Anthraquinones against Cryptococcus neoformans sensu stricto: antifungal interaction, biofilm inhibition and pathogenicity in the Caenorhabditis elegans model.

Introduction. Cryptococcal biofilms have been associated with persistent infections and antifungal resistance. Therefore, strategies, such as the association of natural compounds and antifungal drugs, have been applied for the prevention of biofilm growth. Moreover, the Caenorhabditis elegans pathogenicity model has been used to investigate the capacity to inhibit the pathogenicity of Cryptococcus neoformans sensu stricto.Hypothesis. Anthraquinones and antifungals are associated with preventing C. neoformans sensu stricto biofilm formation and disrupting these communities. Antraquinones reduced the C. neoformans sensu stricto pathogenicity in the C. elegans model.Aim. This study aimed to evaluate the in vitro interaction between aloe emodin, barbaloin or chrysophanol and itraconazole or amphotericin B against growing and mature biofilms of C. neoformans sensu stricto.Methodology. Compounds and antifungal drugs were added during biofilm formation or after 72 h of growth. Then, the metabolic activity was evaluated by the MTT reduction assay, the biomass by crystal-violet staining and the biofilm morphology by confocal laser scanning microscopy. C. neoformans sensu stricto's pathogenicity was investigated using the nematode C. elegans. Finally, pathogenicity inhibition by aloe emodin, barbarloin and chrysophanol was investigated using this model.Results. Anthraquinone-antifungal combinations affected the development of biofilms with a reduction of over 60 % in metabolic activity and above 50 % in biomass. Aloe emodin and barbaloin increased the anti-biofilm activity of antifungal drugs. Chrysophanol potentiated the effect of itraconazole against C. neoformans sensu stricto biofilms. The C. elegans mortality rate reached 76.7 % after the worms were exposed to C. neoformans sensu stricto for 96 h. Aloe emodin, barbaloin and chrysophanol reduced the C. elegans pathogenicity with mortality rates of 61.12 %, 65 % and 53.34 %, respectively, after the worms were exposed for 96 h to C. neoformans sensu stricto and these compounds at same time.Conclusion. These results highlight the potential activity of anthraquinones to increase the effectiveness of antifungal drugs against cryptococcal biofilms.

Antibiofilm activity of promethazine, deferiprone, and Manuka honey in an ex vivo wound model.

This study evaluated the antibiofilm activity of promethazine, deferiprone, and Manuka honey against Staphylococcus aureus and Pseudomonas aeruginosa in vitro and ex vivo in a wound model on porcine skin. The minimum inhibitory concentrations (MICs) and the effects of the compounds on biofilms were evaluated. Then, counting colony-forming units (CFUs) and confocal microscopy were performed on biofilms cultivated on porcine skin for evaluation of the compounds. For promethazine, MICs ranging from 97.66 to 781.25 µg/ml and minimum biofilm eradication concentration (MBEC) values ranging from 195.31 to 1562.5 µg/ml were found. In addition to reducing the biomass of both species' biofilms. As for deferiprone, the MICs were 512 and >1024 µg/ml, the MBECs were ≥1024 µg/ml, and it reduced the biomass of biofilms. Manuka honey had MICs of 10%-40%, MBECs of 20 to >40% and reduced the biomass of S. aureus biofilms only. Concerning the analyses in the ex vivo model, the compounds reduced (P < .05) CFU counts for both bacterial species, altering the biofilm architecture. The action of the compounds on biofilms in in vitro and ex vivo tests raises the possibility of using them against biofilm-associated wounds. However, further studies are needed to characterize the mechanisms of action and their effectiveness on biofilms in vivo.

Chitosan microparticles loaded with essential oils inhibit duo-biofilms of Candida albicans and Streptococcus mutans.

OBJECTIVE: Oral candidiasis is a common fungal infection that affects the oral mucosa, and happens when Candida albicans interacts with bacteria in the oral microbiota, such as Streptococcus mutans, causing severe early childhood caries. C. albicans and S. mutans mixed biofilms are challenging to treat with conventional antimicrobial therapies, thus, new anti-infective drugs are required. This study aimed to test a drug delivery system based on chitosan microparticles loaded with geranium and lemongrass essential oils to inhibit C. albicans and S. mutans mixed biofilms. METHODOLOGY: Chitosan microparticles loaded with essential oils (CM-EOs) were obtained by spray-drying. Susceptibility of planktonic were performed according CLSI at 4 to 2,048 µg/mL. Mixed biofilms were incubated at 37ºC for 48 h and exposed to CM-EOs at 256 to 4,096 µg/mL. The antimicrobial effect was evaluated using the MTT assay, with biofilm architectural changes analyzed by scanning electron microscopy. RAW 264.7 cell was used to evaluate compound cytotoxicity. RESULTS: CM-EOs had better planktonic activity against C. albicans than S. mutans. All samples reduced the metabolic activity of mixed C. albicans and S. mutans biofilms, with encapsulated oils showing better activity than raw chitosan or oils. The microparticles reduced the biofilm on the slides. The essential oils showed cytotoxic effects against RAW 264.7 cells, but encapsulation into chitosan microparticles decreased their toxicity. CONCLUSION: This study demonstrates that chitosan loaded with essential oils may provide an alternative method for treating diseases caused by C. albicans and S. mutans mixed biofilm, such as dental caries.

ß-Estradiol and progesterone enhance biofilm development and persister cell formation in monospecies and microcosms biofilms derived from vulvovaginal candidiasis.

The present study aimed to: (1) evaluate the influence of the steroid hormones (SH) on biofilm development; (2) investigate the formation of persister cells (PC) in biofilms; and (3) investigate the influence of SH on PC formation. Biofilms were derived from vulvovaginal candidiasis (VVC) samples and evaluated by three models: microcosm biofilms grown in Vaginal Fluid Simulator Medium (MiB-VFSM); monospecies biofilms grown in VFSM (MoB-VFSM) and RPMI media (MoB-RPMI). SH altered cell counting and biomass of biofilms grown in VSFM; MoB-RPMI were negatively affected by SH. SH stimulated the formation of PC in MiB-VFSM but not MoB-VFSM; MoB-RPMI showed a lower number of PC in the presence of SH. The results showed that SH altered the dynamics of biofilm formation and development, depending on the study model. The data suggest the influence of hormones on the physiology of Candida biofilms and reinforce the importance of PC in the pathogenesis of VVC.

Antifungal activity of sustainable histone deacetylase inhibitors against planktonic cells and biofilms of Candida spp. and Cryptococcusneoformans.

The limited therapeutic options for fungal infections and the increased incidence of fungal strains resistant to antifungal drugs, especially Candida spp., require the development of new antifungal drugs and strategies. Histone deacetylase inhibitors (HDACi), like vorinostat, have been studied in cancer treatment and have antifungal effects, acting alone or synergistically with classical antifungals. Here we investigated the antifungal activity of two novel sustainable HDACi (LDT compounds) based on vorinostat structure. Molecular docking simulation studies reveal that LDT compounds can bind to Class-I HDACs of Candida albicans, C. tropicalis, and Cryptococcus neoformans, which showed similar binding mode to vorinostat. LDT compounds showed moderate activity when tested alone against fungi but act synergistically with antifungal azoles against Candida spp. They reduced biofilm formation by more than 50% in C. albicans (4 µg/mL), with the main action in fungal filamentation. Cytotoxicity of the LDT compounds against RAW264.7 cells was evaluated and LDT536 demonstrated cytotoxicity only at the concentration of 200 µmol/L, while LDT537 showed IC50 values of 29.12 µmol/L. Our data indicated that these sustainable and inexpensive HDACi have potential antifungal and antibiofilm activities, with better results than vorinostat, although further studies are necessary to better understand the mechanism against fungal cells.

Fungal infections are neglected diseases that affect more than a billion people worldwide. Some histone deacetylase inhibitors can act against fungal cells. Our data reveal that HDACi LDT536 and LDT537 have potential antibiofilm and antifungal activities.

Promethazine inhibits efflux, enhances antifungal susceptibility and disrupts biofilm structure and functioning in Trichosporon.

Trichosporon spp. are emerging opportunistic fungi associated with invasive infections, especially in patients with haematological malignancies. The present study investigated the in vitro inhibition of efflux pumps by promethazine (PMZ) as a strategy to control T. asahii and T. inkin. Planktonic cells were evaluated for antifungal susceptibility to PMZ, as well as inhibition of efflux. The effect of PMZ was also studied in Trichosporon biofilms. PMZ inhibited T. asahii and T. inkin planktonic cells at concentrations ranging from 32 to 256 µg ml-1. Subinhibitory concentrations of PMZ inhibited efflux activity in Trichosporon. Biofilms were completely eradicated by PMZ. PMZ potentiated the action of antifungals, affected the morphology, changed the amount of carbohydrates and proteins and reduced the amount of persister cells inside biofilms. The results showed indirect evidences of the occurrence of efflux pumps in Trichosporon and opens a perspective for the use of this target in the control of trichosporonosis.

Effect of promethazine on biofilms of gram-positive cocci associated with infectious endocarditis.

This study evaluated the antimicrobial activity of promethazine against Staphylococcus aureus, Staphylococcus epidermidis and Streptococcus mutans and its effect on the antimicrobial susceptibility of biofilms grown in vitro and ex vivo on porcine heart valves. Promethazine was evaluated alone and in combination with vancomycin and oxacillin against Staphylococcus spp. and vancomycin and ceftriaxone against S. mutans in planktonic form and biofilms grown in vitro and ex vivo. Promethazine minimum inhibitory concentration range was 24.4-95.31 µg/mL and minimum biofilm eradication concentration range was 781.25-3.125 µg/mL. Promethazine interacted synergistically with vancomycin, oxacillin and ceftriaxone against biofilms in vitro. Promethazine alone reduced (p < 0.05) the CFU-counts of biofilms grown on heart valves for Staphylococcus spp., but not for S. mutans, and increased (p < 0.05) the activity of vancomycin, oxacillin and ceftriaxone against biofilms of Gram-positive cocci grown ex vivo. These findings bring perspectives for repurposing promethazine as adjuvant in the treatment of infective endocarditis.

Standardization of in vitro dual-species biofilms of Staphylococcus pseudintermedius and Malassezia pachydermatis: a strategy to establish an ex vivo biofilm model.

Ex vivo experiments have been performed aiming at mimicking in vivo environments. The main aim of this research was to standardize in vitro dual-species biofilm formation by Staphylococcus pseudintermedius and Malassezia pachydermatis as a strategy to establish an ex vivo biofilm model. Initially, the in vitro formation of biofilms in co-culture was established, using YPD medium, inoculum turbidity of 0.5 on the McFarland scale and maturation periods of 96 h for M. pachydermatis and 48 h for S. pseudintermedius. Subsequently, biofilms were formed on porcine skin using the same conditions, under which a greater number of cells/ml was observed in in vitro dual-species than in in vitro mono-species biofilms. Furthermore, ex vivo biofilm images demonstrated the formation of a highly structured biofilm with the presence of cocci and yeasts surrounded by the matrix. Thus, these conditions optimized the growth of both microorganisms within biofilms in vitro and ex vivo.

In vitro effect of the iron chelator deferiprone on the antimicrobial susceptibility and biofilms of Burkholderia pseudomallei.

This study evaluated the effect of the iron chelator deferiprone (DFP) on antimicrobial susceptibility and biofilm formation and maintenance by Burkholderia pseudomallei. Planktonic susceptibility to DFP alone and in combination with antibiotics was evaluated by broth microdilution and biofilm metabolic activity was determined with resazurin. DFP minimum inhibitory concentration (MIC) range was 4-64 µg/mL and in combination reduced the MIC for amoxicillin/clavulanate and meropenem. DFP reduced the biomass of biofilms by 21 and 12% at MIC and MIC/2, respectively. As for mature biofilms, DFP reduced the biomass by 47%, 59%, 52% and 30% at 512, 256, 128 and 64 µg/mL, respectively, but did not affect B. pseudomallei biofilm viability nor increased biofilm susceptibility to amoxicillin/clavulanate, meropenem and doxycycline. DFP inhibits planktonic growth and potentiates the effect of ß-lactams against B. pseudomallei in the planktonic state and reduces biofilm formation and the biomass of B. pseudomallei biofilms.

Ex vivo wound model on porcine skin for the evaluation of the antibiofilm activity of polyhexamethylene biguanide and ciprofloxacin.

This study aimed to standardize the use of an ex vivo wound model for the evaluation of compounds with antibiofilm activity. The in vitro susceptibility of Staphylococcus aureus ATCC 29213 and Pseudomonas aeruginosa ATCC 27853 to ciprofloxacin and polyhexamethylene biguanide (PHMB) was evaluated in planktonic and biofilm growth. The effects of ciprofloxacin and PHMB on biofilms grown on porcine skin explants were evaluated by colony-forming unit (CFU) counting and confocal microscopy. Minimum inhibitory concentrations (MICs) against S. aureus and P. aeruginosa were, respectively, 0.5 and 0.25 µg mL-1 for ciprofloxacin, and 0.78 and 6.25 µg mL-1 for PHMB. Minimum biofilm eradication concentrations (MBECs) against S. aureus and P. aeruginosa were, respectively, 2 and 8 µg mL-1 for ciprofloxacin, and 12.5 and >25 µg mL-1 for PHMB. Ciprofloxacin reduced (P < 0.05) log CFU counts of the biofilms grown ex vivo by 3 and 0.96 for S. aureus and P. aeruginosa, respectively, at MBEC, and by 0.58 and 8.12 against S. aureus and P. aeruginosa, respectively, at 2xMBEC. PHMB (100 µg/mL) reduced (P < 0.05) log CFU counts by 0.52 for S. aureus and 0.68 log for P. aeruginosa, leading to an overall decrease (P < 0.05) in biofilm biomass. The proposed methodology to evaluate the susceptibility of biofilms grown ex vivo led to reproducible and reliable results.

Virulence factors of Gram-negative bacteria from free-ranging Amazon river dolphins (Inia geoffrensis).

Freshwater cetaceans play a significant role as sentinel animals, providing important data on animal species and aquatic ecosystem health. They also may serve as potential reservoirs of emerging pathogens and host virulence genes in their microbiota. In this study, we evaluated virulence factors produced by Gram-negative bacteria recovered from individuals belonging to two populations of free-ranging Amazon river dolphins (Inia geoffrensis). A total of 132 isolates recovered from the oral cavity, blowhole, genital opening and rectum of 21 river dolphins, 13 from Negro River and 8 from Tapajós River, Brazil, were evaluated for the production of virulence factors, such as biofilms and exoproducts (proteases, hemolysins and siderophores), in planktonic and biofilm forms. In planktonic form, 81.1% (107/132) of the tested bacteria of free-ranging Amazon river dolphins were able to produce virulence factors, with 44/132 (33.4%), 65/132 (49,2%) and 54/132 (40,9%) positive for protease, hemolysin and siderophore production, respectively. Overall, 57/132 (43.2%) of the isolates produced biofilms and, under this form of growth, 66/132 (50%), 88/132 (66.7%) and 80/132 (60.6%) of the isolates were positive for protease, hemolysin and siderophore production. In general, the isolates showed a higher release of exoproducts in biofilm than in planktonic form (P < 0.001). The present findings show that Amazon river dolphins harbor potentially pathogenic bacteria in their microbiota, highlighting the importance of monitoring the micro-organisms from wild animals, as they may emerge as pathogens for humans and other animals.

Malassezia spp. and Candida spp. from patients with psoriasis exhibit reduced susceptibility to antifungals.

INTRODUCTION: Psoriasis is a chronic inflammatory disease that affects over 125 million people worldwide. Many studies have shown the importance of the microbiome for psoriasis exacerbation. AIM: Explore the fungal load and species composition of cultivable yeasts on the skin of psoriatic patients (PP) and healthy volunteers living in a tropical area and evaluate the susceptibility to antifungals. METHODOLOGY: A cross-sectional study with 61 participants (35 patients and 26 healthy controls) was performed during August 2018 and May 2019. Clinical data were collected from patient interviewing and/or medical records review. Samples were collected by swabbing in up to five anatomic sites. Suggestive yeast colonies were counted and further identified by phenotypical tests, PCR-REA, and/or MALDI-TOF. Susceptibility of Malassezia spp. and Candida spp. to azoles, terbinafine, and amphotericin B was evaluated by broth microdilution. RESULTS: Nearly 50% of the patients had moderate to severe psoriasis, and plaque-type psoriasis was the most common clinical form. Yeast colonies count was significantly more abundant among PP than healthy controls. Malassezia and Candida were the most abundant genus detected in all participants. Higher MIC values for ketoconazole and terbinafine were observed in Malassezia strains obtained from PP. Approximately 42% of Candida isolates from PP showed resistance to itraconazole in contrast to 12.5% of isolates from healthy controls. MIC values for fluconazole and amphotericin B were significantly different among Candida isolates from PP and healthy individuals. CONCLUSION: This study showed that Malassezia and Candida strains from PP presented higher MIC values to widespread antifungal drugs than healthy individuals.

Chitosan microparticles loaded with essential oils inhibit duo-biofilms of Candida albicans and Streptococcus mutans

Abstract Oral candidiasis is a common fungal infection that affects the oral mucosa, and happens when Candida albicans interacts with bacteria in the oral microbiota, such as Streptococcus mutans, causing severe early childhood caries. C. albicans and S. mutans mixed biofilms are challenging to treat with conventional antimicrobial therapies, thus, new anti-infective drugs are required. Objective This study aimed to test a drug delivery system based on chitosan microparticles loaded with geranium and lemongrass essential oils to inhibit C. albicans and S. mutans mixed biofilms. Methodology Chitosan microparticles loaded with essential oils (CM-EOs) were obtained by spray-drying. Susceptibility of planktonic were performed according CLSI at 4 to 2,048 µg/mL. Mixed biofilms were incubated at 37ºC for 48 h and exposed to CM-EOs at 256 to 4,096 µg/mL. The antimicrobial effect was evaluated using the MTT assay, with biofilm architectural changes analyzed by scanning electron microscopy. RAW 264.7 cell was used to evaluate compound cytotoxicity. Results CM-EOs had better planktonic activity against C. albicans than S. mutans. All samples reduced the metabolic activity of mixed C. albicans and S. mutans biofilms, with encapsulated oils showing better activity than raw chitosan or oils. The microparticles reduced the biofilm on the slides. The essential oils showed cytotoxic effects against RAW 264.7 cells, but encapsulation into chitosan microparticles decreased their toxicity. Conclusion This study demonstrates that chitosan loaded with essential oils may provide an alternative method for treating diseases caused by C. albicans and S. mutans mixed biofilm, such as dental caries.

The Potential of Phenothiazines against Endodontic Pathogens: A Focus on Enterococcus-Candida Dual-Species Biofilm.

Persistent apical periodontitis occurs when the endodontic treatment fails to eradicate the intraradicular infection, and is mainly caused by Gram-positive bacteria and yeasts, such as Enterococcus faecalis and Candida albicans, respectively. Phenothiazines have been described as potential antimicrobials against bacteria and fungi. This study aimed to investigate the antimicrobial potential of promethazine (PMZ) and chlorpromazine (CPZ) against E. faecalis and C. albicans dual-species biofilms. The susceptibility of planktonic cells to phenothiazines, chlorhexidine (CHX) and sodium hypochlorite (NaOCl) was initially analyzed by broth microdilution. Interaction between phenothiazines and CHX was examined by chequerboard assay. The effect of NaOCl, PMZ, CPZ, CHX, PMZ + CHX, and CPZ + CHX on biofilms was investigated by susceptibility assays, biochemical and morphological analyses. Results were evaluated through one-way ANOVA and Tukey's multiple comparison post-test. PMZ, alone or in combination with irrigants, was the most efficient phenothiazine, capable of reducing cell counts, biomass, biovolume, carbohydrate and protein contents of dual-species biofilms. Neither PMZ nor CPZ increased the antimicrobial activity of CHX. Further investigations of the properties of phenothiazines should be performed to encourage their use in endodontic clinical practice.

Enterococcus faecalis and Candida albicans dual-species biofilm: establishment of an in vitro protocol and characterization.

Enterococcus faecalis is the most important agent of persistent apical periodontitis, and recently, Candida albicans has also been implicated in periapical infections. This study aimed to optimize an in vitro E. faecalis and C. albicans dual-species biofilm protocol for endodontic research. Different physicochemical conditions for biofilm formation were tested. Susceptibility assays to antimicrobials, biochemical composition and an ultra-morphological structure analyses were performed. Reproducible dual-species biofilms were established in BHI medium at 35 °C, for 48 h and in a microaerophilic atmosphere. An increase in biomass and chitin content was detected after vancomycin treatment. Structural analysis revealed that the dual-species biofilm was formed by both microorganisms adhered to the substrate. The proposed protocol could be useful for the study of interkingdom relationships and help to find new strategies against periapical infections.

Inhibitory effect of proteinase K against dermatophyte biofilms: an alternative for increasing the antifungal effects of terbinafine and griseofulvin.

This study aimed to evaluate the effect of proteinase K on mature biofilms of dermatophytes, by assays of metabolic activity and biomass. In addition, the proteinase K-terbinafine and proteinase K-griseofulvin interactions against these biofilms were investigated by the checkerboard assay and scanning electron and confocal microscopy. The biofilms exposed to 32 µg ml-1 of proteinase K had lower metabolic activity and biomass, by 39% and 38%, respectively. Drug interactions were synergistic, with proteinase K reducing the minimum inhibitory concentration of antifungals against dermatophyte biofilms at a concentration of 32 µg ml-1 combined with 128-256 µg ml-1 of terbinafine and griseofulvin. Microscopic images showed a reduction in biofilms exposed to proteinase K, proteinase K-terbinafine and proteinase K-griseofulvin combinations. These findings demonstrate that proteinase K has activity against biofilms of dermatophytes, and the interactions of proteinase K with terbinafine and griseofulvin improve the activity of drugs against mature dermatophyte biofilms.

The herbicide paraquat alters growth and melanin production on the Cryptococcus neoformans/Cryptococcus gattii species complex.

Paraquat (1,10-dimethyl-4,4-bipyridinium dichloride; PQ) is a free-radical producing herbicide that affects cell membranes and can upset the environmental balance of microorganisms present in soil, such as Cryptococcus spp. This study aimed to evaluate the in vitro activity of PQ against Cryptococcus spp. in planktonic and biofilm forms, as well as the protective effect of antioxidant agents against the antifungal effect of PQ and the kinetics of melanin production in response to PQ. Susceptibility to PQ was evaluated by microdilution. Cryptococcus sp. strains exposed to PQ were grown in media with ascorbic acid (AA) and glutathione (GSH). Melanin production was assessed in the presence of l-3,4-dihydroxyphenylalanine (l-DOPA) + PQ. The minimum inhibitory concentration of PQ against Cryptococcus spp. ranged from 8 to 256 µg/mL. Furthermore, PQ reduced biofilm formation. AA and GSH restored the fungal growth of Cryptococcus spp. exposed to PQ. In addition, l-DOPA + PQ delayed melanin production by 24 and 48 h for C. deuterogattii and C. neoformans sensu lato, respectively, suggesting that PQ induces a fitness trade-off in melanin production. Taken together, our data suggest that the antifungal effect of PQ against Cryptococcus spp. possibly exerts selective pressures interfering with biofilm formation and melanin production by these yeasts.

Collateral consequences of agricultural fungicides on pathogenic yeasts: A One Health perspective to tackle azole resistance.

Candida and Cryptococcus affect millions of people yearly, being responsible for a wide array of clinical presentations, including life-threatening diseases. Interestingly, most human pathogenic yeasts are not restricted to the clinical setting, as they are also ubiquitous in the environment. Recent studies raise concern regarding the potential impact of agricultural use of azoles on resistance to medical antifungals in yeasts, as previously outlined with Aspergillus fumigatus. Thus, we undertook a narrative review of the literature and provide lines of evidence suggesting that an alternative, environmental route of azole resistance, may develop in pathogenic yeasts, in addition to patient route. However, it warrants sound evidence to support that pathogenic yeasts cross border between plants, animals and humans and that environmental reservoirs may contribute to azole resistance in Candida or other yeasts for humans. As these possibilities could concern public health, we propose a road map for future studies under the One Health perspective.

One Health Implications of Antimicrobial Resistance in Bacteria from Amazon River Dolphins.

Studies on the microbiota of freshwater cetaceans are scarce and may provide important data on animal and environmental health. This study aimed to evaluate the antimicrobial susceptibility of Gram-negative bacteria recovered from two populations of free-ranging Amazon river dolphins (Inia geoffrensis). Twenty-one animals were captured and released, 13 from Negro River and 8 from Tapajós River, Brazil. Swab samples were obtained from the oral cavity, blowhole, genital opening and rectum and were cultured on MacConkey agar. Isolates were biochemically identified, and antimicrobial susceptibility was assessed by disk diffusion method. Overall, 132 isolates were recovered, of which 71 were recovered from animals from Negro River and 61 from Tapajós River. The most commonly recovered bacterial species were Enterobacter cloacae, Morganella morganii, Klebsiella pneumoniae and Pseudomonas aeruginosa. Overall, 51.6% (63/122) of the isolates were not-susceptible (intermediate resistance and resistance), of which 28/122 (22.9%) were resistant to at least one antimicrobial. Cephalothin, cefuroxime and cefepime were the drugs to which more resistant and intermediate results were observed (P < 0.001). The results indicate that free-ranging Amazon river dolphins host resistant bacteria, contributing for their maintenance in the environment. This study highlights the importance of the One Health approach to monitor the emergence of antimicrobial resistance. Summary Gram-negative bacteria recovered from 21 free-ranging Amazon river dolphins (Inia geoffrensis) from the Negro River and the Tapajós River populations were evaluated for their antimicrobial susceptibility. Overall, 51.6% (63/122) of the isolates were not-susceptible (intermediate resistance and resistance), of which 28/122 (22.9%) were resistant to at least one antimicrobial. Cephalothin, cefuroxime and cefepime were the drugs to which more resistant and intermediate results were observed. Thus, free-ranging Amazon river dolphins, never treated with antimicrobials, host resistant bacteria, contributing for their maintenance in the environment and highlighting the importance of the One Health approach to monitor the emergence of antimicrobial resistance.

Anthraquinones from Aloe spp. inhibit Cryptococcus neoformans sensu stricto: effects against growing and mature biofilms.

This study aimed to evaluate the in vitro effect of aloe emodin, barbaloin and chrysophanol on growing and mature biofilms of Cryptococcus neoformans sensu stricto. The compounds were added at the moment of inducing biofilm growth or after growth for 72 h to evaluate their effects on growing and mature biofilms, respectively. Then, biofilm biomass was evaluated by crystal violet staining and metabolic activity by the XTT reduction assay. Morphological alterations were also evaluated by laser scanning confocal microscopy. Aloe emodin and barbaloin affected growing biofilms and disrupted mature biofilms, reducing metabolic activity by > 60% and biomass by > 70%. Chrysophanol only inhibited mature biofilms, but to a lesser extent. In conclusion, anthraquinones, especially aloe emodin and barbaloin, show a relevant effect against growing and mature biofilms of C. neoformans sensu stricto.