Dual-species Cultures Research Articles

Concomitant Inhibition and Collaring of Dual-Species Biofilms Formed by Candida auris and Staphylococcus aureus by Triazole Based Small Molecule Inhibitors

Background/Objectives: Biofilm-associated infections, particularly those involving Candida auris and Staphylococcus aureus, pose significant challenges in clinical settings due to their resilience and resistance to conventional treatments. This study aimed to synthesize novel triazole derivatives containing a piperazine ring via click chemistry and evaluate their efficacy in disrupting biofilms formed by these pathogens. Methods: Triazole derivatives were synthesized using click chemistry techniques. The antimicrobial activity of the compounds was tested against planktonic cells of C. auris and S. aureus in single and dual-species culture conditions. Biofilm disruption efficacy was assessed, alongside the evaluation of physicochemical properties, oral bioavailability potential, and toxicity profiles. Results: The compound T3 demonstrated potent antimicrobial activity against planktonic cells of C. auris and S. aureus in both single and dual-species cultures. T3 exhibited significant efficacy in reducing microbial viability within biofilms formed by these pathogens. Physicochemical analyses revealed favorable solubility and permeability profiles, supporting its potential for oral bioavailability. Toxicity assessments showed a non-toxic profile, highlighting a promising safety margin for further development. Conclusions: This study underscores the anti-biofilm properties of novel triazole-piperazine derivatives, particularly T3, against single and dual-species biofilms of C. auris and S. aureus. These findings position T3 as a promising candidate for developing therapies targeting polymicrobial infections and provide a foundation for future research into alternative strategies for combating biofilm-associated infections.

Ceragenin-mediated disruption of Pseudomonas aeruginosa biofilms.

Microbial biofilms, as a hallmark of cystic fibrosis (CF) lung disease and other chronic infections, remain a desirable target for antimicrobial therapy. These biopolymer-based viscoelastic structures protect pathogenic organisms from immune responses and antibiotics. Consequently, treatments directed at disrupting biofilms represent a promising strategy for combating biofilm-associated infections. In CF patients, the viscoelasticity of biofilms is determined mainly by their polymicrobial nature and species-specific traits, such as Pseudomonas aeruginosa filamentous (Pf) bacteriophages. Therefore, we examined the impact of microbicidal ceragenins (CSAs) supported by mucolytic agents-DNase I and poly-aspartic acid (pASP), on the viability and viscoelasticity of mono- and bispecies biofilms formed by Pf-positive and Pf-negative P. aeruginosa strains co-cultured with Staphylococcus aureus or Candida albicans. The in vitro antimicrobial activity of ceragenins against P. aeruginosa in mono- and dual-species cultures was assessed by determining minimum inhibitory concentration (MIC) and minimum bactericidal/fungicidal concentration (MBC/MFC). Inhibition of P. aeruginosa mono- and dual-species biofilms formation by ceragenins alone and in combination with DNase I or poly-aspartic acid (pASP) was estimated by the crystal violet assay. Additionally, the viability of the biofilms was measured by colony-forming unit (CFU) counting. Finally, the biofilms' viscoelastic properties characterized by shear storage (G') and loss moduli (G"), were analyzed with a rotational rheometer. Our results demonstrated that ceragenin CSA-13 inhibits biofilm formation and increases its fluidity regardless of the Pf-profile and species composition; however, the Pf-positive biofilms are characterized by elevated viscosity and elasticity parameters. Due to its microbicidal and viscoelasticity-modifying properties, CSA-13 displays therapeutic potential in biofilm-associated infections, especially when combined with mucolytic agents.

Modeling interactions of Clostridium cadaveris and Clostridium sporogenes in anaerobic acidogenesis of glucose and peptone

This study focuses on developing a mathematical model to assess interaction among acidogenic bacteria during the anaerobic degradation of two substrates. Clostridium cadaveris and Clostridium sporogenes were cultured in various combinations with glucose and peptone. Parameter estimates are given for both conventional Monod parameters from single substrate-single species cultures and sum kinetics with interaction parameters obtained from dual substrate-single species cultures. The presence of multiple substrates led to both inhibitory and enhancing effects on biodegradation rates for dual substrates compared to single substrate cultures. A new model of interspecies interaction was developed within the framework of Lotka-Volterra incorporating substrate interaction parameters, with a focus on accuracy, realism, simplicity, and biological significance. The model demonstrated competitive interaction for resource sharing and the additional non-linearity parameter eliminated the constraint of the linear relationship between growth rate and population density.

Does Streptococcus oralis supernatant influence on the proliferation and virulence of Candida albicans?

ObjectiveTo evaluate the influence of Streptococcus oralis supernatant on the proliferation and virulence of Candida albicans. DesignS. oralis supernatant was obtained by filtration of overnight cultures. Single or dual-species cultures of C. albicans and S. oralis were cultivated in both planktonic and biofilm-based models. Planktonic culture growth was measured, and mature biofilms formed on resin disks were collected to measure biofilm metabolic activity, total biomass, and cell counts. Hyphae formation (virulence factor) and biofilm thickness were analyzed by confocal laser scanning microscopy. Data were analyzed by a one-way ANOVA test followed by the Tukey posthoc test (α = 0.05). ResultsWe found that S. oralis supernatant did not influence C. albicans proliferation in planktonic cultures. However, biofilms containing S. oralis supernatant showed higher cell metabolism than C. albicans monoculture biofilms and C. albicans–S. oralis dual-culture biofilms (p < 0.05). Though S. oralis supernatants did increase biofilm metabolic activity, they did not affect the total biomass and cell counts of C. albicans (p > 0.05). However, biofilm imaging revealed enhanced C. albicans hyphae formation in biofilms containing S. oralis supernatant compared to C. albicans monoculture biofilms. ConclusionsSecreted metabolites in S. oralis supernatant may contribute to C. albicans metabolism and virulence.

A sustainable approach to control biofilms infections and reduce medical waste: Catheters coated with antibiotics inhibit single and dual-species biofilms

Catheter-Associated Urinary Tract Infections (CAUTIs) are one of the major diseases that cause severe illness and death among the wider population. More than 30,000 deaths are reported each year due to CAUTI. These infections are caused due to different biofilm- forming species such as Escherichia coli (E. coli), Enterococcus faecalis (E. faecalis), Candida spp, Klebsiella spp, Proteus spp, Bacteroides spp, Staphylococcus aureus, and Enterobacter spp. This study examined the growth of two major uropathogenic (E. coli and E. faecalis) strains on catheter surfaces under antibiotic-treated and untreated conditions. The antibiotics used for this study are Ciprofloxacin and Doxycycline, which are considered to be broad-spectrum antibiotics. The Minimum Inhibitory Concentration (MIC) assay was performed to identify the concentration at which these antibiotics show efficient inhibition. The results show that both antibiotics have an inhibitory effect on single and dual-species biofilms. However, E. coli is more resistant to Doxycycline (MIC: 100 μg/mL), whereas E. faecalis is more resistant to Ciprofloxacin (MIC: 50 μg/mL). Interestingly, the dual-species cultures are more susceptible to both antibiotics at lower concentrations, 5μg/mL. Furthermore, a CFU assay was performed to quantify the results obtained, and a similar trend could be observed with around a 4-fold reduction in bacterial colonies when catheters are coated with antibiotics. In addition, the antibiotic-coated catheters contribute no pathogen contamination to the environment.

Influence of interspecies interactions on biomass and extracellular polymeric substances of bacterial biofilms

BackgroundStudies with emphasis on pure and mixed-species biofilms are of significant importance in understanding biofilm formation mechanisms during microbial infections. This research aims to evaluate pure- and dual-species biofilms of Escherichia coli (code A), Staphylococcus aureus (code B), Klebsiella pneumoniae (code C) and Pseudomonas aeruginosa (code D) pathogenic bacterial species and their production of biofilm exopolysaccharides at laboratory scale. MethodsBiofilm biomass (A595) of pure- and dual-species cultures was determined by means of a microtiter plate assay in triplicate using a microplate photometer (Fisher Scientific, type-357). Extracellular polymeric substances (EPS) and soluble microbial products (SMP) were extracted from the biofilm cells (pure- and dual-species cultures) using the alkaline-heat extraction method. Dry weights (g/L) of EPS and SMP were determined by drying the samples at 105 °C for 8 hours. ResultsKlebsiella pneumoniae biofilm biomass accounted for a 28-72% greater biofilm biomass than the other bacteria. Experimental values of dual-species biofilm biomasses were in the range of 6% to 30% over theoretical values. The experimental value of one dual-species (bacteria B + D) biofilm biomass was 30% higher than its expected value. Decrease or increase in the dual-species biofilm biomass of either bacteria A+C or bacteria B+C combinations was totally dependent on the cell density of bacteria C. Conclusions Biofilm biomass of pure-species cultures was totally species-dependent, and the biofilm biomass of four species was in the following order: bacteria C &gt; D &gt; A &gt; B. Relation between biofilm biomasses and SMP or EPS was inconsistent.

High biofilm-forming Pseudomonas strains isolated from poultry slaughterhouse surfaces: Their importance in the persistence of Salmonella enteritidis in slaughterhouses

The surfaces of poultry slaughterhouse equipment are significant sources of contamination with Pseudomonas strains, which leads to spoilage of poultry meat during subsequent refrigerated storage. In this study, Pseudomonas strains with high biofilm-forming ability were isolated from different surfaces of the poultry slaughterhouse equipment, identified based on molecular data, and characterized their biofilm-forming ability. After 24 h of incubation at 25 °C, 54 out of 58 Pseudomonas strains produced biofilm in vitro on polystyrene microplates. Seven isolates with high-ability to produce biofilm were identified as P. fragi (three strains), P. fluorescens (two strains), P. lundensis and P. cedrina. Despite their differences, these strains produced high amounts of biofilm in pure- and dual-species cultures with S. enteritidis on stainless steel surfaces. However, their ability to produce dual-species biofilms with S. enteritidis depends on whether S. enteritidis form the biofilm simultaneously with the Pseudomonas strains or whether Pseudomonas strains have already formed a biofilm. In concurrent inoculation, S. enteritidis participated in biofilm formation with all seven Pseudomonas strains with varying percent contributions. However, in delayed inoculation, S. enteritidis did not contribute in the biofilm formed by P. lundensis R26, P. fragi R39, and P. fluorescens R47. In addition to highlighting the complexity of bacterial interactions associated with Pseudomonas strains, these results showed that Pseudomonas strains can be implicated in Salmonella persistence in poultry slaughterhouses.

Increasing the Efficacy of Treatment of Staphylococcus aureus-Candida albicans Mixed Infections with Myrtenol.

Infectious diseases caused by various nosocomial microorganisms affect worldwide both immunocompromised and relatively healthy persons. Bacteria and fungi have different tools to evade antimicrobials, such as hydrolysis damaging the drug, efflux systems, and the formation of biofilm that significantly complicates the treatment of the infection. Here, we show that myrtenol potentiates the antimicrobial and biofilm-preventing activity of conventional drugs against S. aureus and C. albicans mono- and dual-species cultures. In our study, the two optical isomers, (-)-myrtenol and (+)-myrtenol, have been tested as either antibacterials, antifungals, or enhancers of conventional drugs. (+)-Myrtenol demonstrated a synergistic effect with amikacin, fluconazole, and benzalkonium chloride on 64-81% of the clinical isolates of S. aureus and C. albicans, including MRSA and fluconazole-resistant fungi, while (-)-myrtenol increased the properties of amikacin and fluconazole to repress biofilm formation in half of the S. aureus and C. albicans isolates. Furthermore, myrtenol was able to potentiate benzalkonium chloride up to sixteen-fold against planktonic cells in an S. aureus-C. albicans mixed culture and repressed the adhesion of S. aureus. The mechanism of both (-)-myrtenol and (+)-myrtenol synergy with conventional drugs was apparently driven by membrane damage since the treatment with both terpenes led to a significant drop in membrane potential similar to the action of benzalkonium chloride. Thus, due to the low toxicity of myrtenol, it seems to be a promising agent to increase the efficiency of the treatment of infections caused by bacteria and be fungi of the genus Candida as well as mixed fungal-bacterial infections, including resistant strains.

Intraspecific and interspecific extracellular metabolites remodel biofilms formed by thermophilic spoilage bacteria.

Thermophilic spoilage bacteria and their biofilms formed during milk powder processing posed threats to safety and quality of dairy products. This research aims to understand more about the bacterial behaviours and their social models in biofilms. Interactional effects from both extracellular metabolites and co-culture on biofilms formation of the contaminating thermophilic bacteria were determined. The results showed that strong biofilm formers always had high AI-2 activities, including Geobacillus stearothermophilus gs1, Bacillus licheniformis bl1 and Thermoactinomyces vulgaris tv1. Metabolites from themself or other species altered their biofilm biomass detected by crystal violet staining. Dual-species cultures observed by confocal laser scanning microscope indicated either synergistic or inhibitory effects between B. circulans bc1 and G. stearothermophilus gs1, as well as B. licheniformis bl1 and G. stearothermophilus gs1. Fourier transform infrared spectrometry results revealed the significant diversities in polysaccharides of the biofilm matrix. Cell communication played an important role on biofilm formation in the complex microbial community. Intraspecific and interspecific extracellular metabolites influenced collective bacterial behaviours under mixed circumstances. This research provided evidences on cell communication and biofilm formation of thermophilic bacteria in dairy industry.

Food web and metabolic interactions of the lung inhabitants Streptococcus pneumoniae and Pseudomonas aeruginosa.

Bacteria that successfully adapt to different substrates and environmental niches within the lung and overcome the immune defence can cause serious lung infections. Such infections are generally complex, and recognized as polymicrobial in nature. Both Pseudomonas aeruginosa and Streptococcus pneumoniae can cause chronic lung infections and were both detected in cystic fibrosis (CF) lung at different stages. In this study, single and dual species cultures of Pseudomonas aeruginosa and Streptococcus pneumoniae were studied under well-controlled planktonic growth conditions. Under pH-controlled conditions, both species apparently benefited from the presence of the other. In co-culture with P. aeruginosa, S. pneumoniae grew efficiently under aerobic conditions, whereas in pure S. pneumoniae culture, growth inhibition occurred in bioreactors with dissolved oxygen concentrations above the microaerobic range. Lactic acid and acetoin that are produced by S. pneumoniae were efficiently utilized by P. aeruginosa. In pH-uncontrolled co-cultures, the low pH triggered by S. pneumoniae assimilation of glucose and lactic acid production negatively affected the growth of both strains. Nevertheless, ammonia production improved significantly, and P. aeruginosa growth dominated at later growth stages. This study revealed unreported metabolic interactions of two important pathogenic microorganisms and shed new lights into pathophysiology of bacterial lung infection.

Synergistic Effects of Fluconazole Combined with Doxycycline Against Dual-Species Cultures of Candida albicans and Staphylococcus epidermidis and the Mechanisms of Action.

Bacterial and fungal coinfections have posed great clinical challenges in recent years, and combination therapy may be a useful way to treat these mixed infections. The objective of this study was to find an effective drug combination to treat dual-species cultures of fungi and bacteria. In this study, we focused on poorly investigated mixed cultures of Candida albicans and Staphylococcus epidermidis. In this research, we investigated the effects of fluconazole (FLC) and doxycycline (DOX) against dual-species cultures of C. albicans and S. epidermidis. Both the fractional inhibitory concentration index model and ΔE model revealed a synergistic antimicrobial effect between FLC and DOX against the four groups of dual-species cultures. Mechanistic studies revealed that the synergism of FLC and DOX against dual-species cultures may be associated with the inhibition of biofilms and calcium dysregulation. Fluconazole+doxycycline appears to be a potential drug combination for the treatment of bacterial and fungal coinfections. These findings are of great significance for overcoming clinical bacterial and fungal coinfections and might provide novel insights into drug discovery for combination therapy.

Interaction of Bdellovibrio bacteriovorus with Gram-Negative and Gram-Positive Bacteria in Dual Species and Polymicrobial Communities.

The interaction of Bdellovibrio bacteriovorus PF13 with mixed bacterial communities, consisting of Gram-negative (Pseudomonas fluorescens and Klebsiella pneumoniae) and Gram-positive (Staphylococcus aureus and Enterococcus faecium) bacteria, was investigated to determine if this wild-type predator preferentially preys on certain bacteria and whether the presence of Gram-positive organisms influences its predation efficiency. In co-culture with P. fluorescens and K. pneumoniae, the cell counts (PFU/mL) of PF13 increased by 5.79 and 5.17 logs (48 h), respectively, while in the dual species assay (P. fluorescens, K. pneumoniae and PF13), the cell counts of PF13 increased by 1.95 logs (24 h). Using ethidium monoazide bromide quantitative polymerase chain reaction (EMA-qPCR), the concentration of PF13 increased by 1.25 to 3.62 logs in the co-culture experiments, by 1.41 to 5.05 logs in dual species cultures and by 2.65 logs in a polymicrobial culture. However, PF13 preferentially preyed on K. pneumoniae in the dual species and polymicrobial cultures, highlighting that the presence of Gram-positive bacteria did not affect the predation efficiency of PF13. This is significant as it implies that the predator can be applied in mixed microbial communities to target Gram-negative pathogens which may pose a health risk to patients, consumers or for the treatment of contaminated water.

Metabolic profile of Candida albicans and Candida parapsilosis interactions within dual-species biofilms.

Within the oral cavity, the ability of Candida species to adhere and form biofilms is well-recognized, especially when Candida albicans is considered. Lately, a knowledge gap has been identified regarding dual-species communication of Candida isolates, as a way to increase virulence, with evidences being collected to support the existence of interactions between C. albicans and Candida parapsilosis. The present work evaluated the synergistic effect of the two Candida species, and explored chemical interactions between cells, evaluating secreted extracellular alcohols and their relation with yeasts' growth and matrix composition. A total of four clinical strains of C. albicans and C. parapsilosis species, isolated from single infections of different patients or from co-infections of a same patient, were tested. It was found that dual-species biofilms negatively impacted the growth of C. parapsilosis and their biofilm matrix, in comparison with mono-species biofilms, and had minor effects on the biofilm biomass. Alcohol secretion revealed to be species- and strain-dependent. However, some dual-species cultures produced much higher amounts of some alcohols (E-nerolidol and E, E-Farnesol) than the respective single cultures, which proves the existence of a synergy between species. These results show evidence that interactions between Candida species affect the biofilm matrix, which is a key element of oral biofilms.

Impact of the Peptide WMR-K on Dual-Species Biofilm Candida albicans/Klebsiella pneumoniae and on the Untargeted Metabolomic Profile.

In recent years, the scientific community has focused on the development of new antibiotics to address the difficulties linked to biofilm-forming microorganisms and drug-resistant infections. In this respect, synthetic antimicrobial peptides (AMPs) are particularly regarded for their therapeutic potential against a broad spectrum of pathogens. In this work, the antimicrobial and antibiofilm activities of the peptide WMR-K towards single and dual species cultures of Candida albicans and Klebsiella pneumoniae were investigated. We found minimum inhibitory concentration (MIC) values for WMR-K of 10 µM for K. pneumoniae and of 200 µM for C. albicans. Furthermore, sub-MIC concentrations of peptide showed an in vitro inhibition of biofilm formation of mono and polymicrobial systems and also a good biofilm eradication even if higher concentrations of it are needed. In order to provide additional evidence for the effect of the examined peptide, a study of changes in extracellular metabolites excreted and/or uptaken from the culture medium (metabolomic footprinting) in the poly-microbial association of C. albicans and K. pneumoniae in presence and absence of WMR-K was performed. Comparing to the untreated dual species biofilm culture, the metabolomic profile of the WMR-K treated culture appears significantly altered. The differentially expressed compounds are mainly related to the primary metabolic pathways, including amino acids, trehalose, pyruvic acid, glycerol and vitamin B6.

Developing New Marine Antifouling Surfaces: Learning from Single-Strain Laboratory Tests

The development of antifouling (AF) technology for marine environments is an area of intense research given the severe economic and ecological effects of marine biofouling. Preliminary data from in vitro assays is frequently used to screen the performance of AF coatings. It is intuitive that microbial composition plays a major role in surface colonization. The rationale behind this study is to investigate whether using a mixed population for the in vitro tests yields substantially different results than using single strains during initial screening. A polymeric coating was tested against single- and dual-species cultures of two common microfouler organisms for 49 days. A bacterium (Pseudoaltermonas tunicata) and a cyanobacterium (Cyanobium sp. LEGE 10375) were used in this study. Linear regression analysis revealed that Cyanobium sp. biofilms were significantly associated with a higher number of cells, wet weight, thickness, and biovolume compared to dual-species biofilms. P. tunicata alone had a biofilm growth kinetics similar to dual-species biofilms, although the P. tunicata–Cyanobium sp. mixture developed less dense and thinner biofilms compared to both single-species biofilms. Cyanobium sp. LEGE 10375 biofilms provided the worst-case scenario, i.e., the conditions that caused higher biofilm amounts on the surface material under test. Therefore, it is likely that assessing the AF performance of new coatings using the most stringent conditions may yield more robust results than using a mixed population, as competition between microfouler organisms may reduce the biofilm formation capacity of the consortium.

Dynamics of Biofilm Formation by Salmonella Typhimurium and Beef Processing Plant Bacteria in Mono- and Dual-Species Cultures.

This study aimed to determine the impact of bacteria from a beef plant conveyor belt on the biofilm formation of Salmonella in dual-species cultures. Beef plant isolates (50) including 18 Gram-negative aerobes (GNA), 8 Gram-positive aerobes (GPA), 5 lactic acid bacteria (LAB), 9 Enterobacteriaceae (EB), and 10 generic Escherichia coli (GEC) were included for developing biofilms in mono- and co-culture with S. Typhimurium at 15°C for 6days. Five selected cultures in planktonic form and in biofilms were tested for susceptibility to two commonly used sanitizers (i.e. E-San and Perox-EPlus). In mono-cultures, ≥ 80, 67, 61, 20, and 13% of GEC, EB, GNA, LAB, and GPA, respectively, developed measurable biofilms after 2days, while all co-culture pairings with S. Typhimurium achieved some level of biofilm production. The predominant effect of EB and only effect of GEC strains on the biofilm formation of S. Typhimuriumwas antagonistic, while that of Gram-positive bacteria was synergistic, with the effect being more prominent on day 6. The effect was highly variable for the GNA isolates. Six aerobic isolates that formed moderate/strong biofilms by day 2 greatly boosted the co-culture biofilm formation. Seven Gram-negative bacteria were antagonistic against the biofilm formation of the co-cultures. Both sanitizers completely inactivated the selected planktonic cultures, but were largely ineffective against biofilms. In conclusion, all beef plant isolates assessed formed biofilms when paired with S. Typhimurium. Aerobic biofilm formers may create a more favorable condition for Salmonella biofilm formation, while some beef plant isolates have potential as a biocontrol strategy for Salmonella biofilms.

Influence of Laboratory Culture Media on in vitro Growth, Adhesion, and Biofilm Formation of Pseudomonas aeruginosa and Staphylococcus aureus

Objective: Pseudomonas aeruginosa and Staphylococcus aureus dual-species biofilm infections are notoriously difficult to manage. This study aimed at investigating the influence of four different culture media on the planktonic growth, adhesion, and biofilm formation of P. aeruginosa and S. aureus. Materials and Methods: We monitored four different culture media including Nutrient Broth, Brain Heart Infusion (BHI) broth, Luria-Bertani broth, and RPMI 1640 medium on the planktonic growth, adhesion, and biofilm formation of P. aeruginosa (ATCC 27853) and S. aureus (ATCC 25923) using MTT assay and scanning electron microscopy (SEM). Results: The most robust growth of the mono- and dual-species cultures was noted in BHI broth. On the contrary, RPMI 1640 medium promoted maximal initial adhesion of both the mono- and dual-species, but BHI broth fostered the maximal biofilm growth. SEM images showed profuse extracellular polysaccharide production in biofilms, particularly in coculture, in BHI medium. Conclusion: Our data demonstrate that BHI broth, relative to the other tested media, is the most conducive for in vitro evaluation of biofilm and planktonic growth kinetics of these two pathogens, both in mono- and coculture.

An in vitro model of Fusobacterium nucleatum and Porphyromonas gingivalis in single- and dual-species biofilms.

PurposeThe goal of this study was to develop and validate a standardized in vitro pathogenic biofilm attached onto saliva-coated surfaces.MethodsFusobacterium nucleatum (F. nucleatum) and Porphyromonas gingivalis (P. gingivalis) strains were grown under anaerobic conditions as single species and in dual-species cultures. Initially, the bacterial biomass was evaluated at 24 and 48 hours to determine the optimal timing for the adhesion phase onto saliva-coated polystyrene surfaces. Thereafter, biofilm development was assessed over time by crystal violet staining and scanning electron microscopy.ResultsThe data showed no significant difference in the overall biomass after 48 hours for P. gingivalis in single- and dual-species conditions. After adhesion, P. gingivalis in single- and dual-species biofilms accumulated a substantially higher biomass after 7 days of incubation than after 3 days, but no significant difference was found between 5 and 7 days. Although the biomass of the F. nucleatum biofilm was higher at 3 days, no difference was found at 3, 5, or 7 days of incubation.ConclusionsPolystyrene substrates from well plates work as a standard surface and provide reproducible results for in vitro biofilm models. Our biofilm model could serve as a reference point for studies investigating biofilms on different surfaces.

Wastewater polishing by consortia of Chlorella vulgaris and activated sludge native bacteria

The increase of human population and anthropogenic activities has resulted in an excessive discharge of wastes into water bodies, contributing to nutrients enrichment and consequent degradation of aquatic environments and freshwater resources. The need for cost-effective systems able to reduce nutrients concentration in wastewaters is therefore a major concern. Microalgal-bacterial consortia can be effectively applied in wastewater polishing, since co-cultivation of these microorganisms can promote mutual growth and nutrients removal from wastewaters. In this study, dual-species cultures of the microalga Chlorella vulgaris and a bacterium isolated from a municipal wastewater treatment plant (Enterobacter asburiae, Klebsiella sp. or Raoultella ornithinolytica) were performed. Studying these consortia aimed the evaluation of their potential in nutrients (nitrogen, phosphorus and chemical oxygen demand) removal from a synthetic medium that mimics a secondary-treated effluent (nitrogen, phosphorus and chemical oxygen demand loads of about 45 mg N L−1, 10 mg P L−1 and 300 mg O2 L−1, respectively). The results have shown that the consortia containing E. asburiae and R. ornithinolytica have significantly improved C. vulgaris growth: when in single cultures maximum cell concentration achieved by C. vulgaris was (8.27 ± 0.79) × 106 cells per mL, whereas in these consortia maximum cell concentrations achieved by C. vulgaris were (19.8 ± 2.0) × 106 and (14.4 ± 2.3) × 106 cells per mL, respectively. The three studied consortia have also contributed to higher nutrients removal, since the time required to achieve the limits in discharged effluents (established by European Union legislation) was reduced to at least half of the value determined for the single C. vulgaris culture. Due to the effectiveness demonstrated by the studied consortia, these microalgal-bacterial systems should be considered as a viable approach for application in the tertiary treatment step of wastewater treatment processes.

Biotechnological potential of Synechocystis salina co-cultures with selected microalgae and cyanobacteria: Nutrients removal, biomass and lipid production

Cultivation of microalgae and cyanobacteria has been the focus of several research studies worldwide, due to the huge biotechnological potential of these photosynthetic microorganisms. However, production of these microorganisms is still not economically viable. One possible alternative to improve the economic feasibility of the process is the use of consortia between microalgae and/or cyanobacteria. In this study, Chlorella vulgaris, Pseudokirchneriella subcapitata and Microcystis aeruginosa were co-cultivated with Synechocystis salina to evaluate how dual-species cultures can influence biomass and lipid production and nutrients removal. Results have shown that the three studied consortia achieved higher biomass productivities than the individual cultures. Additionally, nitrogen and phosphorus consumption rates by the consortia provided final concentrations below the values established by European Union legislation for these nutrients. In the case of lipid productivities, higher values were determined when S. salina was co-cultivated with P. subcapitata and M. aeruginosa.