Decreasing Enterobacter sakazakii (Cronobacter spp.) food contamination level with bacteriophages: prospects and problems

Enterobacter sakazakii (Cronobacter spp.) is an opportunistic pathogen, which can cause rare, but life‐threatening infections in neonates and infants through feeding of a contaminated milk formula. We isolated 67 phages from environmental samples and tested their lytic host range on a representative collection of 40 E. sakazakii strains. A cocktail of five phages prevented the outgrowth of 35 out of 40 test strains in artificially contaminated infant formula. Two E. sakazakii phages represented prolate head Myoviridae. Molecular tests identified them as close relatives of Escherichia coli phage T4. The remaining three phages represented isometric head Myoviridae with large genome size of 140 and 200 kb, respectively, which belonged to two different DNA hybridization groups. A high dose of 10⁸ pfu ml^?1 of phage could effectively sterilize a broth contaminated with both high and low pathogen counts (10⁶ and 10² cfu ml^?1). In contrast, broth inoculated with 10⁴ phage and 10² bacteria per ml first showed normal bacterial growth until reaching a cell titre of 10⁵ cfu ml^?1. Only when crossing this threshold, phage replication started, but it could not reduce the contamination level below 100 cfu ml^?1. Phages could be produced with titres of 10¹⁰ pfu ml^?1 in broth culture, but they were not stable upon freeze‐drying. Addition of trehalose or milk formula stabilized the phage preparation, which then showed excellent storage stability even at elevated temperature.     

Rapid detection of bacteriophages in starter culture using water-in-oil-in-water emulsion microdroplets

Bacteriophage contamination of starter culture and raw material poses a major problem in the fermentation industry. In this study, a rapid detection of lytic phage contamination in starter culture using water-in-oil-in-water (W/O/W) emulsion microdroplets was described. A model bacteria with varying concentrations of lytic phages were encapsulated in W/O/W emulsion microdroplets using a simple needle-in-tube setup. The detection of lytic phage contamination was accomplished in 1 h using the propidium iodide labeling of the phage-infected bacteria inside the W/O/W emulsion microdroplets. Using this approach, a detection limit of 10² PFU/mL of phages was achieved quantitatively, while 10⁴ PFU/mL of phages could be detected qualitatively based on visual comparison of the fluorescence images. Given the simplicity and sensitivity of this approach, it is anticipated that this method can be adapted to any strains of bacteria and lytic phages that are commonly used for fermentation, and has potential for a rapid detection of lytic phage contamination in the fermentation industry.     

Phage Therapy as an Approach to Prevent Vibrio anguillarum Infections in Fish Larvae Production

Fish larvae in aquaculture have high mortality rates due to pathogenic bacteria, especially the Vibrio species, and ineffective prophylactic strategies. Vaccination is not feasible in larvae and antibiotics have reduced efficacy against multidrug resistant bacteria. A novel approach to controlling Vibrio infections in aquaculture is needed. The potential of phage therapy to combat vibriosis in fish larvae production has not yet been examined. We describe the isolation and characterization of two bacteriophages capable of infecting pathogenic Vibrio and their application to prevent bacterial infection in fish larvae. Two groups of zebrafish larvae were infected with V. anguillarum (∼10⁶ CFU mL⁻¹) and one was later treated with a phage lysate (∼10⁸ PFU mL⁻¹). A third group was only added with phages. A fourth group received neither bacteria nor phages (fish control). Larvae mortality, after 72 h, in the infected and treated group was similar to normal levels and significantly lower than that of the infected but not treated group, indicating that phage treatment was effective. Thus, directly supplying phages to the culture water could be an effective and inexpensive approach toward reducing the negative impact of vibriosis in larviculture.     

Characterization of Novel Virulent Broad-Host-Range Phages of Xylella fastidiosa and Xanthomonas

The xylem-limited bacterium Xylella fastidiosa is the causal agent of several plant diseases, most notably Pierce''s disease of grape and citrus variegated chlorosis. We report the isolation and characterization of the first virulent phages for X. fastidiosa, siphophages Sano and Salvo and podophages Prado and Paz, with a host range that includes Xanthomonas spp. Phages propagated on homologous hosts had observed adsorption rate constants of ∼4 × 10⁻¹² ml cell⁻¹ min⁻¹ for X. fastidiosa strain Temecula 1 and ∼5 × 10⁻¹⁰ to 7 × 10⁻¹⁰ ml cell⁻¹ min⁻¹ for Xanthomonas strain EC-12. Sano and Salvo exhibit >80% nucleotide identity to each other in aligned regions and are syntenic to phage BcepNazgul. We propose that phage BcepNazgul is the founding member of a novel phage type, to which Sano and Salvo belong. The lysis genes of the Nazgul-like phage type include a gene that encodes an outer membrane lipoprotein endolysin and also spanin gene families that provide insight into the evolution of the lysis pathway for phages of Gram-negative hosts. Prado and Paz, although exhibiting no significant DNA homology to each other, are new members of the phiKMV-like phage type, based on the position of the single-subunit RNA polymerase gene. The four phages are type IV pilus dependent for infection of both X. fastidiosa and Xanthomonas. The phages may be useful as agents for an effective and environmentally responsible strategy for the control of diseases caused by X. fastidiosa.     

Argentinean Lactococcus lactis bacteriophages: genetic characterization and adsorption studies

Aims: Characterization of four virulent Lactococcus lactis phages (CHD, QF9, QF12 and QP4) isolated from whey samples obtained from Argentinean cheese plants. Methods and Results: Phages were characterized by means of electron microscopy, host range and DNA studies. The influence of Ca²⁺, physiological cell state, pH and temperature on cell adsorption was also investigated. The double‐stranded DNA genomes of these lactococcal phages showed distinctive restriction patterns. Using a multiplex PCR, phage QP4 was classified as a member of the P335 polythetic species while the three others belong to the 936 group. Ca²⁺ was not needed for phage adsorption but indispensable to complete cell lysis by phage QF9. The lactococci phages adsorbed normally between pH 5 and pH 8, and from 0°C to 40°C, with the exception of phage QF12 which had an adsorption rate significantly lower at pH 8 and 0°C. Conclusions: Lactococcal phages from Argentina belong to the same predominant groups of phages found in other countries and they have the same general characteristics. Significance and Impact of the Study: This work is the first study to characterize Argentinean L. lactis bacteriophages.     

Acetobacter senegalensis isolated from mango fruits: Its polyphasic characterization and adaptation to protect against stressors in the industrial production of vinegar: A review

It has been more than a decade since Acetobacter senegalensis was isolated, identified and described as a thermotolerant strain of acetic acid bacteria. It was isolated from mango fruits in Senegal and used for industrial vinegar production in developing countries, mainly in sub-Saharan Africa. The strain was tested during several spirit vinegar fermentation processes at relatively high temperatures in accordance with African acclimation. The upstream fermentation process had significant stress factors, which are highlighted in this review so that the fermentation process can be better controlled. Due to its high industrial potential, this strain was extensively investigated by diverse industrial microbiologists worldwide; they concentrated on its microbiological, physiological and genomic features. A research group based in Belgium proposed an important project for the investigation of the whole-genome sequence of A. senegalensis. It would use a 454-pyrosequencing technique to determine and corroborate features that could give this strain significant diverse bio-industrial applications. For instance, its application in cocoa bean fermentation has made it a more suitable acetic acid bacterium for the making of chocolate than Acetobacter pasteurianus. Therefore, in this paper, we present a review that summarizes the current research on A. senegalensis at its microbial and genomic levels and also its specific bio-industrial applications, which can provide economic opportunities for African agribusiness. This review summarizes the physiological and genomic characteristics of Acetobacter senegalensis, a thermotolerant strain isolated from mango fruits and intended to be used in industrial vinegar fermentation processes. It also explores other bio-industrial applications such as cocoa fermentation. Vinegar fermentation is usually performed with mesophilic strains in temperate regions of the world. Developing countries, such as Senegal, import vinegar or make ‘fake’ vinegar by diluting acetic acid obtained from petrochemicals. The use of a thermotolerant Acetobacter senegalensis strain as a solid functional starter culture, as well as the design of a new adapted bioreactor, has significantly contributed to food security and the creation of small- to medium-sized enterprises that produce mango vinegar in West Africa.     

Virulent Bacteriophage for Efficient Biocontrol of Listeria monocytogenes in Ready-To-Eat Foods

Food-borne Listeria monocytogenes is a serious threat to human health, and new strategies to combat this opportunistic pathogen in foods are needed. Bacteriophages are natural enemies of bacteria and are suitable candidates for the environmentally friendly biocontrol of these pathogens. In a comprehensive set of experiments, we have evaluated the virulent, broad-host-range phages A511 and P100 for control of L. monocytogenes strains Scott A (serovar 4b) and WSLC 1001 (serovar 1/2a) in different ready-to-eat (RTE) foods known to frequently carry the pathogen. Food samples were spiked with bacteria (1 × 10³ CFU/g), phage added thereafter (3 × 10⁶ to 3 × 10⁸ PFU/g), and samples stored at 6°C for 6 days. In liquid foods, such as chocolate milk and mozzarella cheese brine, bacterial counts rapidly dropped below the level of direct detection. On solid foods (hot dogs, sliced turkey meat, smoked salmon, seafood, sliced cabbage, and lettuce leaves), phages could reduce bacterial counts by up to 5 log units. Variation of the experimental conditions (extended storage over 13 days or storage at 20°C) yielded similar results. In general, the application of more phage particles (3 × 10⁸ PFU/g) was more effective than lower doses. The added phages retained most of their infectivity during storage in foods of animal origin, whereas plant material caused inactivation by more than 1 log₁₀. In conclusion, our data demonstrate that virulent broad-host-range phages, such as A511 and P100, can be very effective for specific biocontrol of L. monocytogenes in contamination-sensitive RTE foods.     

Direct Electron Microscopy Study on the Morphological Diversity of Bacteriophage Populations in Lake Plußsee

Direct electron microscopy of bacteriophages adsorbed to a carbon film without prior enrichment by specific host strains or concentration by physical or chemical methods was used to study the morphological diversity of natural bacteriophage assemblages in a North German lake. All samples contained a mixture of morphologically different tailed viruses, which were regarded as bacteriophages. Most of them had isometric heads and long noncontractile tails, belonging to morphotype B1 (Siphoviridae). In addition, members of morphotypes A1 (Myoviridae), B2 (Siphoviridae with elongated heads), and C1 (Podoviridae) were present in lower numbers. Only one cubic virus was detected, while no filamentous or pleomorphic phages were found. Up to 11 different phages per sample, and a total of 39 phages when all samples were considered together, could be distinguished by morphological criteria. The total number of phages was estimated to be on the order of 10⁸/ml.     

Genome Sequence and Characteristics of Lrm1, a Prophage from Industrial Lactobacillus rhamnosus Strain M1

Prophage Lrm1 was induced with mitomycin C from an industrial Lactobacillus rhamnosus starter culture, M1. Electron microscopy of the lysate revealed relatively few intact bacteriophage particles among empty heads and disassociated tails. The defective Siphoviridae phage had an isometric head of approximately 55 nm and noncontractile tail of about 275 nm with a small baseplate. In repeated attempts, the prophage could not be cured from L. rhamnosus M1, nor could a sensitive host be identified. Sequencing of the phage Lrm1 DNA revealed a genome of 39,989 bp and a G+C content of 45.5%. A similar genomic organization and mosaic pattern of identities align Lrm1 among the closely related Lactobacillus casei temperate phages A2, ΦAT3, and LcaI and with L. rhamnosus virulent phage Lu-Nu. Of the 54 open reading frames (ORFs) identified, all but 8 shared homology with other phages of this group. Five unknown ORFs were identified that had no homologies in the databases nor predicted functions. Notably, Lrm1 encodes a putative endonuclease and a putative DNA methylase with homology to a methylase in Lactococcus lactis phage Tuc2009. Possibly, the DNA methylase, endonuclease, or other Lrm1 genes provide a function crucial to L. rhamnosus M1 survival, resulting in the stability of the defective prophage in its lysogenic state. The presence of a defective prophage in an industrial strain could provide superinfection immunity to the host but could also contribute DNA in recombination events to produce new phages potentially infective for the host strain in a large-scale fermentation environment.     

Morphology of the Bacteriophages of Lactic Streptococci

Electron microscope studies have been made of a number of phages of lactic streptococci, seven of which were phages of Streptococcus lactis C10. Two of the phages are thought to be identical; five have been classified by the method of Tikhonenko as belonging to group IV (phages with noncontractile tails) with type III tail plates; one belongs to group V (phages with tails possessing a contractile sheath). Both prolate polyhedral heads and isometric polyhedral heads are represented among the group IV phages. The phage drc3 of S. diacetilactis DRC3 has been shown to have similar structure to the group IV phages of S. lactis C10 with prolate polyhedral heads. The phages ml1, hp, c11, and z8 of the S. cremoris strains ML1, HP, C11, and Z8, respectively, were shown to belong to the group IV phages with type III tail plates by the method of Tikhonenko. All had octahedral heads and tended to be larger than most of the other phages studied.     

Tumor specific phage particles promote tumor regression in a mouse melanoma model

Within cancer research, phage display libraries have been widely used for the identification of tumor targeting peptides and antibodies. Additionally, phages are known to be highly immunogenic; therefore we evaluated the immunotherapeutic potential of tumor specific phages to treat established solid tumors in a mouse model of melanoma. We developed two tumor specific phages, one derived from a peptide phage display library and one Fab expressing phage with known specificity, for the treatment of mice bearing palpable B16-F10 or B16/A2K^b tumors. Therapy in B16-F10 tumor bearing mice with tumor specific phages was superior to treatment with non-tumor specific phages and lead to delayed tumor growth and increased survival. In B16/A2K^b tumor bearing mice, therapy with tumor specific phages resulted in complete tumor regression and long-term survival in 50% of the mice. Histological analysis of tumors undergoing treatment with tumor specific phages revealed that phage administration induced a massive infiltration of polymorphonuclear neutrophils. Furthermore, phages induced secretion of IL-12 (p70) and IFN-γ as measured in mouse splenocyte culture supernatants. These results demonstrate a novel, immunotherapeutic cancer treatment showing that tumor specific phages can promote regression of established tumors by recruitment of inflammatory cells and induction of Th1 cytokines.     

Enumeration of Bacteriophages and Host Bacteria in Sewage and the Activated-Sludge Treatment Process

Bacteriophage populations in an activated-sludge sewage treatment plant were enumerated. A newly developed assay for quantitation of total phages, employing direct electron microscopic counts, was used in conjunction with the plaque assay. The total concentration of phages was significantly higher in reactor mixed liquor and effluent than in influent sewage, indicating a net production of phages within the reactor. Maximum total phage concentrations in the fluid phase of sewage, activated-sludge mixed liquor, and reactor effluent were 2.2 × 10⁷, 9.5 × 10⁷, and 8.4 × 10⁷/ml, respectively. Conditions were optimized for isolation of predominant heterotrophic aerobic bacteria from sewage and mixed liquor. Blending at ice water temperatures was superior to ultrasound or enzyme treatments for maximum release of viable bacteria from microbial floc. A solidified extract of mixed liquor was superior to standard media for cultivating maximum numbers of heterotrophic bacteria. The highest culture counts for sewage and mixed liquor were 1.4 × 10⁷ and 1.3 × 10⁹/ml, respectively, which represented only 3 and 6.8% of the total microscopic cell counts. Only 3 out of 48 dominant bacterial isolates from either mixed liquor or sewage were hosts for phages present in the system. The sum of phage populations infecting these three hosts accounted for, at best, 3.8% (sewage) and 0.2% (mixed liquor) of the total number of phages present. Generally, specific phage titers were lower in mixed liquor than in sewage, indicating that these hosts were not responsible for the net production of phages in the reactor. This study emphasizes the limitations of the plaque assay for ecological studies of phages, and it suggests that bacteria responsible for phage production in activated-sludge mixed liquor are either minor components of the heterotrophic population, floc-producing strains, or members of other physiological groups.     

Resistance against Industrial Bacteriophages Conferred on Lactococci by Plasmid pAJ1106 and Related Plasmids

Plasmid pAJ1106 and its deletion derivative, plasmid pAJ2074, conferred lactose-fermenting ability (Lac) and bacteriophage resistance (Hsp) at 30°C to Lac⁻ proteinase (Prt)-negative Lactococcus lactis subsp. lactis and L. lactis subsp. lactis var. diacetylactis recipient strains. An additional plasmid, pAJ331, isolated from the original source strain of pAJ1106, retained Hsp and conjugative ability without Lac. pAJ331 was conjugally transferred to two L. lactis subsp. lactis and one L. lactis subsp. cremoris starter strains. The transconjugants from such crosses acquired resistance to the phages which propagated on the parent recipient strains. Of 10 transconjugant strains carrying pAJ1106 or one of the related plasmids, 8 remained insensitive to phages through five activity test cycles in which cultures were exposed to a large number of industrial phages at incubation temperatures used in lactic casein manufacture. Three of ten strains remained phage insensitive through five cycles of a cheesemaking activity test in which cultures were exposed to approximately 80 different phages through cheesemaking temperatures. Three phages which propagated on transconjugant strains during cheesemaking activity tests were studied in detail. Two were similar (prolate) in morphology and by DNA homology to phages which were shown to be sensitive to the plasmid-encoded phage resistance mechanism. The third phage was a long-tailed, small isometric phage of a type rarely found in New Zealand cheese wheys. The phage resistance mechanism was partially inactivated in most strains at 37°C.     

Detection and Quantification of Flavobacterium psychrophilum-Specific Bacteriophages In Vivo in Rainbow Trout upon Oral Administration: Implications for Disease Control in Aquaculture

The use of bacteriophages in the treatment and prevention of infections by the fish pathogen Flavobacterium psychrophilum has attracted increased attention in recent years. It has been shown recently that phage delivery via the parenteral route resulted in immediate distribution of phages to the circulatory system and the different organs. However, little is known about phage dispersal and survival in vivo in rainbow trout after delivery via the oral route. Here we examined the dispersal and survival of F. psychrophilum phage FpV-9 in vivo in juvenile rainbow trout after administration by three different methods—bath, oral intubation into the stomach, and phage-coated feed—with special emphasis on the oral route of delivery. Phages could be detected in all the organs investigated (intestine, spleen, brain, and kidney) 0.5 h postadministration, reaching concentrations as high as ∼10⁵ PFU mg intestine⁻¹ and ∼10³ PFU mg spleen⁻¹ within the first 24 h following the bath and ∼10⁷ PFU mg intestine⁻¹ and ∼10⁴ PFU mg spleen⁻¹ within the first 24 h following oral intubation. The phages were most persistent in the organs for the first 24 h and then decreased exponentially; no phages were detected after 83 h in the organs investigated. Phage administration via feed resulted in the detection of phages in the intestine, spleen, and kidney 1 h after feeding. Average concentrations of ∼10⁴ PFU mg intestine⁻¹ and ∼10¹ PFU mg spleen⁻¹ were found throughout the experimental period (200 h) following continuous delivery of phages with feed. These experiments clearly demonstrate the ability of the phages to survive passage through the fish stomach and to penetrate the intestinal barrier and enter the circulatory system after oral delivery, although the quantity of phages found in the spleen was 100- to 1,000-fold lower than that in the intestine. It was also shown that phages could tolerate long periods of desiccation on the feed pellets, with 60% survival after storage at −80°C, and 10% survival after storage at 5°C, for ∼8 months. Continuous delivery of phages via coated feed pellets constitutes a promising method of treatment and especially prevention of rainbow trout fry syndrome.     

Interactions of Lactobacillus bulgaricus Temperate Bacteriophage 0448 with Host Strains

Lactobacillus bulgaricus LT4(0448) is a lysogenic strain from which a temperate bacteriophage can be induced by mitomycin C or UV irradiation. Lactobacillus lactis CNRZ 326 is an indicator strain for the temperate phage 0448, but this strain lyses only in the presence of Ca²⁺ ions. A resistant culture developed secondarily after phage lysis and grew normally in MRS broth but again lysed abruptly if Ca²⁺ ions were added after two or three transfers. This behavior of the secondary culture and its subcultures is explained by a heterogeneous and fluctuating bacterial population, including clones identical to L. lactis 326, which were sensitive to 0448 and which formed rough colonies, as does the indicator. The proportion of these clones increased in the course of transfers in MRS, explaining lysis when Ca²⁺ was added. The population also included clones which formed smooth colonies (S clones). SI clones, which could not be induced by mitomycin C, were the major type in the initial culture, although they were sensitive to temperate phage 0448. The SI population then decreased and was gradually replaced by SII clones, inducible by mitomycin C and resistant to 0448. These SII clones were lysogenized clones, 326(0448), whose stability was confirmed by growth in the presence of an antiphage serum. When L. bulgaricus LT4(0448) was treated with mitomycin C, several cured LT4 clones were obtained that were related to the clones of the indicator L. lactis 326; they formed rough colonies. They also became sensitive to lytic phages or temperate phages active against L. lactis 326 and insensitive to lytic phages which lysed L. bulgaricus LT4(0448). This suggests that phage 0448 can lead to a lysogenic conversion of host strain LT4.     

A Raman-spectroscopy-based approach for detection and discrimination of <Emphasis Type="Italic">Streptococcus thermophilus</Emphasis> and <Emphasis Type="Italic">Lactobacillus bulgaricus</Emphasis> phages at low titer in raw milk

In this study, a method combining Raman spectroscopy with chemometric analysis was developed for detection of phage presence in raw milk and discrimination of Streptococcus thermophilus and Lactobacillus bulgaricus phages which are among the main phages causing problems in dairy industry. For this purpose, S. thermophilus and L. bulgaricus phages were added into raw milk separately, and then some pretreatments such as fat separation, removal of casein, and filtration were applied to the raw milk samples. Raman spectra of the samples were collected and then analyzed using principal component analysis in order to discriminate these phages in raw milk. In the next step, dilutions of S. thermophilus phages in pretreated raw milk were prepared, and Raman spectra were collected. These spectra were analyzed by using partial least squares method to quantify phages in low titer. Consequently, it has been demonstrated that S. thermophilus and L. bulgaricus phages, which have titers sufficient to fail the fermentation (~?10⁷ pfu/mL) and have lower titers (10²–10³ pfu/mL), could be discriminated from antibiotic and each other. Additionally, low concentrations of S. thermophilus phages (10² pfu/mL) could be detected through Raman spectroscopy with a short analysis time (60 min) and high coefficient of determination (R²) values for both calibration (0.985) and validation (0.906) with a root mean square error of calibration of 70.54 and root mean square error of prediction of 165.47. However, a lower success was achieved with L. bulgaricus phages and the obtained coefficient of determination values were not sufficiently high (0.649).     

Amplification and Purification of T4-Like Escherichia coli Phages for Phage Therapy: from Laboratory to Pilot Scale

We investigated the amplification and purification of phage preparations with respect to titer, contamination level, stability, and technical affordability. Using various production systems (wave bags, stirred-tank reactors, and Erlenmeyer flasks), we obtained peak titers of 10⁹ to 10¹⁰ PFU/ml for T4-like coliphages. Phage lysates could be sterilized through 0.22-μm membrane filters without titer loss. Phages concentrated by differential centrifugation were not contaminated with cellular debris or bacterial proteins, as assessed by electron microscopy and mass spectrometry, respectively. Titer losses occurred by high-speed pelleting of phages but could be decreased by sedimentation through a sucrose cushion. Alternative phage concentration methods are prolonged medium-speed centrifugation, strong anion-exchange chromatography, and ultrafiltration, but the latter still allowed elevated lipopolysaccharide contamination. T4-like phages could not be pasteurized but maintained their infectivity titer in the cold chain. In the presence of 10 mM magnesium ions, phages showed no loss of titer over 1 month at 30°C.     

High-level production of arachidonic acid by fed-batch culture of <Emphasis Type="Italic"> Mortierella alpina</Emphasis> using NH<Subscript>4</Subscript>OH as a nitrogen source and pH control

Mortierella alpina was grown in a fed-batch culture using a 12-l jar fermenter with an initial 8-l working volume containing 20 g glucose l⁻¹ and 10 g corn-steep powder l⁻¹. Glucose was intermittently fed to give 32 g l⁻¹ at each time. The pH of culture was maintained using 14% (v/v) NH₄OH, which also acted as a nitrogen source. A final cell density of 72.5 g l⁻¹ was reached after 12.5 days with a content of arachidonic acid (ARA) at 18.8 g l⁻¹. These values were 4 and 1.8 times higher than the respective values in batch culture. Our results suggest that the combined feeding of glucose and NH₄⁺ to the growth of M. alpina could be applied for the industrial scale production of ARA.     

Studies of the Stability of Microbial Association Use in Industrial Biofiltering of Gaseous Discharges

Results of industrial exploitation of a biofiltration plant tailored for purifying gaseous discharges of hazardous organic components such as toluene, cyclohexane, and xylene, are examined. Both numerical and compositional variations were monitored for a long-term (more than 1.5 years) utilization process in an association of microorganisms decomposing organic pollutants. A population of microbial association composed by one yeast and two bacterial strains in the biofilm on the surface of filtering sheets was abundant (10⁸–10⁹ yeast cells/cm² and 10¹⁰–10¹¹ bacterial cells/cm²) and stable during the whole period of monitoring. A microbial association in the culture medium averaging 10⁶ yeast cells/l and 10⁸ bacterial cells/l is more susceptible to technogenic impacts and seasonal fluctuations. Overall, the biofilter as an open and autonomic system maintained its microbial association, thereby providing high-degree (93–98%) purification of industrial gaseous discharges from organic pollutants.     

The phage‐driven microbial loop in petroleum bioremediation

During the drilling process and transport of crude oil, water mixes with the petroleum. At oil terminals, the water settles to the bottom of storage tanks. This drainage water is contaminated with emulsified oil and water-soluble hydrocarbons and must be treated before it can be released into the environment. In this study, we tested the efficiency of a continuous flow, two-stage bioreactor for treating drainage water from an Israeli oil terminal. The bioreactor removed all of the ammonia, 93% of the sulfide and converted 90% of the total organic carbon (TOC) into carbon dioxide. SYBR Gold staining indicated that reactor 1 contained 1.7 × 10⁸ bacteria and 3.7 × 10⁸ phages per millilitre, and reactor 2 contained 1.3 × 10⁸ bacteria and 1.7 × 10⁹ phages per millilitre. The unexpectedly high mineralization of TOC and high concentration of phage in reactor 2 support the concept of a phage-driven microbial loop in the bioremediation of the drainage water. In general, application of this concept in bioremediation of contaminated water has the potential to increase the efficiency of processes.