Dynamics of the cytosol soluble carbohydrates and membrane lipids in response to ambient pH in alkaliphilic and alkalitolerant fungi

Comparative composition of lipids and cytosol soluble carbohydrates at different ambient pH values was studied for two obligately alkaliphilic fungi (Sodiomyces magadii and S. alkalinus) and for two alkalitolerant ones (Acrostalagmus luteoalbus and Chordomyces antarcticus). The differences and common patterns were revealed in responses to pH stress for the fungi with different types of adaptation to ambient pH. While trehalose was one of the major cytosol carbohydrates in alkaliphilic fungi under optimal growth conditions (pH 10.2), pH decrease to 7.0 resulted in doubling its content. In alkalitolerant fungi trehalose was a minor component and its level did not change significantly at different pH. In alkalitolerant fungi, arabitol and mannitol were the major carbohydrate components, with their highest ratio observed under alkaline conditions and the lowest one, under neutral and acidic conditions. In alkaliphiles, significant levels of arabitol were revealed only under alkaline conditions, which indicated importance of trehalose and arabitol for alkaliphily. Decreased pH resulted in the doubling of the proportion of phosphatidic acids among the membrane lipids, which was accompanied by a decrease in the fractions of phosphatidylcholines and sterols. Alkalitolerant fungi also exhibited a decrease in sterol level at decreased pH, but against the background of increased proportion of one of phospholipids. Decreased unsaturation degree in the fatty acids of the major phospholipids was a common response to decreased ambient pH.     

Alkalitolerant micromycetes in acidic and neutral soils of the temperate zone

A wide diversity of micromycetes from various taxonomic groups in acidic and neutral soils is known from the literature data. In the present work, the fungi isolated from these soils and capable of growth at high pH are analyzed. The fungi were isolated from acidic sod-podzol and neutral cultivated soils by plating on alkaline agar (pH 10.0–10.5). Their identification was carried out using morphological, cultural, and molecular genetic criteria. Phylogenetic analysis was performed and the rates of linear growth within a broad pH range (4.0–10.4) were determined. The isolates represented a polyphyletic group of ascomycetes (Sordariomycetes), which included members of Plectosphaerellaceae (5 species) and various families of Hypocreales (4 species). The most common species were Gibellulopsis nigrescens, Acrostalagmus luteoalbus, Chordomyces antarcticum, and Plectosphaerella spp. Investigation of fungal growth at different pH values revealed all isolates to be alkalitolerant, with no alkaliphilic fungi isolated from acidic sod-podzol and neutral cultivated soils. Although the group of isolates was polyphyletic and its members originated from different ecological and trophic niches, most alkalitolerant isolates exhibited common morphological traits with acremonium- and verticillium-like conidial spore formation, abundant slime formation, and a tendency for aggregation of their mycelium in bundles. Our research confirmed the presence of fungi with alkalitolerant adaptation to external pH in the sod-podzolic and cultivated soils of the Moscow region.     

Ecophysiology and polymorphism of the unicellular extremely natronophilic cyanobacterium <Emphasis Type="Italic">Euhalothece</Emphasis> sp. Z-M001 from Lake Magadi

Strain Z-M001 of a unicellular cyanobacterium, assigned by analysis of the 16S rRNA gene sequence to the phylogenetic group of the generic level Euhalothece, was isolated from soda Lake Magadi. It was shown that strain Z-M001, unlike all other known cultured and uncultured organisms of the Euhalothece group, is extremely natronophilic, and it was named accordingly “Euhalothece natronophila”. In its ecophysiological characteristics, it is comparable to extremely alkaliphilic organotrophic natronobacteria, which is essential for soda ecosystems, because cyanobacteria belong to primary producers. E. natronophila exhibits considerable morphological variability depending on the concentration of carbonates in the medium. The polymorphism of “ E. natronophila” is primarily connected to limitation by utilizable forms of carbon.     

Nimesulide inhibits pathogenic fungi: PGE2-dependent mechanisms

Certain non-steroidal anti-inflammatory drugs can inhibit fungal growth, fungal prostaglandin E2 production, and enzyme activation. This study aims to investigate the antifungal effect of nimesulide against pathogenic filamentous fungi and yeast. The experiments detailed below were also designed to investigate whether the action is dependent on E2 fungal prostaglandins. Our data showed that nimesulide exhibited potent antifungal activity, mainly against Trichophyton mentagrophytes (ATCC 9533) and Cryptococcus neoformans with MIC values of 2 and 62 μg/mL, respectively. This drug was also able to inhibit the growth of clinic isolates of filamentous fungi, such as Aspergillus fumigatus, and dermatophytes, such as T. rubrum, T. mentagrophytes, Epidermophyton floccosum, Microsporum canis, and M. gypseum, with MIC values ranging from 112 to 770 μg/mL. Our data also showed that the inhibition of fungal growth by nimesulide was mediated by a mechanism dependent on PGE2, which led to the inhibition of essential fungal enzymes. Thus, we concluded that nimesulide exerts a fungicidal effect against pathogenic filamentous fungi and yeast, involving the inhibition of fungal prostaglandins and fungal enzymes important to the fungal growth and colonization.     

Phylogenetic characterization and morphological and physiological aspects of a novel acidotolerant and halotolerant microalga <Emphasis Type="Italic">Coccomyxa onubensis</Emphasis> sp. nov. (Chlorophyta,Trebouxiophyceae)

The genus Coccomyxa comprises green microalgae, which can be found worldwide in remarkably versatile aquatic and terrestrial ecosystems including symbiotic associations with a number of different hosts. In this study, we describe a new species, Coccomyxa onubensis, based on 18S and ITS ribosomal DNA (rDNA) sequence data. Coccomyxa onubensis was isolated from acidic water, and its ability to adapt to a wide range of acidic and alkaline pH values and to high salinity was analyzed. The long-term adaptation capacity of the microalga to such extreme conditions was evaluated by performing continuous repeated batches at selected salt concentrations and pH values. Adapted cultures of C. onubensis were found to yield high biomass productivities from pH 2.5 to 9, with maximum yields at acidic pH between 2.5 and 4.5. Moreover, C. onubensis was also found to adapt to salinities as high as 0.5 M NaCl, reaching biomass productivities that were similar to those of control cultures. Ultrastructural analysis by transmission electron microscopy of C. onubensis cells adapted to high salinity showed a robust response to hyperosmotic shock. Thus, C. onubensis was found to be acidotolerant and halotolerant. High biomass productivity over a wide range of pH and salinities denotes C. onubensis as an interesting candidate for various biotechnological applications including outdoor biomass production.     

Ecology of <Emphasis Type="Italic">Scedosporium</Emphasis> Species: Present Knowledge and Future Research

The genus Scedosporium, which comprises at least five clinically relevant species, i.e. Scedosporium apiospermum, Scedosporium boydii, Scedosporium aurantiacum, Scedosporium dehoogii and Scedosporium minutisporum, ranks the second among the filamentous fungi colonizing the airways of patients with cystic fibrosis (CF). This colonization of the airways is thought to contribute to the inflammatory reaction leading to a progressive deterioration of the lung function. Additionally, these colonizing fungi may lead to severe disseminated infections in case of lung transplantation. Therefore, considering the low susceptibility of Scedosporium species to all current antifungal drugs, preventive measures should be defined to reduce the risk of exposure to these fungi for non-colonized CF patients. With this in mind, several studies have been conducted to elucidate the ecology of these fungi and to define possible sources of patient contamination. This review will summarize the major outcomes of those studies, including: the clear demonstration that ecological niches of Scedosporium species are strongly impacted by human activities, and the ability of Scedosporium species to degrade aliphatic and aromatic pollutants which supports the high occurrence of these species in contaminated soils and polluted waters and makes them promising candidates for bioremediation purposes. Finally, prospects for future research in this field are proposed.     

A laboratory study of host use by the cuckoo catfish <Emphasis Type="Italic">Synodontis multipunctatus</Emphasis>

The only known non-avian vertebrate obligate brood parasite is the cuckoo catfish (Synodontis multipunctatus), a Lake Tanganyikan endemic. The cuckoo catfish parasitizes Tanganyikan mouthbrooding cichlids, and under captive conditions, will also parasitize cichlids from other Rift Valley lakes. Here we examine the frequency of parasitism by the cuckoo catfish of Ctenochromis horei from Lake Tanganyika and three species from Lake Malawi and the greater Lake Victorian system in a laboratory setting. C. horei was parasitized significantly less (17%) than the allopatric species Haplochromis latifasciatus, Haplochromis nubilus, and Metriaclima estherae (combined parasitism rate of 28%). The lower rates of parasitism in C. horei may be due to differences in the mating ritual, oviposition (e.g., long periods of pseudo-spawning before actual oviposition), and behavioral adaptations (e.g., increased aggression towards the cuckoo catfish). The number of catfish eggs per parasitized brood was similar between C. horei, H. latifasciatus, H. nubilus, and M. estherae. Our results are comparable to findings from the field for C. horei parasitism frequency and number of cuckoo catfish per brood. We also analyzed the parasitism rate of the albino morph of Metriaclima zebra, a domestic strain. Parasitism rates and number of catfish per brood were the highest in the albino morphotype suggesting that the higher levels of parasitism may be related to lower aggressive behavior, lower visual acuity, or captive influence. Cuckoo catfish and mouthbrooding cichlids provide a model system for testing brood parasitism in a laboratory setting.     

Interactions of the manganese hyperaccumulator <Emphasis Type="Italic">Phytolacca americana</Emphasis> L. with soil pH and phosphate

Hyperaccumulators are plants that store exceptionally high concentrations of heavy metals or metalloids in their leaves. Phytolacca americana is one of the few species known to hyperaccumulate manganese (Mn); however, it is a common weedy species and has no specific association with high-Mn soils. Neither the mechanism by which P. americana hyperaccumulates Mn nor the ecological significance of this trait are well understood. It has recently been suggested that P. americana secretes acids into the rhizosphere as a means of acquiring phosphate, which might coincidentally increase Mn uptake. To determine whether P. americana acidifies the surrounding soil, plants were grown in rhizoboxes providing access to living roots. A thin layer of agar containing bromocresol green pH indicator dye was placed on the roots to observe color changes indicating acidification. Comparative studies showed that P. americana acidifies the rhizosphere significantly more than the non-accumulating plant Acalypha rhomboidea. A second experiment studied whether adjustment of soil pH and phosphate affect foliar Mn concentrations of P. americana. Concentrations of Mn in leaves were highest when plants were grown in acidified soils but were significantly lower in soils that were alkaline and/or enriched with phosphate. These results suggest that Mn hyperaccumulation may be a side effect of rhizosphere acidification as a phosphorus-acquisition mechanism, rather than an adaptation in its own right. The findings provide fundamental information about hyperaccumulator physiology and evolution, and may be relevant to attempts to utilize P. americana for phytoremediation.     

Comprehensive analysis of fungal diversity and enzyme activity in <Emphasis Type="Italic">nuruk</Emphasis>, a Korean fermenting starter,for acquiring useful fungi

Nuruk is a fermenting starter that is involved in the production of alcoholic beverages, and has been used in South Korea for a very long time. To analyze the fungal diversity, we collected a total of 59 nuruk samples from several companies and persons in 2013 to 2014, and obtained 364 isolates. All of the single isolated fungi were identified, both morphologically and molecularly, based on the sequences of ribosomal RNA gene [18S, ITS1-5.8S-ITS2, and 26S (D1/D2 region)]. In 46 nuruk samples out of 59 (78%), Saccharomycopsis fibuligera, a dimorphic yeast, was most frequently isolated. Among the filamentous fungi, Aspergillus and Lichtheimia were found in more than 50% of the samples with lower colony forming unit (CFU/g of sample) than those of yeasts. The yeasts S. fibuligera and Wickerhamomyces anomalus were counted with maximum 1.3–1.8 × 10⁸ CFU/g. Among Mucorales fungi, Lichtheimia and Mucor were isolated in much higher numbers than Rhizopus and Rhizomucor. Overall, the home-made nuruks tend to contain more diverse filamentous fungi than the commercial nuruks. To acquire industrially useful filamentous fungi and yeasts, we analyzed the enzyme activities of α-amylase, glucoamylase and acid protease associated with brewing properties for 131 strains. Aspergillus oryzae and S. fibuligera had high α- and glucoamylase activities and most isolates of Lichtheimia ramosa had high acid protease activity. For further applications, 27 fungal strains were chosen based on isolation frequencies from nuruk, and the ability to produce useful enzyme.     

Progress in Definition,Prevention and Treatment of Fungal Infections in Cystic Fibrosis

Cystic fibrosis (CF) is a chronic lethal multi-system condition; however, most of the morbidity and mortality is dependent on the status of the respiratory system. Progressive respiratory decline is mediated by chronic infection and inflammation, punctuated by important acute events known as pulmonary exacerbations which can lead to accelerated decline. The main bacterial species causing infections include Pseudomonas aeruginosa, Staphylococcus aureus, Haemophilus influenzae and Achromobacter xylosoxidans. In addition to bacteria, fungi are detected in a significant number of patients. The impact of fungal colonization of the airways is still not completely elucidated, but an increasing body of evidence suggests an important role for moulds and yeasts. Although fungal infections are rare, fungi can cause severe pneumonia requiring appropriate targeted treatment. The most common fungi in respiratory samples of patients with CF are Aspergillus fumigatus, Aspergillus terreus and Scedosporium species for filamentous fungi, and yeasts such as Candida albicans and Candida glabrata. Therapeutic strategies depend on the detected fungus and the underlying clinical status of the patient. The antifungal therapy can range from a simple monotherapy up to a combination of three different drugs. Treatment course may be indicated in some patients for two weeks and in others for up to six months, and in rare cases even longer. New antifungal drugs have been developed and are being tested in clinical studies offering the hope of therapeutic alternatives to existing drugs. Identifying relevant risk factors and diagnostic criteria for fungal colonization and infection is crucial to enabling an adequate prevention, diagnosis and treatment.     

Phylogenetic analysis of <Emphasis Type="Italic">Plectosphaerella</Emphasis> species based on multi-locus DNA sequences and description of <Emphasis Type="Italic">P. sinensis</Emphasis> sp. nov.

Species in Plectosphaerella are well known as pathogens of several plant species causing fruit, root and collar rot and collapse. In an investigation of endophytic fungi associated with cucurbit plants in China, we isolated 77 strains belonging to the genus Plectosphaerella. To identify the isolated strains, we collected the type or reference strains of all currently accepted species in Plectosphaerella except P. oratosquillae and conducted a phylogenetic analysis. Phylogenetic analysis of the partial 28S rDNA sequences showed that all species in Plectosphaerella were located in one clade of Plectosphaerellaceae. Based on multi-locus phylogenetic analysis of the ITS, CaM, EF1, TUB and morphological characteristics, all species in Plectosphaerella were well separated. Three endophytic strains from stems of Cucurbita moschata, Citrullus lanatus and Cucumis melo from North China were assigned to a new species described as P. sinensis in this paper. The new species differs morphologically from other Plectosphaerella species by irregular chlamydospores, and the dimensions of phialides and conidia. The other endophytic strains from several cucurbit plants were identified as P. cucumerina.     

Effects of shrub encroachment on vertical changes in soil organic carbon in Mongolian grasslands: using a multi-biomarker approach

Background

This study started from typical replant disease symptoms limited to specific foci within three multi-generation orchards showing homogeneous growth.

Methods

A plant growth assay was conducted using soil from symptomatic and asymptomatic tree root zones along planted rows and from strip-rows. Root colonizing fungal communities were investigated, then the study turned to pathogenicity of Cylindrocarpon-like fungi (Dactylonectria and Ilyonectria spp) and their extracellular exudates.

Results

Growth of apple rootstock plantlets in soil from symptomatic trees was significantly lower than in those observed in soil from asymptomatic trees and from strip-rows. Among the main group of endophytic filamentous fungi isolated from roots, Cylindrocapon-like fungi (Dactylonectria torresensis and, to a lesser extent, Ilyonectria robusta), along with binucleate Rhizoctonia spp., prevailed mostly in plantlets grown in soil collected from planted rows. On the other hand, Fusarium spp. prevailed in plantlets grown in soil from the strip-rows. Cylindrocarpon-like fungi was found to be the most negatively correlated with plant growth. As findings of artificial inoculation with main root colonizing fungal species were not in line with what was observed in native soils, a further investigation was performed on secondary metabolites through which Dactylonectria torresensis exerts pathogenicity; this analysis revealed that both phytotoxins (tentoxin, HC toxin and zearalenone) and cytotoxic compounds (rabelomycin and nidulin) may be involved.

Conclusion

Findings suggest that extracellular compounds released by D. torresensis may have contributed to the severe growth reduction associated with replant disease-like symptoms.

     

<Emphasis Type="Italic">Rhodobaca barguzinensis</Emphasis> sp. nov., a new alkaliphilic purple nonsulfur bacterium isolated from a soda lake of the Barguzin Valley (Buryat Republic,Eastern Siberia)

A novel strain, alga-05, of alkaliphilic purple nonsulfur bacteria was isolated from sediments of a small saline (60 g/l) soda lake near Lake Algin (Barguzin Valley, Buryat Republic, Russia). These bacteria contain bacteriochlorophyll a and carotenoids of the alternative spirilloxanthin group with predominating demethylspheroidenone. They are facultative anaerobes; their photosynthetic structures are of the vesicular type and arranged along the cell periphery. Growth of this strain is possible in a salinity range of 5–80 g/l NaCl, with an optimum at 20 g/l NaCl. Best growth occurred at 20–35°C. Analysis of the 16S rRNA gene sequences demonstrated that the studied isolate is closely related to the alkaliphilic purple nonsulfur bacterium Rhodobaca bogoriensis (99% similarity) isolated from soda lakes of the African Rift Zone. According to the results of DNA-DNA hybridization, strain alga-05 has a 52% similarity with the type species of the genus Rhodobaca. On the basis of the obtained genotypic data and some phenotypic properties (dwelling in a hypersaline soda lake of Siberia, moderate halophily, ability to grow at relatively low temperatures, etc.), the isolated strain of purple bacteria was described as a new species of the genus Rhodobaca, Rca. barguzinensis sp. nov.     

Metabolic activity of <Emphasis Type="Italic">Aspergillus niger</Emphasis> and <Emphasis Type="Italic">Fusarium lateritium</Emphasis> induced by ethoxylated oleyl cetyl alcohol and their bioremediation and biotechnological potential

The effect of ethoxylated oleyl cetyl alcohol (Henkel, Serbia) on the growth and metabolic activity of Aspergillus niger and Fusarium lateritium was in the focus of this paper. The fungi were isolated from wastewater of Lepenica River (Kragujevac, Serbia) at a place where municipal wastewater discharged into the river. The fungi were grown in Czapek-Dox liquid nutrient medium without and with addition of 0.5% pollutant. The physico-chemical and biochemical changes of pH, total biomass dry weight, quantity of free and total organic acids, proteolytic activity and quality of carbohydrates were evaluated from 4-th to 19-th day of fungal growth. The capacity of fungi to decrease concentration of pollutant in medium was determined by cobalt thiocyanate method. The pollutant caused an inhibitory effect on biomass dry weight of A. niger and F. lateritium for 8.50 and 30.61%, respectively. Among tested fungi, A. niger had the better biodegradation capacity (83%) than F. lateritium (65%). Alkaline protease activity of A. niger enhanced in the presence of pollutant for 7.6% whereas the enzyme of F. lateritium retained about 62.2% activity. Overall, the obtained results indicate the potential application of tested fungi in wastewater treatment, detergent industry and biotechnology.     

Is bigger better? A possibility for adaptation of <Emphasis Type="Italic">Daphnia</Emphasis> to filamentous cyanobacteria in the face of global warming

Body size is a key determinant of fitness in Daphnia. Bigger size means higher feeding efficiency and reproduction. However, filamentous cyanobacteria have a more detrimental effect on larger daphnids. Predicted global warming is expected to reduce size of Daphnia and facilitate frequent occurrence of cyanobacteria. Therefore, we asked two questions: (i) does elevated temperature cause reduced size of daphnids, and (ii) is temperature-dependent size reduction adaptive in an environment dominated by filamentous cyanobacteria? To address these issues, we obtained 8 clones of the Daphnia longispina complex from artificially heated lakes and 8 clones from control lakes, and exposed them to a favorable food Scenedesmus obliquus and a mixture of S. obliquus and the filamentous cyanobacterium Cylindrospermopsis raciborskii in life table experiments at 16, 20, and 24°C. Individuals from heated lakes attained larger body size than those from control lakes. Moreover, exposure to the filamentous cyanobacterium led to reduced fecundity of Daphnia from the non-heated lakes and did not affect reproduction of Daphnia from the heated lakes. We conclude that Daphnia display some evolutionary adaptations to cope with filamentous cyanobacteria, linked to long-term exposition to elevated temperature. Our results violate broadly accepted assumptions that ectotherms are smaller in warmer environments.     

<Emphasis Type="Italic">Methylophaga muralis</Emphasis> Bur 1, a haloalkaliphilic methylotroph isolated from the Khilganta soda lake (Southern Transbaikalia,Buryat Republic)

A haloalkaliphilic restricted facultative methylotroph, strain Bur 1, which used methanol, methylated amines, and fructose as carbon and energy sources, was isolated from the soda Lake Khilganta (Buryat Republic, Russia). The cells were gram-negative non-spore-forming, motile rods reproducing by binary fission. The organism was aerobic, reduced nitrates to nitrites. Growth occurred at temperatures from 4 to 37°C (optimum at 25–29°C), pH 7.5–10.5 (optimum at 8.5–9.5), and NaCl concentration in the medium from 0.05 to 10.0% NaCl (optimum at 3–4%). Ectoine, glutamate, and sucrose were accumulated as osmoprotectants. Activity of the enzymes of de novo ectoine biosynthesis were detected. The organism utilized C₁ compounds via the KDPG variant of the ribulose monophosphate pathway. The DNA G + C content was 44.67 mol %. Based on the similarity of the 16S rRNA gene sequences (94.7–99.1%) and the results of DNA–DNA hybridization (24–74%) with type strains of the neutrophilic and alkaliphilic Methylophaga species, the isolate was identified as Methylophaga muralis Bur 1 (VKM B-3046 = DSM 103617). The genome of M. muralis Bur 1 contained 2585 protein-encoding genes; 634 proteins with unidentified functions were predicted. Three rRNAs (5S, 16S, and 23S) and 38 tRNAs were identified. Apart from the mxaFJGIRSACKLDEH classical cluster of methanol oxidation genes, the xoxF gene was found. Methylamine was oxidized to formaldehyde by methylamine dehydrogenase and via the N-methylglutamate pathway. Orthologs of type III glutamine synthetases were revealed in the genome. The operons of ectoine and sucrose biosynthesis, ectRABC-ask and sps-spp-fruK-ams, were found. The genomes of M. muralis Bur 1 and M. lonarensis MPL^T, unlike that of M. nitratireducenticrescens JAM1^T, were found to contain the genes encoding the proteins of bicarbonate transport.