated sludge process (biofilm-activated sludge--BAS) has been shown to be very successful for the treatment of different types of pulp and paper mill effluents. The robust biofilm pre-treatment in combination with activated sludge results in a stable, compact and highly efficient process. Recent findings have shown that nutrient limited operation of the biofilm process greatly improves the sludge characteristics in the following activated sludge stage, while minimising sludge production and effluent discharge of nutrients. The nutrient limited BAS process was implemented at full scale at the Södra Cell Värö kraft mill and taken into operation in July 2002. After start-up and optimisation over about 5 months, the process meets all effluent discharge limits. The removal of COD is close to 70% and the removal of EDTA greater than 90%. Typical effluent concentrations of suspended solids and nutrients during stable operations have been 20-30 mg/L TSS, 0.3-0.5 mg/L phosphorus and 3-5 mg/L nitrogen. The sludge production was 0.09 kgSS/kg COD removed and the sludge volume index was 50-100 mL/g.

An investigation was conducted to explore the lignin-degrading capacity of attached-growth white-rot fungi. Five white-rot fungi, Phanerochaete chrysosporium, Pleurotus ostreatus, Lentinus edodes, Trametes versicolor and S22, grown on a porous plastic media, were individually used to treat black liquor from a pulp and paper mill. Over 71% of lignin and 48% of chemical oxygen demand (COD) were removed from the wastewater. Several factors, including pH, concentrations of carbon, nitrogen and trace elements in wastewater, all had significant effects on the degradation of lignin and the removal of COD. Three white-rot fungi, P. chrysosporium, P. ostreatus and S22, showed high capacity for lignin degradation at pH 9.0-11.0. The addition of 1 g l-1 glucose and 0.2 g l-1 ammonium tartrate was beneficial for the degradation of lignin by the white-rot fungi studied.

A process of advanced oxidation treatment discussed here has been developed in the last seven years. A technical plant for treatment of paper mill wastewater started operation in 1999. Advanced oxidation treatment is defined here as a combination of chemical and biochemical oxidation applied to a completely biodegraded effluent. The process is a combination of ozonation with following biodegradation in a biofilm reactor. The combination makes use of the effect of partial oxidation in which with reduced use of expensive chemical oxidants persistent COD becomes biodegradable. A far-reaching elimination of AOX, colour and other disturbing substances like complexing agents is achieved simultaneously. The partially oxidized compounds, now biodegradable, are eliminated in a following biofilm reactor. Using single-stage systems, COD elimination rates of up to 60% are achieved with 0.4 to 1.0 g ozone per g COD eliminated in the combined process. With a two-stage system COD elimination of 85% with 0.65 g/g has been achieved in pilot tests. The quality of the treated effluent suits fresh water requirements for most types of paper production, so that either a total closure of the water circuits or - due to increasing concentrations of electrolytes, which are not eliminated by this process - a substantial closure to about 20% of the original amount of fresh water can be achieved.

A two-stage pilot-scale thermophilic aerobic suspended carrier biofilm process (SCBP) was set up for the on-site treatment of pulp and paper mill whitewater lining. The microbial diversity in this process was analyzed by length heterogeneity analysis of PCR-amplified 16S ribosomal DNA. The primer pair selected for PCR amplification was first evaluated by a computational analysis of fragment lengths in ten main phylogenetical eubacterial groups. The fragment contained the first third of the 16S rRNA gene, which was shown to vary naturally between 465 and 563 bp in length. The length heterogeneity analysis of polymerase chain reaction (LH-PCR) profile of the biomass attached to carrier elements was found to be diverse in both stages of the SCBP. During normal operating conditions, sequences belonging to beta-Proteobacteria, Cytophaga/Flexibacter/Bacteroides group and gamma-Proteobacteria were assigned to the most prominent LH-PCR peak. Samples from the suspended biomass consisted of completely different bacterial populations, which were, however, similar in the serial reactors. The pilot process experienced alkaline shocks, after which Bacillus-like sequences were detected in both the biofilm and suspended biomass. However, when the conditions were reversed, the normal microbial population in the biofilm recovered rapidly without further biomass inoculations. This study shows that LH-PCR is a valuable method for profiling microbial diversity and dynamics in industrial wastewater processes.

In this paper we describe an alternative flow-chart for full treatment of wastewaters rich in organic substrates, ammonia (or organic nitrogen), and sulfate, such as those generated in fish cannery industries. Biogas generated during anaerobic pretreatment of these wastewaters is rich in hydrogen sulfide that needs to be removed to enable application of the biogas. Nitrogen elimination is traditionally achieved by subsequent nitrification and denitrification of the effluent of the anaerobic reactor. Alternatively, the hydrogen sulfide in the biogas can be applied as an electron donor in an autotrophic post-denitrification step. In order to study whether sufficient hydrogen sulfide containing biogas for denitrification was produced in the anaerobic reactor, the biogas composition as a function of the anaerobic reactor-pH was estimated based on a typical wastewater composition and chemical equilibrium equations. It is demonstrated that typical sulfate and nitrogen concentrations in fish cannery wastewater are highly appropriate for application of autotrophic post-denitrification. A literature review furthermore suggested that the kinetic parameters for autotrophic denitrification by Thiobacillus denitrificans represent no bottleneck for its application. Initial experimental studies in fixed-film reactors were conducted with sodium sulfide and nitrate as an electron donor-acceptor couple. The results revealed that only moderate volumetric treatment capacities (1 g-NO3- N l(-1) day(-1)) could be achieved. Mass balances suggested that incomplete sulfide oxidation to elemental sulfur occurred, limiting biomass retention and the treatment capacity of the reactor. Future research should clarify the questions concerning product formation from sulfide oxidation.

Paper machine biofilms formed in situ on stainless steel surfaces were studied. A robust flow cell was fitted to side stream (1.8 m s(-1)) of the spray water circuit of a paper machine. This on-site tool allowed for assessing the efficacy of antifoulants and the adequacy of steel polishing under mill conditions. A rapid fluorescence-based assay was developed to quantify the biomass of shallow biofilms on machine steel. The fluorescence matched the ATP content measured for the same biofilms. Electrolytic polishing reduced the tendency of biofouling of 500 grit surface steel. Biofilm grew under machine conditions as clusters on the steels, showing uniformly coccoid, filaments or short rods; only one cell type in each cluster. The biofilm clusters excluded latex beads of 0.02 microm with hydrophilic or with hydrophobic surfaces from penetrating more than three to four layers of cells. Under the high hydraulic flow at the machine (1.8 m s(-1)), the biofilm grew in 7 days 6-10 microm thick. The high flow rate guided the shape of the biofilm clusters emerging after the primary attachment of cells. Adhered individual bacteria were the platform on steel to which solids such as paper machine fines then accumulated.

Biofilms of paper mill bacteria were cultivated in paper mill white water-simulating conditions on glass slides or stainless steel coupons in a laboratory culture system. The sugar content and composition of the biofilms were analysed and compared with the sugar composition of paper mill slimes. Acid methanolysis followed by gas chromatography revealed that Burkholderia was the major biofilm producer in pure culture, producing up to 50 microg of biofilm sugar cm(-2) in 5 days in rich medium and 10 microg in paper mill simulating medium. A mixture of simulated paper mill water with a culture medium yielded more biofilm (100 microg cm(-2)) than either of the media alone, so the biofilm accumulation was not proportional to the available substrate. More biofilm accumulated on stainless steel coupons than on glass slides, and the steel-coupon biofilms contained slightly more uronic acids. The biofilm sugars contained mainly galactose, glucose, mannose, and rhamnose. In paper mill medium, the Burkholderia biofilm contained more galactose and glucose, and less rhamnose, than in rich laboratory medium. The sugar composition of paper mill slimes was quite similar to those of steel-cultured Burkholderia cepacia biofilms. This suggests that Burkholderia cepacia is responsible for much of the slime in the paper mill.

Mechanisms for the undesired persistence of Bacillus species in paper machine slimes were investigated. Biofilm formation was measured for industrial Bacillus isolates under paper machine wet-end-simulating conditions (white water, pH 7, agitated at 45°C for 1–2 days). None of the 40 tested strains of seven Bacillus species formed biofilm on polished stainless steel or on polystyrene surfaces as a monoculture. Under the same conditions, Deinococcus geothermalis E50051 covered all test surfaces as a patchy thick biofilm. The paper machine bacilli, however, formed mixed biofilms with D. geothermalis E50051 as revealed by confocal microscopy. Biofilm interactions between the bacilli and the deinococci varied from synergism to antagonism. Synergism in biofilm formation of D. geothermalis E50051 was strongest with Bacillus coagulans D50192, and with the type strains of B. coagulans, B. amyloliquefaciens or B. pumilus. Two B. licheniformis, one B. amyloliquefaciens, one B. pumilus and four B. cereus strains antagonized biofilm production by D. geothermalis. B. licheniformis D50141 and the type strain of B. licheniformis were the strongest antagonists. These bacteria inhibited deinococcal growth by emitting heat-stable, methanol-soluble metabolite(s). We conclude that the persistence of Bacillus species in paper machine slimes relates to their ability to conquer biofilms formed by primary colonizers, such as D. geothermalis. Journal of Industrial Microbiology & Biotechnology (2001) 27, 343–351.