Inhibitory Effects of Cysteine and Cysteine Derivatives on Germination of Sporangiospores and Hyphal Growth of Different Zygomycetes

The in vitro antifungal activity of cysteine (d- and l-cysteine) and its four derivatives (l-cysteine-methyl-ester, N-acetyl-cysteine, N-isobutyryl-d-cysteine, and N-isobutyryl-l-cysteine) were investigated on 20 fungal isolates representing 16 genera (Absidia, Actinomucor, Backusella, Gilbertella, Micromucor, Mortierella, Mucor, Mycotypha, Phycomyces, Rhizomucor, Rhizopus, Saksenaea, Syncephalastrum, Thamnostylum, Umbellopsis, and Zygorynchus). The inhibitory potential of different concentrations of these compounds, ranging from 0.625 to 10 mM, were investigated on the germination of sporangiospores as well as on hyphal extension, using broth microdilution method and agar plate test. Treatment with cysteine and its derivatives resulted in a strong inhibition in most studied strains. At 10 mM of compounds, complete blockage of growth was observed for some isolates. Sensitive species exhibited severe changes in colony morphology in the presence of 10 mM l-cysteine, N-acetyl-cysteine, and N-isobutyryl-l-cysteine. Microscopic observations revealed that 10 mM N-acetyl-cysteine induced dramatic modifications in the structural organization of the hyphae. Results suggest that cysteine and its derivatives have a therapeutic potential against fungal infections caused by Zygomycetes species.     

Degradation of long-chain n-alkanes by mucorales

Summary The composition of fatty acids in the lipids ofAbsidia spinosa, Cunninghamella echinulata, andMortierella isabellina was determined after growth on glucose and some n-alkanes. Mycelia metabolizing odd-carbon n-alkanes contained, among others, mono- and polyenoic heptadecanoic acids, that is, C_17:1(9)-, C_{17:2(9, 12)}-, and C_{17:3(6, 9, 12)}-acids. From the amounts of the isolated lipids it can be deduced thatAbsidia andCunninghamella may mainly degrade the alkanes in a terminal oxidation pathway whileMortierella prefers a subterminal way.Dedicated to Professor Dr. L. Acker on the occasion of his 65th birthday     

Acetylsalicylic acid as antifungal in <Emphasis Type="Italic">Eremothecium</Emphasis> and other yeasts

Interesting distribution patterns of acetylsalicylic acid (ASA, aspirin) sensitive 3-hydroxy (OH) oxylipins were previously reported in some representatives of the yeast genus Eremothecium—an important group of plant pathogens. Using immunofluorescence microscopy and 3-OH oxylipin specific antibodies in this study, we were able to map the presence of these compounds also in other Eremothecium species. In Eremothecium cymbalariae, these oxylipins were found to cover mostly the spiky tips of narrowly triangular ascospores while in Eremothecium gossypii, oxylipins covered the whole spindle-shaped ascospore with terminal appendages. The presence of these oxylipins was confirmed by chemical analysis. When ASA, a 3-OH oxylipin inhibitor, was added to these yeasts in increasing concentrations, the sexual stage was found to be the most sensitive. Our results suggest that 3-OH oxylipins, produced by mitochondria through incomplete β-oxidation, are associated with the development of the sexual stages in both yeasts. Strikingly, preliminary studies on yeast growth suggest that yeasts, characterized by mainly an aerobic respiration rather than a fermentative pathway, are more sensitive to ASA than yeasts characterized by both pathways. These data further support the role of mitochondria in sexual as well as asexual reproduction of yeasts and its role to serve as a target for ASA antifungal action.     

Performance of Rhizopus,Rhizomucor, and Mucor in ethanol production from glucose,xylose, and wood hydrolyzates

In searching for ethanol producing microorganisms also capable of fermenting pentoses, nine zygomycetes strains including three strains of Rhizopus oryzae, Mucor corticolous, M. hiemalis, M. indicus, Rhizomucor pusillus, R. miehei, and zygomycete IT were examined. Each strain was cultivated on glucose, xylose or dilute-acid hydrolyzate (DAH) as carbon sources, and the production of ethanol, lactic acid, glycerol, xylitol, and succinic acid were investigated. Great similarities but also conspicuous differences were seen between the species, to some extent linked to the genera. All strains were capable of growing on glucose or xylose as single carbon source. With the exception of the two Rhizomucor strains, all produced ethanol. All the strains produced glycerol as by-product, while Rhizopus and Rhizomucor but not Mucor produced lactic acid in significant amounts. All Mucor and Rhizopus strains and one strain of Rhizomucor produced xylitol in the xylose medium, but no xylitol was detected after growth on DAH. All Mucor and two R. oryzae strains were capable of growing on DAH. Two Mucor species, M. hiemalis and M. indicus showed greater ethanol production than the other strains. The ethanol yields by M. hiemalis on glucose, xylose, and DAH were 0.39, 0.18, and 0.44 g/g, respectively, whereas the corresponding results for M. indicus were 0.39, 0.22, and 0.44 g/g. The strains also rapidly consumed hydroxymethyl furfural present in DAH.     

Production of arachidonic acid by Mortierella fungi

Summary Various Mortierella fungi were assayed for their productivity of arachidonic acid (ARA). Only strains belonging to the subgenus Mortierella accumulated detectable amounts of ARA together with dihomo--linolenic acid. None of the strains belonging to the subgenus Micromucor tested accumulated these C-20 fatty acids, although they produced a C-18 fatty acid, -linolenic enic acid. A soil isolate, M. alpina 1S-4, was found to grow well in a liquid medium containing glucose and yeast extract as carbon and nitrogen sources, respectively. Addition of several natural oils such as olive and soybean oils to the medium increased the accumulation of ARA. Under optimal culture conditions in a 5-1 bench-scale fermentor, the fungus produced 3.6 g/l of ARA in 7 days. On cultivation for 10 days at 28°C in a 2000-1 fermentor, the same fungus produced 22.5 kg/kl mycelia (dry weight) containing 9.9 kg lipids, in which ARA comprised 31.0% of the total fatty acids. On standing the harvested mycelia for a further 6 days, major mycelial fatty acids (i.e. palmitic acid, oleic acid, linoleic acid, etc.) other than ARA rapidly decomposed and the ARA content of the total fatty acids reached nearly 70%.     

Revision of the genus Absidia (Mucorales, Zygomycetes) based on physiological, phylogenetic, and morphological characters; thermotolerant Absidia spp. form a coherent group, Mycocladiaceae fam. nov.

The genus Absidia comprises ubiquitously distributed soil fungi inhabiting different growth temperature optima ranging from 20–42 °C. Some of the mesophilic species are important biotechnologically in the biotransformation of steroids or as producers of rennin-like components, whereas species with higher growth temperature optima are of clinical relevance as opportunistic human pathogens. The aim of this study was to investigate the phylogenetic relationships between these species and to establish a revision of their systematics. For this purpose single and combined genealogies based on distance, MP, ML, and Bayesian analyses of aligned nucleotide sequences of the nuclear-encoded genes for actin (act) and for the 5.8S ribosomal RNA flanked by the ITS regions 1 and 2 (comprising 807 and 828 characters, respectively) of 16 Absidia species were reconstructed. The phylogenetic reconstructions suggest a trichotomy of the Absidia genus consisting of a mesophilic, a fast-growing thermotolerant, and a slowly-growing mycoparasitic Absidia group. The trichotomous phylogenetic grouping is concordant with the morphology of the zygospores, which are zygotes resulting from sexual conjugation between two compatible mating partners. Whereas the mesophilic group comprises the majority of absidiaceaeous species forming sterile hair-like, mycelial appendages on the suspensors of their zygospores, the thermotolerant group is characterised by the formation of smooth-walled zygospores, and the mycoparasitic group, namely Absidia parricida and A. zychae, by Mucor-like rough-walled zygospores. Based on the phylogenetic coherence of mesophilic and thermotolerant Absidia species, we propose that the two groups are separated into two distinct genera, Absidia for the mesophilic Absidia species resembling the Absidiaceae and Mycocladus for the thermotolerant species A. corymbifera, A. blakesleeana and A. hyalospora. Because Mycocladus is physiologically, phylogenetically, and morphologically distinct from the Absidiaceae s. str. we suggest that they are classified as a separate family, Mycocladiaceae fam. nov., which comprises the three species M. corymbifer, M. blakesleeanus and M. hyalospora.     

Survey of soil fungi from the Sudan Gezira

One hundred and twenty sites, from six localities from the Sudan Gezira were examined for soil mycoflora. The lower fungi were mostly represented by mucoraceous genera, such as Rhizopus, Mucor and Cunninghamella. Oomycetes were rare. Ascomycetes, other than the perfect states of Aspergillus, were not common. Only Chaetomium globosum was isolated from the six localities. Monascus, Thermoascus, Neocosmospora and Pyronema were recorded for the first time from Sudanese soils. The majority of the isolates were Hyphomycetes. The bulk of the fungal population were species of Aspergillus, of which A. niger was the most common, followed by A. terreus, A. flavus, A. nidulans and A. fumigatus. Other Aspergilli were sporadic. A. niveus and A. sejunctus were not previously reported from Sudanese soil.     

A Case of Rhinoorbital Mucormycosis in a Leukemic Patient with a Literature Review from Turkey

Mucormycosis (Zygomycosis) is a rare, invasive, opportunistic fungal infection of the paranasal sinuses, caused by a fungus of the order Mucorales. We report a case of rhinoorbital mucormycosis caused by Rhizopus oryzae in an acute lymphoblastic leukemia patient and review the 79 Mucormycosis cases reported in the last decade from Turkey. In our case, the diagnosis was made with endoscopic appearance, computerized tomography of the paranasal sinuses, and culture of the surgical materials. Following aggressive surgical debridement and parenteral amphotericin B therapy, the patient recovered completely. In Turkish literature, rhinocerebral manifestations were the most common form of the mucormycosis (64 cases), followed by pulmonary form (6 cases). The most common risk factor was hematologic malignancies (32 cases) and diabetes mellitus (32 cases), similar to those reported from the rest of the world. The etiologic agents responsible for the review cases were Rhizopus sp., Mucor spp., Rhizomucor spp., Rhizopus oryzae, Mucor circinelloides, and Lichtheimia corymbifera. Although various treatment modalities were used, amphotericin B was the mainstay of therapy. Mortality rate was found to be 49.4% in review cases. It seems that strong clinical suspicion and early diagnosis, along with aggressive antifungal therapy and endoscopic sinus surgery, have great importance for better prognosis in mucormycosis.     

Aspartic proteinases from <Emphasis Type="Italic">Mucor</Emphasis> spp. in cheese manufacturing

Filamentous fungi belonging to the order of Mucorales are well known as producers of aspartic proteinases depicting milk-clotting activity. The biosynthesis level, the biochemical characteristics, and the technological properties of the resulting proteinases are affected by the producer strain and the mode of cultivation. While the milk-clotting enzymes produced by the Rhizomucor spp. have been extensively studied in the past, much less is known on the properties and potential applications of the aspartic proteinases obtained for Mucor spp. Indeed, several Mucor spp. strains have been reported as a potential source of milk-clotting enzymes having unique technological properties. Both submerged fermentation and solid substrate cultivation are proven alternatives for the production of Mucor spp. aspartic proteinases. This review provides an overview on the bioprocessing routes to obtain large amounts of these enzymes, on their structural characteristics as related to their functional properties, and on their industrial applications with focus on cheese manufacturing.     

Ampelomyces quisqualis Ces., ein Parasit auf Mucorineen

Zusammenfassung Ampelomyces quisqualis Ces., ein bisher für Mucorineae noch unbekannter Parasit, wurde in seiner Entwicklung und seinem Befallsbild untersucht. Neun Gattungen der Mucoraceae und vier Gattungen anderer Mucorineen-Familien wurden durch Infektionsversuche auf Agar-Platten (Ausnahme Actinomucor) getestet. Bei allen wurde eine meist starke Schädigung festgestellt. Nur die Sektion Pusilla der Gattung Mortierella war weitgehend resistent. Im einzelnen sei als Folge des Befalls hingewiesen auf das Auftreten homothallischer Zygoten bei Absidia glauca, auf die Bildung von Zwergsporangien bei Phycomyces und auf die Pilaira-artige Ausbildung des Sporangienträgers bei Pilobolus.

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Ampelomyces quisqualis Ces., a Parasite on Mucorineae

Summary Ampelomyces quisqualis Ces., an hitherto unknown parasite on Mucorineae, is described in its development and its growth on the hosts. 9 genera of Mucoraceae and 4 genera of other families of Mucorineae were tested as potential hosts for Ampelomyces quisqualis on malt-agarplates. A violent damage to most species was found. Only the sectio Pusilla of the genus Mortierella was rather resitent. As sequence to the attack homothallic zygospores appeared in Absidia glauca, dwarfish sporangia in Phycomyces, and the sporangiophores of Pilobolus became similar in appearance to the sporangiophores of Pilaira.

     

Metabolism of the Soy Isoflavones Daidzein and Genistein by Fungi Used in the Preparation of Various Fermented Soybean Foods

The ability of fungi used in the preparation of fermented soybean foods to metabolize the soy isoflavones daidzein and genistein was investigated. A total of 21 fungal strains from dou-chi, miso, sake, soy sauce, and sufu were screened. The genera of the tested fungi included Actinomucor, Aspergillus, Candida, Debaryomyces, Monascus, Mucor, Rhizopus, Saccharomyces, and Zygosaccharomyces. The results were that all tested Aspergillus strains from these soybean foods, including five A. oryzae strains, one A. sojae strain, and one A. tamarii strain, metabolized both daidzein and genistein. In contrast, no other tested fungi from the fermented soybean foods metabolized either daidzein or genistein. The metabolites of daidzein and genistein by Aspergillus strains were identified as 8-hydroxydaidzein and 8-hydroxygenistein, respectively, based on their mass, ¹H-, and ¹³C-NMR spectra.     

New insights into the pollen morphology of the genus Gunnera (Gunneraceae)

Pollen grains from ten species of Gunnera, chosen to represent the six different subgenera in the genus, were examined using light and scanning electron microscopy. The aim of the study was to explore characters that have the potential to define different types of pollen within Gunnera and to study the evolution of these characters in light of the phylogeny of the genus. According to our results, there are three main types of pollen in the examined species of Gunnera. Type 1, unique for the South American species G. herteri (subgenus Ostenigunnera), is characterised by an imperfect reticulum with sinuous undulating-creasted muri. A reticulum with equidimensional polygonal lumina is typical for the plesiomorphic type of pollen (type 2) present in subgenera Gunnera, Misandra and Panke. Lastly, pollen grains of subgenera Pseudogunnera and Milligania are characterised by a reticulum with lumina of variable shape and size (type 3). In G. macrophylla (subgenus Pseudogunnera), the lumina in the apocolpia are of a different shape and size from the lumina in the mesocolpia (type 3a), while in G. dentata, G. monoica and G. cordifolia (subgenus Milligania), the lumina are identical in the apocolpium and the mesocolpium (type 3b).

The identification of pollen types will possibly allow the interpretation of the different specimens of Tricolpites reticulatus, the fossil species believed to be allied to the extant Gunnera.

In addition to the revision on the pollen of Gunnera, a brief comparison between the pollen of this genus and its sister group Myrothamnus is reported.     

The role of wing geometric morphometrics in the identification of sandflies within the subgenus Lutzomyia

The Lutzomyia subgenus (Diptera: Psychodidae) includes sibling species with morphologically indistinguishable females. The aims of this study were to analyse variations in the size and shape of wings of species within the Lutzomyia subgenus and to assess whether these analyses might be useful in their identification. Wings (n = 733) of 18 species deposited in Brazilian collections were analysed by geometric morphometrics, using other genera and subgenera as outgroups. Shape variation was summarized in multivariate analyses and differences in wing size among species were tested by analysis of variance. The results showed significant variation in the sizes and shapes of wings of different Lutzomyia species. Two clusters within the Lutzomyia subgenus were distinguished in analyses of both males and females. In Cluster 1 (Lutzomyia ischnacantha, Lutzomyia cavernicola, Lutzomyia almerioi, Lutzomyia forattinii, Lutzomyia renei and Lutzomyia battistinii), scores for correct reclassification were high (females, kappa = 0.91; males, kappa = 0.90), whereas in Cluster 2 (Lutzomyia alencari, Lutzomyia ischyracantha, Lutzomyia cruzi, Lutzomyia longipalpis, Lutzomyia gaminarai and Lutzomyia lichyi), scores for correct reclassification were low (females, kappa = 0.42; males, kappa = 0.48). Wing geometry was useful in the identification of some species of the Lutzomyia subgenus, but did not allow the identification of sibling species such as L. longipalpis and L. cruzi.     

核酸碱基组分与毛霉目的分类

本文对毛霉目中6科16属40个种共60株真菌DNA的G+C含量与分布作了系统的研究。在提取DNA时选用了Storck等人在Marmur提取细菌DNA方法基础上发展起来的真菌DNA提取法,经过一些修改后成功地提取了毛霉目真菌的DNA。经该法提取的DNA片段长、纯度高,在热变性时DNA的增色效应一般都大于35%。各个种的GC含量大致有一固定值。根据所测毛霉目16属各属真菌的平均GC含量,可将它们分为明显的三组:Gongronella, Haplosporangium, Mortierella及Syncephalastrum为一组,GC含量最高为46.0-49.8%; Cunninghamella单独成一组,GC含量最低只有28.8%;其他各属包括Absidia, Mucor, Rhizopus等GC含量介乎这两组之间,分布于34.9-41.9%。这一次序除Helicostylum与Circinella的数值低于他人的报道外,其余和文献报道值一致。一般属内GC含量变化小于10%,种内变化小于2%。所测得的G+C mol%对分Syncephalastrum monosporum Zheng et al.和Mortierella ramanniana (Moeller) Linneman的合理归属提供了佐证。对某些所测结果与文献报道值有出入的原因作了讨论和分析。对采用Mandel等1970年建议的公式GC=(Tm 0.1×SSC/50.2)-0.990来计算GC含量的依据也作了讨论。     

柯拉斯那沉香的生物活性成分研究

为了解柯拉斯那(Aquilaria crassna)的化学成分,从其所产沉香中分离得到10个化合物,经波谱分析分别鉴定为:6,8-羟基-2-(2-苯乙基)色酮(1),6,8-二羟基-2-[2-(4-甲氧基苯)乙基]色酮(2),rel-(1a R,2R,3R,7b S)-1a,2,3,7b-tetrahydro-2,3-dihydroxy-5-(2-phenylethyl)-7H-oxireno[f][1]benzopyran-7-one(3),rel-(1a R,2R,3R,7b S)-1a,2,3,7b-tetrahydro-2,3-dihydroxy-[2-(4-methoxyphenyl)-ethyl]-7H-oxireno[f][1]benzopyran-7-one(4),rel-(1a R,2R,3R,7b S)-1a,2,3,7b-tetrahydro-2,3-dihydroxy-5-[2-(3-hydroxy-4-methoxyphenyl)-ethyl]-7H-oxireno[f][1]benzopyran-7-one(5),oxidoagarochromone B(6),oxidoagarochromone C(7),(5S,6R,7S,8R)-2-[2-(3′-hydroxy-4′-methoxyphenyl)ethyl]-5,6,7,8-tetrahydroxy-5,6,7,8-tetrahydrochromone(8),6,7-cis-dihydroxy-2-(2-phenylethyl)-5,6,7,8-tetrahydrochromone(9),N-trans-feruloyltyramine(10)。化合物3~5和8~10为首次从柯拉斯那沉香中分离得到。化合物1,3,6,7,9和10对乙酰胆碱酯酶具有一定的抑制活性,化合物4对人慢性髓原白血病细胞株K-562和人胃癌细胞株SGC-7901均具有较小的抑制作用,化合物1和3对人肝癌细胞株BEL-7402也有抑制活性。     

A new β-glucosidase gene from the zygomycete fungus Rhizomucor miehei

In this study, a β-glucosidase coding gene (bgl) of the zygomycete fungus Rhizomucor miehei has been cloned and characterized. The gene comprises a total of 2,826 bp including the coding sequence of a 717 amino acids length putative protein and 10 introns dispersed in the whole coding region. The putative N-and C-terminal catalytic domains (aa 68 to aa 274 and aa 358–601, respectively) were identified; the two domains are connected with a 84-amino-acids linker. The catalytic region showed an extensive sequence homology with other fungal β-glucosidases classified as family 3 glycoside hydrolases. The isolated Rhizomucor gene was expressed in the related fungus Mucor circinelloides. Transformant Mucor strains maintained the introduced plasmid in an autoreplicative manner and showed significantly higher cellobiase activity than the recipient strain.     

Biotoxic activity in the Mucorales

The toxigenicity of representatives of 15 species of Mucorales (Absidia glauca, Actinomucor elegans, Cunninghamella elegans, Helicostylum piriforme, Mortierella isabellina, Mortierella (Mucor) rammaniana, Mucor hiemalis, Mucor mucedo, Mucor spinosus, Phycomyces blakesleeanus, Rhizopus oligosporus, Rhizopus stolonifer, Syncephalastrum racemosum, Thamnidium elegans, Zygorhynchus moelleri) towards the larvae of brine shrimp (Artemia salina) and the growth of pea seedlings (Pisum sativum) and tobacco plants (Nicotiana tabacum) was evaluated. The fungi were cultivated on malt extract agar and aqueous solutions of the cultures were tested.Thamnidium elegans showed a marked toxic action towards brine shrimp (mortality: 74.1%) andPhycomyces, Actinomucor andSyncephalastrum were only weakly toxic. Length and weight of stems of pea seedlings were moderately reduced by extracts ofAbsidia, Cunninghamella, Zygorhynchus andThamnidium and to a lesser degree byMucor spinosus. Cunninghamella andMucor spinosus also inhibited the development of pea hypocotyls. The length of tobacco stems was reduced byMortierella ramanniana, Rhizopus stolonifer andCunninghamella elegans. Wilting or other toxic phenomena were never observed with both test plants. Considering the present results and data from literature it is suggested that species of Mucorales have only a weak toxigenicity.     

Synthesis of Annonaceous Acetogenins from Muricatacin

Synthetic studies of annonaceous acetogenins starting from (?)-muricatacin (1a) or (+)-muricatacin are described, involving (?)-muricatacin (1a), mono-THF acetogenin, solamin (2), reticulatacin (3), (15R, 16R, 19S, 20S)-cis-solamin (4a) and (15S, 16S, 19R, 20R)-cis-solamin (4b), non-adjacent bis-THF acetogenin, 4-deoxygigantecin (5), and epoxide-bearing acetogenin, (15S, 16R, 19S, 20R)-diepomuricanin (6a).     

Synthesis of Both the Enantiomers of 3-Octanol,the Pheromone of Various Species of Ants

(2S,3R,1′S,2′S)-Serricorole (1) and (2S,3R,1′R)-serricorone (2), sex pheromone components of the cigarette beetle (Lasioderma serricorne F.), were synthesized, starting from the enantiomers of methyl 3-hydroxypentanoate. The stereochemistry of the naturally occurring 1 was determined to be 2S,3R,1′S,2′S, and that of 2 to be 2S,3R,1′RS by comparing between the CD spectra of the natural and synthetic samples.     

Antibacterial Effect of 2R,3R-dihydromyricetin on the Cellular Functions of Staphylococcus aureus

The C-3-OH, C-4 carbonyl oxygen and hydrogenation of C2=C3 bond on the C-ring of 2R,3R-dihydromyricetin (DMY) proved to be not necessary for the antibacterial activity against Staphylococcus aureus. DMY significantly decreased the intracellular ATP of S. aureus cells but had few effects on pH_in, proline oxidation, succinate dehydrogenase activity or malate dehydrogenase activity.