A simple culture medium for phenotypic characterization and long-term storage of medically relevant fusarioid fungi.

Fusarioid fungi, particularly Neocosmospora solani and Fusarium oxysporum, are emerging as significant human pathogens, causing infections ranging from localized mycoses to life-threatening systemic diseases. Accurate identification and preservation of these fungi in clinical laboratories remain challenging because of their diverse morphologies and specific growth requirements. This study evaluated a novel milk-honey and malt agar (MHM) against conventional media for cultivating and preserving 60 clinical fusarioid isolates, including Neocosmospora spp. (n = 47), Bisifusarium spp. (n = 5), and Fusarium spp. (n = 8). Compared with Sabouraud dextrose 2 % agar (SDA) and malt extract agar (ME2), MHM significantly increased conidia production (p < 0.0001, mean = 3.4 × 103, standard deviation (SD) = ±1.3 × 103), with results similar to those of carnation leaf agar (CLA). MHM facilitated superior preservation of fusarioid viability for up to one year at room temperature on slant cultures and over two years on swabs in Amies gel with charcoal, outperforming current methods such as Castellani (water) or cryopreservation. Morphological characterization of fusarioid fungi grown on MHM revealed distinct growth patterns and conidial structures for Neocosmospora, Bisifusarium, and Fusarium species, aiding in identifying these genera. The superior performance of MHM in stimulating conidiation, maintaining viability, and preserving morphology underscore its potential as a reference medium for medically relevant fusarioid fungi, with broad implications for clinical mycology laboratories and resource-limited settings.

Development of quaternized agar-based materials for the coronavirus inactivation.

The COVID-19 pandemic has revealed weaknesses in healthcare systems and underscored the need for advanced antimicrobial materials. This study investigates the quaternization of agar, a seaweed-derived polysaccharide, and the development of electrospun membranes for air filtration in facemasks and biomedical applications. Using the betacoronavirus MHV-3 as a model, quaternized agar and membranes achieved a 90-99.99 % reduction in viral load, without associated cytotoxicity. The quaternization process reduced the viscosity of the solution from 1.19 ± 0.005 to 0.64 ± 0.005 Pa.s and consequently the electrospun fiber diameter ranged from 360 to 185 nm. Membranes synthesized based on polyvinyl alcohol and thermally cross-linked with citric acid exhibited lower water permeability. Avoiding organic solvents in the electrospinning technique ensured eco-friendly production. This approach offers a promising way to develop biocompatible and functional materials for healthcare and environmental applications.

Unusual plant-extract based media for the differentiation between species in the Candida albicans complex: A comparative study.

Candida albicans is the most common agent in human fungal infections; nevertheless, in the last decades, the closely related yeasts Candida dubliniensis and Candida africana have emerged as pathogens. The purpose of this study was to compare tobacco agar with another five agars prepared from plant extracts (Origanum vulgare, Rosmarinus officinalis, Solanum rudepannum, Solanum oblongifolium and Brugmansia arborea) on the differentiation of C. albicans complex. The hyphae and chlamyconidia formation and the color and margin of the colonies of 200 clinical isolates of C. albicans, C. dubliniensis and C. africana were evaluated. After seven days of incubation at 28 °C, Tobacco agar, S. rudepannum and B. arborea agars allowed the differentiation of 100 % C. dubliniensis. Additionally, 24% of C. africana isolates produced brownish colonies in the medium prepared from Rosmarinus officinalis (rosemary) extract. These results indicate that S. rudepannun, B. arborea and rosemary agar could be used as screening for the phenotypic differentiation between the species of C. albicans complex. Rosemary agar could be used to aid in the differentiation of C. albicans from C. africana. These culture media based on plants, could be used as simple and inexpensive screening methods in the phenotypic differentiation of C. dubliniensis and C. africana.

Modeling the growth of Aspergillus brasiliensis affected by a nonthermal plasma.

AIM: The main objective of the study was to develop and validate a model for the growth of Aspergillus brasiliensis on surfaces, specifically on agar culture medium. An additional aim was to determine conditions for complete growth inhibition of this micromycete using two different nonthermal plasma (NTP) sources. METHODS AND RESULTS: The developed model uses two key parameters, namely the growth rate and growth delay, which depend on the cultivation temperature and the amount of inoculum. These parameters well describe the growth of A. brasiliensis and the effect of NTP on it. For complete fungus inactivation, a single 10-minute exposure to a diffuse coplanar surface barrier discharge was sufficient, while a point-to-ring corona discharge required several repeated 10-minute exposures at 24-h intervals. CONCLUSIONS: The article presents a model for simulating the surface growth of A. brasiliensis and evaluates the effectiveness of two NTP sources in deactivating fungi on agar media.

Identification and antimicrobial susceptibility profile of bacteria isolated from primary endodontic infections.

This study aimed to identify and characterize the antimicrobial susceptibility profile of bacteria found in primary endodontic infections in the teeth of patients treated at the Dental Clinic of the University of Ribeirão Preto, São Paulo, Brazil. From September to December 2019, samples were obtained from 21 patients with primary endodontic infections. The collections were carried out in triplicate using paper cones placed close to the total length of the root canal. Bacterial isolation was performed in Brain Heart Infusion agar, Blood agar, and other selective culture media cultured at 37°C for up to 48 h under aerobiosis and microaerophilic conditions. The bacterial species were identified using the Vitek 2 automated system. The disk diffusion method on agar Müeller-Hinton was used to assess antimicrobial susceptibility with the recommended antimicrobials for each identified bacterial species. A total of 49 antibiotics were evaluated. Fifteen of the 21 samples collected showed bacterial growth, and 17 bacterial isolates were found. There were 10 different bacterial species identified: Enterococcus faecalis (four isolates), Streptococcus mitis/oralis (three isolates), Streptococcus anginosus (three isolates) being the most common, followed by Staphylococcus epidermidis, Enterococcus faecium, Streptococcus constellatus, Streptococcus alactolyticus, Enterobacter cloacae, Klebsiella variicola, and Providencia rettgeri (one isolate of each species). The analysis demonstrated significant susceptibility to most of the tested antibiotics. However, some Enterococcus isolates resisted the antibiotic's erythromycin, ciprofloxacin, and tetracycline. A Staphylococcus epidermidis isolate was characterized as multidrug-resistant. Five Streptococcus isolates were non-susceptible to all antibiotics tested.

Performance evaluation of Trichoderma reseei in tolerance and biodegradation of diuron herbicide in agar plate, liquid culture and solid-state fermentation.

The present study evaluated the performance of the fungus Trichoderma reesei to tolerate and biodegrade the herbicide diuron in its agrochemical presentation in agar plates, liquid culture, and solid-state fermentation. The tolerance of T. reesei to diuron was characterized through a non-competitive inhibition model of the fungal radial growth on the PDA agar plate and growth in liquid culture with glucose and ammonium nitrate, showing a higher tolerance to diuron on the PDA agar plate (inhibition constant 98.63 mg L-1) than in liquid culture (inhibition constant 39.4 mg L-1). Diuron biodegradation by T. reesei was characterized through model inhibition by the substrate on agar plate and liquid culture. In liquid culture, the fungus biotransformed diuron into 3,4-dichloroaniline using the amide group from the diuron structure as a carbon and nitrogen source, yielding 0.154 mg of biomass per mg of diuron. A mixture of barley straw and agrolite was used as the support and substrate for solid-state fermentation. The diuron removal percentage in solid-state fermentation was fitted by non-multiple linear regression to a parabolic surface response model and reached the higher removal (97.26%) with a specific aeration rate of 1.0 vkgm and inoculum of 2.6 × 108 spores g-1. The diuron removal in solid-state fermentation by sorption on barley straw and agrolite was discarded compared to the removal magnitude of the biosorption and biodegradation mechanisms of Trichoderma reesei. The findings in this work about the tolerance and capability of Trichoderma reesei to remove diuron in liquid and solid culture media demonstrate the potential of the fungus to be implemented in bioremediation technologies of herbicide-polluted sites.

Agar dehydration: a simple method for long-term storage of Acanthamoeba spp. collection at room temperature.

This study describes dehydration of agar containing cysts as a novel and inexpensive method for long-term storage of Acanthamoeba spp. collections at room temperature. Five hundred microliters of axenically cultured Acanthamoeba spp. trophozoites (106 cells/mL) in PYG media or 150 µl of amoeba suspension (106 cells or cysts/mL) from monoxenic plate culture was spread onto the surface of non-nutritive agar (NNA, 2-3-mm thick) without or with a layer of heat-inactivated Escherichia coli, respectively. The plates were sealed and incubated at 30 °C. After the encystment, the Parafilm® was removed, and the plates were kept at the same temperature until the NNA was completely dehydrated. The dehydrated cyst-containing NNA was cut in rectangles and stored in airtight tubes at room temperature for up to 3 years. Cyst viability was assessed by inoculating them in fresh NNA with a layer of E. coli and in PYG followed by incubation at 30 °C. One hundred percent of samples from all specimens (19) stored over the 3 years allowed new cultures to be re-established; however, two strains showed reduced viability, at 66.7% and 62.5%, after 2 years of room temperature storage. One hundred percent of the cyst samples produced axenically and maintained in dry NNA allowed the re-establishment of axenic cultures through direct incubation in PYG, with excystment occurring within 24 or 48 h. For the first time, we report the dehydration of cyst-containing agar as an economical and effective method for the long-term storage of Acanthamoeba spp. collections at room temperature. It enables the creation of large collections using reduced space and economical transport of Acanthamoeba strains, in addition to allowing better organization of the collection.

Anti-biofilm effectiveness of protocols for cleaning complete dentures in hospitalized patients: a randomized controlled trial.

BACKGROUND: Denture biofilm acts as a potential reservoir for respiratory pathogens, considerably increasing the risk of lung infections, specifically aspiration pneumonia, mainly 48h after hospital admission. The establishment of a straightforward, affordable, and applicable hygiene protocol in a hospital environment for the effective control of denture biofilm can be particularly useful to prevent respiratory infections or reduce the course of established lung disease. OBJECTIVES: To evaluate the anti-biofilm effectiveness of denture cleaning protocols in hospitalized patients. METHODOLOGY: The maxillary complete dentures (MCDs) of 340 hospitalized participants were randomly cleaned once using one of the following 17 protocols (n=20): brushing with distilled water, toothpaste, or neutral liquid soap (controls); immersion in chemical solutions (1% sodium hypochlorite, alkaline peroxide, 0.12% or 2% chlorhexidine digluconate), or microwave irradiation (650 W for 3 min) combined or not with brushing. Before and after the application of the protocols, the biofilm of the intaglio surface of the MCDs was evaluated using two methods: denture biofilm coverage area (%) and microbiological quantitative cultures on blood agar and Sabouraud Dextrose Agar (CFU/mL). Data were subjected to the Wilcoxon and Kruskal-Wallis tests (α=0.05). RESULTS: All 17 protocols significantly reduced the percentage area of denture biofilm and microbial and fungal load (P<0.05). The highest percentage reductions in the area of denture biofilm were observed for 1% hypochlorite solution with or without brushing and for 2% chlorhexidine solution and microwave irradiation only in association with brushing (P<0.05). The greatest reductions in microbial and fungal load were found for the groups that used solutions of 2% chlorhexidine and 1% hypochlorite and microwave irradiation, regardless of the association with brushing (P<0.05). CONCLUSIONS: A single immersion for 10 min in 1% sodium hypochlorite, even in the absence of brushing, proved to be a straightforward, rapid, low-cost, and effective protocol for cleaning the dentures of hospitalized patients.

Micropropagation of Encyclia cordigera (Kunth) Dressler in Ebb-and-Flow Bioreactor.

The use of new technologies for micropropagation such as temporary immersion systems (TISs) is important, because it reduces costs by 40% lowering labor, agar and containers. TISs are containers designed for large-scale, semiautomatic production of plants in a liquid medium, which has been used in propagation of commercial orchids. This tool has high potential for application in micropropagation of medicinal and endangered orchids for conservation and commercial purposes. In this chapter, we describe a detailed protocol for propagation and development of Encyclia cordigera to be used in research projects for small-scale production. This protocol comprises all steps from explant preparation to the establishment orchids plantlets.

Modifications to the parasitological technique of Rugai increase the diagnostic sensitivity for strongyloidiasis.

Strongyloidiasis is a neglected tropical disease caused mainly by Strongyloides stercoralis, a nematode that can persist for decades in the human host with a very low parasitic burden and without specific symptoms. Hence, it is difficult to diagnose and control. Larval concentration and culture methods with fecal samples show higher sensitivity for the diagnosis of Strongyloides-infected individuals; however, these techniques are not routinely used, primarily due to the challenges associated with processing a substantial volume of fecal samples. In the current study, we comparatively evaluated the sensitivity and applicability of modifications made to the Rugai parasitological method for the diagnosis of strongyloidiasis in fecal samples of experimentally infected rats and in 68 individuals from an urban community close to Maceió, Brazil. The presence and quantity of parasite larvae in the feces were comparatively evaluated using different parasitological techniques. In the experimental model, we demonstrated that the modified Rugai technique (RMOD) allowed for significantly higher recovery of larvae than the original Rugai technique (RO). Moreover, the sediment was cleaner and easier to evaluate using optical microscopy. Compared to other parasitological techniques, such as agar-plate culture (A-PC) and spontaneous sedimentation (SS), the RMOD technique showed higher sensitivity in the detection of larvae in all infected groups and presented comparatively better performance, especially in rats with a low parasite burden. In the human population, among the 68 stool samples evaluated, Strongyloides larvae were detected in the feces of six individuals with an estimated prevalence of 8.82%. However, the performance of each parasitological method was remarkably different. SS identified Strongyloides larvae in only two individuals and A-PC in three, whereas RMOD was able to identify six infected individuals, resulting in sensitivities of 33.3%, 50%, and 100%, respectively. In conclusion, the modifications introduced to the Rugai technique resulted in improved sensitivity for the detection of Strongyloides spp. infections, especially in stool samples with a low parasite burden, in comparison with other routinely used parasitological techniques.

Agreement of methods to assess antimicrobial susceptibility using Escherichia coli isolates as target models.

Antimicrobial susceptibility tests (AST) conducted in vitro offer a range of methods to assess the antimicrobial resistance (AMR) of microorganisms. Escherichia coli, a widely distributed bacterium, is closely linked to the issue of AMR. In this way, the present study aimed to assess the agreement among different in vitro AST methods, including disk diffusion in agar, broth dilution, and agar dilution method. A total of 100 E. coli isolates were analyzed for their resistance levels against six antibiotics: amoxicillin, ceftiofur, ciprofloxacin, chloramphenicol, tetracycline, and sulfamethoxazole + trimethoprim, using the aforementioned AST methods. Standard breakpoint values were employed to classify isolates as resistant, intermediate, or susceptible, and comparisons among the AST methods were conducted by McNemar's test (P < .05). The obtained data demonstrated equivalence among the AST methods, highlighting the reliability of these standardized classical methodologies. This standardization aids in preventing the inappropriate use of antimicrobials and the dissemination of antimicrobial-resistant microorganisms.

Antifungal susceptibility testing following the CLSI M27 document, along with the measurement of MFC/MIC ratio, could be the optimal approach to detect amphotericin B resistance in Clavispora (Candida) lusitaniae. Susceptibility patterns of contemporary isolates of this species.

Antifungal susceptibility testing (AST) is crucial in clinical settings to guide appropriate therapy. Nevertheless, discrepancies between treatment response and some results still persist, particularly in detecting resistance to amphotericin B (AMB) in Clavispora (Candida) lusitaniae. This study aimed to assess the susceptibility patterns of 48 recent isolates of C. lusitaniae to 9 antifungal agents and explore the feasibility of using a CLSI reference-based method to identify AMB resistance. Microdilution techniques revealed a wide range of minimal inhibitory concentration (MIC) values for azole antifungals, while echinocandins and AMB exhibited a narrow range of MIC values, with all strains considered wild-type for the tested polyene and echinocandins. However, when agar diffusion (ellipsometry) was employed for AST, certain strains displayed colonies within the inhibition ellipse, indicating potential resistance. Interestingly, these strains did not respond to AMB treatment and were isolated during AMB treatment (breakthrough). Moreover, the evaluation of AMB minimum fungicidal concentrations (MFCs) indicated that only the strains with colonies inside the ellipse had MFC/MIC ratios ≥ 4, suggesting reduced fungicidal activity. In conclusion, this study confirms the effectiveness of ellipsometry with RPMI-1640 2% glucose agar for detecting AMB resistance in C. lusitaniae. Additionally, the proposed approach of culturing "clear" wells in the microdilution method can aid in uncovering resistant strains. The findings highlight the importance of appropriate AST methods to guide effective treatment strategies for deep-seated candidiasis caused by C. lusitaniae. Further collaborative studies are warranted to validate these findings and improve the detection of AMB clinical resistance.

Morphological diversity of actinobacteria isolated from oil palm compost (Elaeis guineensis).

Composting is a natural process of decomposition of organic matter that occurs by the action of microorganisms such as fungi, bacteria, and actinobacteria. The actinobacteria are present throughout the process due to their resistance to different environmental conditions. They are Gram-positive, filamentous bacteria with a high capacity for producing secondary metabolites of biotechnological importance. Thus, the objective of this work was to isolate and characterize actinobacteria from industrial composting soil of oil palm (Elaeis guineensis) in the municipality of Igarapé-Açu, Pará. Ten samples of the material were collected and seeded on soy tryptone agar, Reasoner's 2A agar, and Columbia agar, using the serial dilution technique. For morphological characterization of the strains, Gram staining and microculture were performed, and for biochemical characterization, the motility, triple sugar iron, Simmons citrate, maltose, phenylalanine, catalase, and DNAse tests were performed. It was observed that compost actinobacteria have a great diversity in morphological and metabolic production, which may be associated with the substrate and cultivation conditions. Therefore, palm oil compost material represents a rich source of bacterial biodiversity, bringing new perspectives for the bioprospecting of actinobacteria of biotechnological importance in little explored environments.

Analysis of fungal microbiota of ambient air in an intensive care unit in Rio Branco, Acre, Western Amazon, Brazil.

Fungi exhibit three adverse effects on human health: inflammatory, allergic and toxic effects, these implications affect mainly immunodepressed patients. The objective of this work was to analyze the fungal microbiota of the ambient air of an Intensive Care Unit. Three collections were carried out in an Intensive Care Unit in the city of Rio Branco, Acre, Western Amazon, Brazil from March to May 2017. 126 Petri dishes were exposed with the culture medium Agar Sabouraud with chloramphenicol and Agar Mycosel, considering the distribution of the 21 air conditioners, split residential model. The plates were incubated for seven days at room temperature and the growth of Colony Forming Units was observed. Colony counting and isolation for the morphological characterization of the granted fungi was performed. After quantification, the concentration of fungi per cubic meters of air (CFU.m-3) was settled. The third collection had a larger number of colony forming units with 48.6%. In the total of the analyzed samples, filamentous fungi (85.5%) and yeasts (14.5%) were isolated. Thirteen genera of fungi were identified, with the most frequent filaments being Cladosporium spp. 33.0%, Aspergillus spp. 30.4% and Penicillium spp. 19.6%, and yeasts Candida spp. 52.6%, Trichosporon spp. 36.9%. The colony-forming unit per cubic meter (CFU.m-3) did not shown any difference between the Cores in the same collection period, however in the 1st and 3rd collection, Core 1 had the highest average. The fungal microbiota of this Unit presented thirteen different genera potentially pathogenic, revealing the need for monitoring microorganisms and prevention actions.

High-throughput bacterial co-encapsulation in microfluidic gel beads for discovery of antibiotic-producing strains.

Bacteria with antagonistic activity inhibit the growth of other bacteria through different mechanisms, including the production of antibiotics. As a result, these microorganisms are a prolific source of such compounds. However, searching for antibiotic-producing strains requires high-throughput techniques due to the vast diversity of microorganisms. Here, we screened and isolated bacteria with antagonistic activity against Escherichia coli expressing the green fluorescent protein (E. coli-GFP). We used microfluidics to co-encapsulate and co-culture single cells from different strains within picoliter gel beads and analyzed them using fluorescence-activated cell sorting (FACS). To test the methodology, we used three bacterial isolates obtained from Mexican maize, which exhibit high, moderate, or no antagonistic activity against E. coli-GFP, as determined previously using agar plate assays. Single cells from each strain were separately co-incubated into gel beads with E. coli-GFP. We monitored the development of the maize bacteria microcolonies and tracked the growth or inhibition of E. coli-GFP using bright-field and fluorescent microscopy. We correlated these images with distinctive light scatter and fluorescence signatures of each incubated bead type using FACS. This analysis enabled us to sort gel beads filled with an antagonistic strain, starting from a mixture of the three different types of maize bacteria and E. coli-GFP. Likewise, culturing the FACS-sorted beads on agar plates confirmed the isolation and recovery of the two antagonistic strains. In addition, enrichment assays demonstrated the methodology's effectiveness in isolating rare antibiotic-producer strains (0.01% abundance) present in a mixture of microorganisms. These results show that associating light side scatter and fluorescent flow cytometry signals with microscopy images provides valuable controls to establish successful high-throughput methods for sorting beads in which microbial interaction assays are performed.

Review of Analytical Methods for Evaluating Azithromycin in the Context of Green Analytical Chemistry.

BACKGROUND: Azithromycin (AZT) is an antimicrobial available in different pharmaceutical forms and many people can have access to this medicine. Therefore, the existence of adequate and reliable analytical methods for evaluating the quality of AZT and AZT-based products is essential. OBJECTIVE/METHODS: The purpose of this review is to discuss the analytical methods for evaluating AZT present in the literature and official compendia in the context of Green Analytical Chemistry (GAC). RESULTS: Among the methods found in the literature for evaluating AZT, the most used method is HPLC (62%) followed by TLC (14%) and the microbiological method by agar diffusion (14%). Even pharmacopoeias recommend the analysis of AZT by HPLC or agar diffusion. Acetonitrile and methanol account for 35% of the most used solvents in the analyses, followed by buffer. CONCLUSION: AZT lacks analytical methods in the context of GAC. Both physical-chemical and microbiological methods can contemplate the environmentally friendly way to analyze AZT and AZT-based products, depending only on the chosen conditions. Ethanol, purified water, acetic acid instead of methanol, acetonitrile, buffer, formic acid in the physical-chemical methods are excellent alternatives. However, in the microbiological method, turbidimetry is a great option instead of agar diffusion.

Arthrobacter vasquezii sp. nov., isolated from a soil sample from Union Glacier, Antarctica.

A Gram-stain-positive, catalase-positive, non-motile bacteria, with a rod-coccus cycle (designated as EH-1B-1T) was isolated from a soil sample from Union Glacier in Ellsworth Mountains, Antarctica. Strain EH-1B-1T had an optimal growth temperature of 28 °C and grew at pH 7-10. The major cellular fatty acids were anteiso-C15 : 0, iso-C15 : 0, C16 : 0 and anteiso-C17 : 0. The G+C content based on the whole genome sequence was 63.1 mol%. Strain EH-1B-1T was most closely related to members of the genus Arthrobacter, namely Arthrobacter subterraneus and Arthrobacter tumbae. The strain grew on tryptic soy agar, Reasoner's 2A agar, lysogeny broth agar and nutrient agar. The average nucleotide identity and digital DNA-DNA hybridization values between strain EH-1B-1T and its closest reference type strains ranged from 78 to 88 % and from 20.9 to 36.3 %, respectively. Based on phenotypic, chemotypic and genotypic evidence, it is proposed that strain EH-1B-1T represents a novel species of Arthrobacter, for which the name Arthrobacter vasquezii sp. nov. is proposed, with strain EH-1B-1T (RGM 3386T=LMG 32961T) as the type strain.

Microbiological evaluation of homemade mayonnaise and self-serve acai sold in Araguaína, Tocantins.

Foodborne diseases are common illnesses caused by the consumption of food contaminated with microorganisms, such as viruses, fungi, bacteria, and protozoa. Every year, 600 million people become ill and 420,000 people die as a result of consuming contaminated food. Therefore, food safety is an important issue. In this study, samples of homemade spiced mayonnaise and self-serve acai sold in the city of Araguaína, Tocantins, Brazil were analyzed for microbiological contaminants. Acai was collected from 10 stores, one sample from each store, and tested for mold, yeast, and coliforms, as well as coliform identification and total and thermotolerant coliform counts. Mayonnaise was collected from 20 snack bars, one sample from each. These samples were inoculated on MacConkey and Salmonella Shigella agar plates, and the plates were analyzed for growth. Salmonella spp. were detected in some Mayonnaise samples, and coliforms were detected in all acai samples; 60% of samples had thermotolerant coliforms, and only 40% were within the limits established by ANVISA. The collected samples of mayonnaise and acai were contaminated with molds and yeasts above the established limit of 103 CFU/g. Thus, the analyzed mayonnaise and acai samples were contaminated and unfit for consumption, demonstrating the importance of hygienic-sanitary measures in food handling.

Biochemical typing and evaluation of pathogenicity in vulvovaginal isolates of Candida albicans complex / Tipificación bioquímica y evaluación de la patogenicidad de aislamientos vulvovaginales del complejo Candida albicans

Introduction: Candida albicans, C. dubliniensis, and C. africana form the Candida albicans complex. Objective: To identify the phenotypic and pathogenic characteristics of isolates of the C. albicans complex preserved in a collection. Materials and methods: Three hundred presumptive strains of the C. albicans complex were evaluated using CHROMagarTM Candida. Germ tube production was determined by three methods, chlamydospores formation was assessed and colonies were characterized in artisanal agars (Rosmarinus officinalis and Nicotiana tabacum). MALDI-TOF was used as the gold standard identification test. To detect pathogenicity factors, we evaluated the hemolytic activity of each isolate and cocultured with Staphylococcus aureus, coagulase enzyme production, and biofilm formation. Results: Out of the 300 isolates, 43.7% produced germ tube in the heart-brain infusion broth and 47% of the isolates produced chlamydospores. In the artisan media, 6% of the isolates produced brown colonies on rosemary agar and 5% did so on tobacco agar. None of the strains hemolyzed the blood agar alone or cocultured with S. aureus. However, 50% of the isolates hemolyzed the potato dextrose agar supplemented with blood. All strains were coagulase producers, and biofilm production was variable. For germ tube production, the human serum method showed the same positivity as the milk broth method. All isolates were identified as C. albicans by MALDI-TOF. Conclusions: The use of proteomics, molecular tests or a combination of methods is required for species identification.

Introducción: Candida albicans, C. dubliniensis y C. africana forman el complejo Candida albicans. Objetivo: Identificar las características fenotípicas y patogénicas de aislamientos del complejo C. albicans conservados en una colección. Materiales y métodos. Se evaluaron 300 aislamientos identificados presuntivamente como del complejo C. albicans, utilizando CHROMagarTM Candida. Se determinó la producción del tubo germinal mediante tres métodos, se evaluó la producción de clamidosporas, se caracterizaron las colonias en agares artesanales (Rosmarinus officinalis y Nicotiana tabacum) y se utilizó MALDI-TOF como prueba de referencia para la identificación. Para detectar factores de patogenicidad, se evaluó la actividad hemolítica de los aislamientos independientes y en cocultivo con Staphylococcus aureus, la producción de enzima coagulasa y la formación de biopelículas. Resultados: El 43,7 % de los aislamientos produjo tubo germinal en caldo de medio infusión de cerebro-corazón y el 47 % generó clamidosporas. En los medios artesanales, en el 6 % de los aislamientos se obtuvieron colonias de color café en agar romero y, en el 5 %, en agar tabaco. Ninguna de las cepas hemolizó el agar sangre comercial (ni en presencia o ausencia de S. aureus), mientras que el 50 % hemolizó el agar papa dextrosa suplementado con sangre. Todos los aislamientos produjeron enzima coagulasa y la producción de biopelículas fue variable. Para la producción de tubo germinal, el método de suero humano mostró igual positividad que el de caldo de leche. Todos los aislamientos fueron identificados como C. albicans por MALDITOF. Conclusiones: Se requieren herramientas de proteómica y pruebas moleculares, o la combinación de métodos, para poder discriminar entre especies.