Antifungal Activity of ZnO Nanoparticles Synthesized from Eichhornia crassipes Extract for Construction Applications.

Fungal growth on construction materials in tropical climates can degrade aesthetics and manifestations on modern and historical sick buildings, affecting the health of their inhabitants. This study synthesized ZnO nanoparticles with enhanced antifungal properties using a precipitation method. Different concentrations (25%, 50%, and 100%) of Eichhornia crassipes aqueous extract were used with Zn(NO3)2·6H2O as the precursor to evaluate their spectroscopic, morphological, textural, and antifungal properties. X-ray diffraction confirmed the hexagonal wurtzite phase of ZnO with crystallite sizes up to 20 nm. Fourier-transform infrared spectroscopy identified absorption bands at 426, 503, and 567 cm-1 for ZnO-100, ZnO-50, and ZnO-25, respectively. Nitrogen physisorption indicated a type II isotherm with macropores and a fractal dimension coefficient near 2 across all concentrations. Polydispersity index analysis showed that ZnO-50 had a higher PDI, indicating a broader size distribution, while ZnO-25 and ZnO-100 exhibited lower PDI values, reflecting uniform and monodisperse particle sizes. FESEM observations revealed semi-spherical ZnO morphologies prone to agglomeration, particularly in ZnO-25. Antifungal tests highlighted ZnO-25 as the most effective, especially against Phoma sp. with an MFC/MIC ratio of 78 µg/mL. Poisoned plate assays demonstrated over 50% inhibition at 312 µg/mL for all tested fungi, outperforming commercial antifungals. The results indicate that ZnO NPs synthesized using E. crassipes extract effectively inhibit fungal growth on construction materials. This procedure offers a practical approach to improving the durability of building aesthetics and may contribute to reducing the health risks associated with exposure to fungal compounds.

Broad-Spectrum Antifungal, Biosurfactants and Bioemulsifier Activity of Bacillus subtilis subsp. spizizenii-A Potential Biocontrol and Bioremediation Agent in Agriculture.

In this study, the antifungal, biosurfactant and bioemulsifying activity of the lipopeptides produced by the marine bacterium Bacillus subtilis subsp. spizizenii MC6B-22 is presented. The kinetics showed that at 84 h, the highest yield of lipopeptides (556 mg/mL) with antifungal, biosurfactant, bioemulsifying and hemolytic activity was detected, finding a relationship with the sporulation of the bacteria. Based on the hemolytic activity, bio-guided purification methods were used to obtain the lipopeptide. By TLC, HPLC and MALDI-TOF, the mycosubtilin was identified as the main lipopeptide, and it was further confirmed by NRPS gene clusters prediction based on the strain's genome sequence, in addition to other genes related to antimicrobial activity. The lipopeptide showed a broad-spectrum activity against ten phytopathogens of tropical crops at a minimum inhibitory concentration of 400 to 25 µg/mL and with a fungicidal mode of action. In addition, it exhibited that biosurfactant and bioemulsifying activities remain stable over a wide range of salinity and pH and it can emulsify different hydrophobic substrates. These results demonstrate the potential of the MC6B-22 strain as a biocontrol agent for agriculture and its application in bioremediation and other biotechnological fields.

Bacteria Halotolerant from Karst Sinkholes as a Source of Biosurfactants and Bioemulsifiers.

Halotolerant bacteria with biosurfactant (BS) and bioemulsifiers (BE) activity can coexist in Karstic sinkholes with marine influence. Two sinkholes in the Yucatan peninsula were selected to isolate bacteria with BE and BS activity stable in NaCl. The optimal time, the effect of nitrogen and carbon source in the medium, and the conditions (agitation, pH and salinity) for the production of BS and BE compounds in planktonic and sessile (stimulate the formation of biofilms in cell roller) culture were determined. Eighty strains showed the highest emulsification activity (EI24 ≥ 50%) and drop-collapse ≥ 4 mm. 87% of the strains are moderately halotolerant, and 21% bordered the limit of extreme halotolerance. Twenty-four strains maintained or improved their BS and BE activity under salinity conditions at 5% and 10%, being the most active genera Bacillus, Paenibacillus and Lysinibacillus, identified by sequencing of the 16S rRNA gene. The results show that the nitrogen source positively affects the BS and BE activity, regardless of the type of culture. The sessile culture markedly stimulated BS activity with significant differences. However, we did not find a greater influence on the culture conditions. The results suggest that halotolerant bacteria from sinkholes could be implemented in bioremediation and other biotechnological applications.

Expression of ion transport proteins and routine metabolism in juveniles of tropical gar (Atractosteus tropicus) exposed to ammonia.

Tropical gar (Atractosteus tropicus) thrives in aquatic habitats with high levels of total nitrogen (TAN) and unionized ammonia (NH3). However, the tolerance of TAN and NH3, the excretion mechanisms involved, and the effects of these chemicals on routine metabolism are still unknown. Therefore, our objectives were to assess the acute toxicity of TAN and NH3 in A. tropicus juveniles after a 96-h exposure (LC50-96 h) to NH4Cl and after chronic exposure to two concentrations (15% and 30% of LC50-96 h TAN) for 12 days, as well as to evaluate the transcriptional effects associated with Rhesus proteins (rhag, rhbg, rhcg) and ion transporters (NHE, NKA, NKCC, and CFTR) in gills and skin; and to determine the effects of TAN and NH3 on routine metabolism through oxygen consumption (µM g-1 h-1) and gill ventilation frequency (beats min-1). LC50-96 h values were 100.20 ± 11.21 mg/L for TAN and 3.756 ± 0.259 mg/L for NH3. The genes encoding Rhesus proteins and ion transporters in gills and skin showed a differential expression according to TAN concentrations and exposure time. Oxygen consumption on day 12 showed significant differences between treatments with 15% and 30% TAN. Gill ventilation frequency on day 12 was higher in fish exposed to 30% TAN. In conclusion, A. tropicus juveniles are highly tolerant to TAN, showing upregulation of the genes involved in TAN excretion through gills and skin, which affects routine oxygen consumption and energetic cost. These findings are relevant for understanding adaptations in the physiological response of a tropical ancestral air-breathing fish.

Enzymatic Bioprospecting of Fungi Isolated from a Tropical Rainforest in Mexico.

The humid tropical environment provides an ideal place for developing a high diversity of plants; this is why it is an interesting site for the enzymatic bioprospecting of fungi that are responsible for the recycling of organic matter in an efficient and accelerated way and whose enzymes could have multiple biotechnological applications. For this study, 1250 isolates of macroscopic and microscopic fungal morphotypes were collected from soil, leaf litter, and wood. One hundred and fifty strains (50 from each source) were selected for the enzymatic screening. From the first phase, 51 strains with positive activity for laccase, protease, amylase, xylanase, and lipase enzymes were evaluated, of which 20 were isolated from leaf litter, 18 from the soil, and 13 from wood. The 10 best strains were selected for the enzymatic quantification, considering the potency index and the production of at least two enzymes. High laccase activity was detected for Trametes villosa FE35 and Marasmius sp. CE25 (1179 and 710.66 U/mg, respectively), while Daedalea flavida PE47 showed laccase (521.85 U/mg) and protease activities (80.66 U/mg). Fusarium spp. PH79 and FS400 strains had amylase (14.0 U/mg, 49.23 U/mg) and xylanase activities (40.05 U/mg, 36.03 U/mg) respectively. These results confirm the enzymatic potential of fungi that inhabit little-explored tropical rainforests with applications in industry.

Effect of salinity on zinc toxicity (ZnCl2 and ZnO nanomaterials) in the mosquitofish (Gambusia sexradiata).

Zn is an essential trace metal in living beings. However, excessive concentrations can cause toxic effects even in the aquatic biota. Zn is widely used in different industrial sectors, which has increased its presence in aquatic environments. To assess the acute toxicity of Zn, bioassays were performed with the fish Gambusia sexradiata for a 96-h exposure using ZnCl2 (0 and 15 salinity) and ZnO nanomaterials (0 salinity). The mean lethal concentrations (LC50-96 h) for ZnCl2 were 25.36 (19.64-32.76) and 177.91 (129.39-244.63) mg Zn L-1 to 0 and 15 salinity, respectively. The increased concentration of ZnCl2 showed a dose-response relationship; similarly, the increase in salinity significantly reduces the toxicity of Zn. Characterisation of ZnO nanomaterials was carried out by FTIR, DRX, SEM, DLS and zeta potential. The FTIR spectra showed the characteristic band of Zn-O vibration at 364 cm-1, while DRX presents the hexagonal wurtzite structure with an average crystallite size of 40 nm. SEM micrographs reveal rod-like shapes with lengths and diameters of 40-350 nm and 90 nm, respectively. Agglomerates of 423 nm in water suspension were obtained by DLS and zeta potential of + 14.4 mV. Under these conditions, no mortality was observed due to the rapid flocculation/precipitation of ZnO nanomaterials, which involved brief interaction periods of Zn in the water column with the fish. Gambusia sexradiata is affected by increased Zn concentrations in hard water conditions, and salinity changes modified Zn toxicity, placing it as a suitable model for toxicity tests for this type of particles.

Molecular Identification of Selected Streptomyces Strains Isolated from Mexican Tropical Soils and their Anti-Candida Activity.

The increasing incidence of Candida albicans infections and resistance to current antifungal therapies has led to the search for new and more effective antifungal compounds. Actinobacterial species from the Streptomyces genus are recognized as some of the major producers of antimicrobial compounds. Therefore, the aims of this study were: (1) the identification of Streptomyces strains isolated from Mexican tropical acidic soils, (2) the evaluation of their antifungal activity on C. albicans, and (3) the exploration of the presence of polyketide synthase genes in their genome and antifungal secondary metabolites in their extracts. Four actinobacterial strains, isolated from previously unexplored soils with antibacterial antecedents, were selected. These strains were identified as Streptomycesangustmyceticus S6A-03, Streptomyces manipurensis S3A-05 and S3A-09, and Streptomyces parvisporogenes S2A-04, according to their molecular analyses. The ethanol extract of the lyophilized supernatant of S. parvisporogenes displayed the most interesting antifungal activity against C. albicans, with a minimum inhibitory concentration (MIC) of 0.5 mg/mL. Type I polyketide synthase (PKS-I) and non-ribosomal peptide synthase (NRPS) genes were detected in all strains. In addition, type II PKS genes (PKS-II) were also found in S.manipurensis S3A-05 and S. parvisporogenes. LC-UV-HRMS analysis of the active organic extract of S. parvisporogenes indicated the presence of the known antifungal compound carbazomycin G as the major component.

Interactions between abundant fungal species influence the fungal community assemblage on limestone.

The assembly of fungal communities on stone materials is mainly influenced by the differential bioreceptivity of such materials and environmental conditions. However, little is known about the role of fungal interactions in the colonization and establishment of fungal species. We analyzed the effects of intra- and interspecific interactions between 11 species of fungi in oligotrophic and copiotrophic media and on limestone coupons. In a previous study, these species were the most frequently isolated in the epilithic biofilms of limestone walls exposed to a subtropical climate. In the culture media, we found a greater frequency of intra- and interspecific inhibitory effects in the oligotrophic medium than in the copiotrophic medium. On the limestone coupons, all fungi were able to establish; however, the colonization success rate varied significantly. Cladosporium cladosporioides had a less extensive colonization in isolation (control) than in dual interactions (coexistence) with other species. Phoma eupyrena exhibited the highest colonization success rate and competitive dominance among all tested species. X-ray diffraction (XRD) and scanning electron microscope (SEM) analyses revealed that Pestalotiopsis maculans and Paraconiothyrium sp. produced calcium oxalate crystals during their growth on coupon surfaces, both in isolation and in dual interactions. Our results demonstrate that interactions between abundant fungal species influence the fungal colonization on substrates, the biomineralization and the fungal community assemblage growing in limestone biofilms.

Bioweathering Potential of Cultivable Fungi Associated with Semi-Arid Surface Microhabitats of Mayan Buildings.

Soil and rock surfaces support microbial communities involved in mineral weathering processes. Using selective isolation, fungi were obtained from limestone surfaces of Mayan monuments in the semi-arid climate at Yucatan, Mexico. A total of 101 isolates representing 53 different taxa were studied. Common fungi such as Fusarium, Pestalotiopsis, Trichoderma, and Penicillium were associated with surfaces and were, probably derived from airborne spores. In contrast, unusual fungi such as Rosellinia, Annulohypoxylon, and Xylaria were predominantly identified from mycelium particles of biofilm biomass. Simulating oligotrophic conditions, agar amended with CaCO3 was inoculated with fungi to test for carbonate activity. A substantial proportion of fungi, in particular those isolated from mycelium (59%), were capable of solubilizing calcium by means of organic acid release, notably oxalic acid as evidenced by ion chromatography. Contrary to our hypothesis, nutrient level was not a variable influencing the CaCO3 solubilization ability among isolates. Particularly active fungi (Annulohypoxylon stygium, Penicillium oxalicum, and Rosellinia sp.) were selected as models for bioweathering experiments with limestone-containing mesocosms to identify if other mineral phases, in addition to oxalates, were linked to bioweathering processes. Fungal biofilms were seen heavily covering the stone surface, while a biomineralized front was also observed at the stone-biofilm interface, where network of hyphae and mycogenic crystals was observed. X-ray diffraction analysis (XRD) identified calcite as the main phase, along with whewellite and wedellite. In addition, lower levels of citrate were detected by Attenuated Total Reflectance-Fourier-Transform Infrared Spectroscopy (ATR-FTIR). Overall, our results suggest that a diverse fungal community is associated with limestone surfaces insemi-arid climates. A subset of this community is geochemically active, excreting organic acids under quasi-oligotrophic conditions, suggesting that the high metabolic cost of exuding organic acids beneficial under nutrient limitation. Oxalic acid release may deteriorate or stabilize limestone surfaces, depending on microclimatic dynamics.

Emulsifying activity and stability of a non-toxic bioemulsifier synthesized by Microbacterium sp. MC3B-10.

A previously reported bacterial bioemulsifier, here termed microbactan, was further analyzed to characterize its lipid component, molecular weight, ionic character and toxicity, along with its bioemulsifying potential for hydrophobic substrates at a range of temperatures, salinities and pH values. Analyses showed that microbactan is a high molecular weight (700 kDa), non-ionic molecule. Gas chromatography of the lipid fraction revealed the presence of palmitic, stearic, and oleic acids; thus microbactan may be considered a glycolipoprotein. Microbactan emulsified aromatic hydrocarbons and oils to various extents; the highest emulsification index was recorded against motor oil (96%). The stability of the microbactan-motor oil emulsion model reached its highest level (94%) at 50 °C, pH 10 and 3.5% NaCl content. It was not toxic to Artemia salina nauplii. Microbactan is, therefore, a non-toxic and non-ionic bioemulsifier of high molecular weight with affinity for a range of oily substrates. Comparative phylogenetic assessment of the 16S rDNA gene of Microbacterium sp. MC3B-10 with genes derived from other marine Microbacterium species suggested that this genus is well represented in coastal zones. The chemical nature and stability of the bioemulsifier suggest its potential application in bioremediation of marine environments and in cosmetics.

Antifungal coatings based on Ca(OH)2 mixed with ZnO/TiO2 nanomaterials for protection of limestone monuments.

The presence and deteriorating action of microbial biofilms on historic stone buildings have received considerable attention in the past few years. Among microorganisms, fungi are one of the most damaging groups. In the present work, antimicrobial surfaces were prepared using suspensions of Ca(OH)2 particles, mixed with ZnO or TiO2 nanoparticles. The antimicrobial surfaces were evaluated for their antifungal activity both in the dark and under simulated natural photoperiod cycles, using Penicillium oxalicum and Aspergillus niger as model organisms, and two limestone lithotypes commonly used in construction and as materials for the restoration of historic buildings. Both Ca(OH)2-ZnO and Ca(OH)2-TiO2 materials displayed antifungal activity: ZnO-based systems had the best antifungal properties, being effective both in the dark and under illumination. In contrast, TiO2-based coatings showed antifungal activity only under photoperiod conditions. Controls with coatings consisting of only Ca(OH)2 were readily colonized by both fungi. The antifungal activity was monitored by direct observation with microscope, X-ray diffraction (XRD), and scanning electron microscopy (SEM), and was found to be different for the two lithotypes, suggesting that the mineral grain distribution and porosity played a role in the activity. XRD was used to investigate the formation of biominerals as indicator of the fungal attack of the limestone materials, while SEM illustrated the influence of porosity of both the limestone material and the coatings on the fungal penetration into the limestone. The coated nanosystems based on Ca(OH)2-50%ZnO and pure zincite nanoparticulate films have promising performance on low porosity limestone, showing good antifungal properties against P. oxalicum and A. niger under simulated photoperiod conditions.

Antimycobacterial and antileishmanial effects of microfungi isolated from tropical regions in México.

A total of 82 fungal extracts were selected and screened against Mycobacterium tuberculosis and promastigotes of Leishmania mexicana strains. Results showed inhibitory activity in 29 % of the fungal strains against at least one of the targets tested. The most significant antituberculosis (antiTB) effects were presented by Cylindrocarpon sp. XH9B, Fusarium sp. TA54, Fusarium XH1Ga, Gliocladium penicillioides TH04 and TH21, Gliocladium sp. TH16, Kutilakesa sp. MR46, and Verticillium sp. TH28 strains (minimal inhibition concentration (MIC) = 1.56-25 µg/ml). Mortality of L. mexicana promastigotes was displayed by only four strains, Fusarium sp. TA50, Fusarium sp. TA54, Verticillium sp. TH28, and the unidentified 2TA2 strain (IC(50) = 14.23-100 µg/ml and IC(100) = 50-100 µg/ml). Seven of these most active strains were defatted and their corresponding fractions evaluated again. The results showed the best antiTB activity in Gliocladium sp. TH16 (MIC = 1.56 µg/ml) and the highest leishmanicidal potential in Fusarium sp. TA54 (IC(50) = 6.36 µg/ml). These results show that fungi living in the tropical regions of México have the ability to produce bioactive metabolites that could be used in the near future as natural products to control neglected tropical diseases.

Succession of fungi colonizing porous and compact limestone exposed to subtropical environments.

Little is known about the dynamics of succession of fungi on limestone exposed in subtropical environments. In this study, the colonization of experimental blocks of compact and porous limestone by a fungal community derived from natural biofilms occurring on Structure X from the archaeological site of Becán (México), was studied using a cultivation-dependent approach after short-term (9 m) exposure in order to provide a preliminary insight of the colonization process under seminatural conditions. Microbial growth seen as the change of colour of stone surfaces to black/dark green was more abundant on the porous limestone. There was a fairly clear difference in microbial colonization between the onset of the experiment and the 6th month for both limestone types, but no significant increase in the colonization of coupons occurred between months 6 and 9. This could be related to the low rainfall expected for this period, corresponding to the dry season. A total of 977 isolates were obtained. From these, 138 sterile fungi were unidentified, 380 could only be assigned to the order Sphaeropsidales; the remaining isolates (459) were grouped into 27 genera and 99 different species. Nearly all detected fungal species belonged to the Ascomycota (90 %). Rare taxa (species represented by one to three isolates) included the recently described genus Elasticomyces, several species of genera Hyalodendron, Monodyctis, Papulospora, Curvularia, and Septoria. Other taxa were Minimedusa and Gliomastix luzulae, which have not been previously described for stone environments. Abundant fungi included several species of the common genera Cladosporium, Alternaria, and Taeniolella typical for a range of habitats. Succession of populations was observed for certain taxa, this shift in the composition of fungal communities was more evident in porous limestone. After 6 m of exposure, species of the genera Scolecobasidium, Hyalodendron, and Taeniolella were predominant, while after 9 m, the predominant species belonged to the genera Curvularia and Alternaria, particularly on porous stone. These results suggest that Curvularia and Alternaria replaced other fungi, due to a higher tolerance towards low levels of available water during the dry season. Higher levels of water within the porous stone, keep longer periods of microbial activity, minimizing the impact of desiccation. This study contributes to understand the diversity of fungal communities in stone surfaces in subtropical settings and the dynamics of colonization on limestone.

Valuable processes and products from marine intertidal microbial communities.

Microbial communities are ubiquitous in marine intertidal environments. These communities, which grow preferentially as biofilms on natural and artificial surfaces, carry out key processes contributing to the functioning of coastal environments and providing valuable services to human society, including carbon cycling, primary productivity, trophic linkage, and transfer and removal of pollutants. In addition, their surface-associated life style greatly influences the integrity and performance of marine infrastructure and archaeological heritage materials. The fluctuating conditions of the intertidal zone make it an extreme environment to which intertidal biofilm organisms must adapt at varying levels. This requirement has probably favored the development and spread of specific microorganisms with particular physiological and metabolic processes. These organisms may have potential biotechnological utility, in that they may provide novel secondary metabolites, biopolymers, lipids, and enzymes and even processes for the production of energy in a sustainable manner.