Production of microbial lipases in a solid state fermentation system

Summary Several filamentous fungi were grown on solid state fermentation systems and selected for their highest yield in lipolytic activity, low protease levels and dairy flavor generation.P. candidum,M. miehei andP. camembertii were the selected ones. A comparison between kinetics in solid and submerged fermentation was performed usingP. candidum, with the solid system giving the highest titers and a stable production.     

Coprophilous fungi of the horse

A total of 1267 microfungi, including 35 Myxomycetes, were recorded from the fecal samples of the 60 horses; of these 395 were found on 20 saddle-horse feces, 363 on 20 race-horses and 509 on 20 working-horses. Eighty two species representing 53 genera were recorded; of these 7 were Zygomycetes, 18 Ascomycetes, 1 Basidiomycetes and 25 Fungi Imperfecti: 2 Myxomycetes. Common coprophilous fungi are in decreasing orderPilobolus kleinii, Saccobolus depauperatus, Mucor hiemalis, Lasiobolus ciliatus, Podospora curvula, Petriella guttulata, M. circinelloides, Coprinus radiatus, Dictyostelium mucoroides, Sordaria fimicola, C. miser, C. stercorarius, Acremonium sp., Coprotus granuliformis, Graphium putredinis, Iodophanus carneus, Chaetomium murorum, Podospora communis, P. inaequalis, P. setosa, Saccobolus versicolor andCladosporium cucumerinum. Species ofMyrothecium verrucaria, Actinomucor elegans, Kernia nitida, Spiculostilbella dendritica andMucorparvispora were found exclusively in working-horses feces.Badhamia sp., Anixiopsis stercoraria, Echinobotryum state ofD. stemonitis, Geotrichum candidum andOidiodendron sp. were found only in saddle-horses feces.Chlamidomyces palmarum andPhilocopra sp. were found exclusively in race-horses feces.Notes on infrequent or interesting fungi includeThamnostylum piriforme, Phialocephala dimorphospora, Rhopalomyces elegans andSpiculostilbella dendritica.     

Revisione del Genere Polyscytalum Riess

Riassunto Si discute la posizione sistematica del generePolyscytalum in relazione ai funghi artrosporei prospettando la necessità di tenerlo distinto daGeotrichum eCylindrium. Sulla scorta di materiali di erbario (tra cui l'autotipo diP. fecundissimum var.macrosporum, si è accertato che i conidi (artrospore) del generePolyscytalum sono unisettati. Il genere appartiene così agli Ifomiceti Ialodidimi e non a quelli Ialoamerosporei. Delle altre specie ristudiate, ilP. sericeum Sacc. (tipo) è una specie del genereFusidium Link (=Cylindrium Bon.) e si propone la combinazioneF. sericeum; ilP. griseum è sinonimo diG. candidum. IlP. sericeum var.conorum Sacc. rientra nel genereFusidium e si propone la nuova combinazioneF. conorum. IlP. chymophilum e ilP. saccardianum sono pure sinonimi diG. candidum. IlP. murinum, isolato in Brasile da un pesce del genereCarassius corrisponde aCylindrium heteronemum Sacc. e pertanto viene proposta la nuova combinazioneFusidium heteronemum (Sacc.)Cif. etCar. In conclusione viene riconvalidato il generePolyscytalum Riess emendandone la diagnosi.

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Summary The taxonomy of the arthrosporic genusPolyscytalum, as related toGeotrichum andCylindrium, is discussed. The study of the authentic material of a variety of the type species (Polyscytalum fecundissimum Riess var.macrosporum Riess) revealed the presence of a central septum, evident only by the phase contrast microscopy. The only species cultivated,P. murinum Oud., is butFusidium heteronemum (Sacc.) (=Cylindrium heteronemum Sacc.), at least in the Brazilian culture from the intestinal content of a fish. In additionP. griseum, P. chymophilum andP. saccardianum are synonyms ofGeotrichum candidum. ForP. sericeum andP. sericeum var.conorum the combinationsF. sericeum (Sacc.) andF. conorum (Sacc.) are proposed.In conclusion the genusPolyscytalum is re-validated, but the diagnosis emended.

     

Some fossil woods from the palaeogene of northern Kyushu,III

Petrified woods obtained from the Oligocene of Tsuyazaki, Fukuoka Prefecture have been studied, and four new species ofPrunus (Rosaceae) are described:Prunus palaeozippeliana, P. ascendentiporulosa, P. uviporulosa andP. polyporulosa. These four species are very similar to each other in gross anatomical features and are characterized by the diffuse porosity, exclusively simple perforation plates, spiral thickenings on vessel walls, heterogeneous rays and the presence of traumatic gum canals. The petrified woods ofPrunus are new records from the Tetiary of Japan.     

Ultrastructural aspects of leaves inFestuca ovina andPoa sphondylodes (C-3 Poaceae)

Structural aspects of the leaves of two common festucoids,Festuca ovina andPoa sphondylodes, have been examined employing the electron microscopy. The nature of vascular bundles and of sheaths that surround vascular tissues was discussed in the study. The festucoids exhibited a non-Kranz C-3 anatomy with more than four mesophyll cells separating the bundle sheaths of a leaf blade. Vascular tissues in theseFestuca andPoa leaves were surrounded by a double sheath: an inner distinct mestome sheath (MST) and an outer indistinctive layer of parenchymatous bundle sheath (PBS) cells. The PBS cells were much larger than the MST and had thin walls. The MST cells were relatively small and rectangular inP. sphondylodes and more or less hexangular in transverse sections ofF. ovina. InP. sphondylodes, MST had conspicuously thickened inner tangential walls with asymmetrically uninterrupted suberized lamellae in radial and tangential walls. In most differentiated MST cells, all walls were highly suberized. During suberin deposition, MST cells were quite vacuolated and most of the cytoplasm was present as a thin peripheral layer. However, MST walls inF. ovina revealed very thin suberized lamellae with translucent striations. No chloroplasts were detected inP. sphondylodes, whereas the MST inF. ovina contained small chloroplasts. Plasmodesmata were well developed in the primary pit fields of walls between MST and vascular cells, and between adjacent MST cells. Plasmodesmata were less frequent in the walls between the inner and outer sheath cells. Suberized lamellae were totally absent from the PBS cell walls in all veins. External to the PBS, the mesophyll comprised thin walled cells with abundant intercellular spaces. Peripherally arranged chloroplasts in the mesophyll were numerous and often larger than those of PBS and MST cells. Characteristics associated with C-3 and other ultrastructural features were also discussed in the study.     

尾叶桉林下5种植物的耐阴性、生物量及其个体消长

为了解尾叶桉林下不同植物种类的生理生态特性和生长适应性,在鹤山站尾叶桉林中选取分布较多且药用价值和经济价值较高的5种林下植物为研究对象,测定各植物的叶片特征、叶绿素含量和光合-光响应曲线,同时结合2006年和2011年尾叶桉林的样方调查数据分析了5种林下植物生物量的变化和个体数量的消长。结果表明:(1)野牡丹叶片的宽度、厚度、单片叶面积等指标最大,梅叶冬青最小,桃金娘、栀子和山苍子处于两者之间;桃金娘、野牡丹和山苍子的光合速率随光照的增强迅速增大,其光饱和点较高;梅叶冬青和栀子的光合速率也随光照的增强而增大,但其斜率比野牡丹和山苍子大,并很快就达到光饱和点。综合聚类分析植物的耐阴程度,5种植物可分为2类:第1类为耐阴性较强的梅叶冬青、栀子,第2类为耐阴性较弱的野牡丹、山苍子和桃金娘。(2)随着尾叶桉林的生长,5种林下植物的生物量都有不同程度的增加,其中增幅较大的是野牡丹和桃金娘分别增加74倍和18倍,增幅最小的是栀子仅1.3倍。野牡丹、桃金娘和山苍子这3个种生物量的增加主要是由个体高度和地径的增长引起,而栀子和梅叶冬青生物量的增加是其个体数量、株高和地径做出的贡献。(3)在尾叶桉林生长过程中,耐阴种类梅叶冬青随着个体密度、株高和地径的增长,其在林内发展成为主要优势灌木种类;栀子的密度虽然有一定的增大,但其株高和地径增加量小,生物量也小,发展为次要灌木种类。野牡丹、桃金娘和山苍子等3种的个体密度都在下降,虽然其地径、株高和生物量也有一定的增长,但其较强的阳生性不能适应林内的荫蔽环境,只适于林窗或林缘光照较强的地方生长。因此,在桉树林经营管理过程中应适当保护和种植梅叶冬青和栀子;在短轮伐期经营模式下,林窗和林缘也可适当保留野牡丹、桃金娘和山苍子。     

Growth of Geotrichum candidum and Penicillium camemberti Cultivated on Liquid Media Correlated with Ammonia and Methanethiol Emission

The growth of Geotrichum candidum and Penicillium camemberti plays an important role in the ripening of Camembert‐type cheeses, but the monitoring of the corresponding kinetics for fungal cocultures on solid media appears difficult. Continuous and non‐intrusive methods to characterize the growth of both species (like the monitoring of the emissions of ammonia and volatile sulphur compounds) may be highly relevant, under the condition that such emissions could be correlated with growth. This would be easier to investigate in submerged culture, since total biomass concentration is known to vary in proportion to broth turbidity. For this reason, growth kinetics, ammonia and flavour gas emission of both Geotrichum candidum and Penicillium camemberti grown separately in submerged cultures under the conditions of low aeration rate and uncontrolled pH were continuously recorded.In the basal medium (peptone+lactate supplemented with both glutamic acid and methionine [1 g/l]each), no significant gas emission was observed during the growth of both fungi. Ammonia and sulphur gas emissions by G. candidum were a little stimulated by supplementing the basal medium with trace elements, and, at a larger extent, by the addition of inorganic phosphate: Such a gaseous emission took place at the end of the growth phase of G. candidum. Irrespective of the basal medium supplementation, no significant emission ofammonia and sulphur gas was observed during the growth of P. camemberti. For the media and strains used, ammonia and volatile sulphur compounds emissions unequivocally showed the growth of Geotrichum candidum.     

A review ofPseudo-nitzschia,with special reference to the Skagerrak,North Atlantic,and adjacent waters

ThePseudo-nitzschia flora of the Skagerrak, North Atlantic, and adjacent waters, comprisingP. pungens, P. multiseries, P. seriata, P. fraudulenta, P. heimii, P. delicatissima, andP. pseudodelicatissima, has been examined. Except forP. australis, allPseudo-nitzschia species shown to produce the toxin domoic acid are present in the area although an outbreak of amnesic shellfish poisoning has never been reported. For comparison of morphological and taxonomic characters,Pseudo-nitzschia seriata f.obtusa, P. australis, P. subfraudulenta, P. subpacifica, P. lineola, P. inflatula, andP. cuspidata have been included in this investigation. Fine details of band structure and poroid occlusions, previously ignored or unresolved, have proven to add to the morphological distinction betweenP. pungens andP. multiseries, P. seriata andP. fraudulenta, P. seriata andP. australis, andP. delicatissima andP. pseudodelicatissima. Additional information on the structure of the proximal mantle compared to that of the valve face has revealed similarities in most of the species but differences betweenP. pungens andP. multiseries. The species seasonal and long-term distributional patterns during the sampling period (October 1978 through September 1993) in the Skagerrak area are outlined. The greatest abundances ofP. seriata, a cold-water species most likely restricted to the northern hemisphere, occurred in the spring, and those of the presumably cosmopolitan diatomsP. pungens, P. multiseries andP. pseudodelicatissima, in the autumn. WhereasP. multiseries seems to have decreased in abundance in the 1990s,P. pseudodelicatissima has apparently increased.     

Diversity and characteristics of terrestrial cyanobacteria near gimhae city,Korea

Cyanobacteria samples were collected from smears and mats growing on the surfaces of buildings, roadside walls, rocks, and monuments near Gimhae City, Korea. These populations were not as thick and dense, nor as diverse, as those found in temperate and tropical regions. Crusts at Gimhae City were dominated by colonial forms of species fromChroococcus, Aphanothece,Aphanocapsa, Gloeofthece,Gloeocapsa, andChroococidiopsis, as well as by filamentous forms from species ofScytonema andTolypothrix. The colonial forms appeared primarily as smears, especially for species ofGloeocapsa andChroococcidiopsis, whereas filamentous forms were the major constituents of mat populations. Invariably, the cells and trichomes of these colonial and filamentous forms were enclosed with a thick, firm, multi-colored laminar sheath, as seen by transmission electron microscopy. Such an association between cell walls and trichomes was stronger in the colonial forms, particularly forGloeocapsa, compared with the filamentous forms, in which the sheath did not detach from the wall even under arid conditions. The organization of cytoplasmic structures, particularly the thylakoid membranes, was least affected by acute dryness.     

Studies on the fungi associated with tomato fruit rots and effects of environment on storage

Seven fungi associated with fruit rot of tomato were isolated includingFusarium equiseti, F. chlamydosporum, Alternaria solani, Geotrichum candidum, Acremonium recifei, Aspergillus flavus andA. niger. They were all pathogenic on tomato fruits, most pathogenic beingGeotrichum candidum followed byA. niger. Least rot was caused byAlternaria solani. The optimum temperature for maximum rotting caused byG. candidum, A. niger andA. flavus was 30°C. The relative humidity for maximum rot ranged from 70–90%. Tomato fruits stored well at 0–10°C and rather poorly at 20–30°C. Fruits stored at 35°C showed blemishes. The best RH for storage ranged between 60 and 90%.     

Seed morphology and variation in the genus<Emphasis Type="Italic"> Pachycereus</Emphasis> (Cactaceae)

Seeds of 13 Pachycereus species and two Stenocereus species that have been suggested as closely related were examined with the scanning electron microscope. Quantitative features were evaluated using multivariate analysis in order to identify characters that distinguish them. Several species groups were recognized on the basis of 16 qualitative characters. All species studied are keeled. Stenocereus aragonii and S. eichlamii share with most Pachycereus species large size, glossy appearance, and a flat relief on periclinal cells in the lateral region. Pachycereus gatesii and P. schottii are unique in having the smallest seeds and a deeply impressed hilum-micropylar region. P. hollianus does not exhibit micro-relief on periclinal walls in the lateral region, and P. fulviceps has no expanded testa border. Multivariate analysis showed that four characters, length, breadth, hilum-micropylar region length, and angle, made the greatest contribution to distinguishing among species groups. More than 80% of P. fulviceps, P. hollianus, P. tepamo, P. weberi, and S. eichlamii seeds could be classified correctly using four seed features and the percentage was even higher using just two or three features for P. gatesii, P. grandis, P. militaris, P. pringlei, and P. schottii. Testa appearance, testa cell-pattern, and position relative to the rim of the hilum-micropylar region were found to be potentially informative and should be combined with other sources of data in future phylogenetic analyses.     

Population structure,gene diversity,and differentiation in natural populations of Cedar pines (Pinus subsect.Cembrae,Pinaceae) in the USSR

Twelve natural populations of four cedar pine species,Pinus sibirica, P. cembra, P. pumila, andP. koraiensis, occurring in the Soviet Union were investigated by starch-gel electrophoresis. Frequencies of 55 alleles at 19 loci were determined. Interpopulation genetic diversity inP. sibirica andP. pumila was only 2–4 per cent of the total genetic diversity. Nei's distance coefficient (Dn) was used to estimate the level of genetic differentiation among conspecific populations and among species. Dn values among populations ranged from 0.006 to 0.038. A dendrogram constructed using Dn values divided cedar pines species into 2 clusters:sibirica-cembra (Dn = 0.030) andpumila-koraiensis (Dn = 0.143). Nei's distance between these clusters was 0.232. On the basis of the data obtained it was possible to draw the following conclusion:P. sibirica, P. pumila, andP. koraiensis are distinct species, whileP. cembra should apparently be regarded as geographicalP. sibirica race.     

Utility of ionic liquid for Geotrichum candidum-catalyzed synthesis of optically active alcohols

Ionic liquids have recognized as a solvent for Geotrichum candidum-catalyzed optical resolution and/or deracemization of racemic secondary alcohols, giving optically active alcohols. The immobilized Geotrichum candidum proceeded the enantioselective oxidation of alcohols, producing chiral alcohols in an ionic liquid. Further, deracemization of racemic alcohols was proceeded to give the corresponding chiral alcohols in high yield with excellent stereoselectivity by the Geotrichum candidum–NaBH₄ system in the mixture of MES buffer solution and ionic liquid.     

On the cell wall composition of the apiculate yeasts

Summary Sonic oscillation was used for the purpose of obtaining clean, chemically intact cell walls. The rate of disruption was determined for cells ofHanseniaspora uvarum andSaccharomyces cerevisiae. The carbohydrate fractions of cell walls ofHanseniaspora uvarum, H. valbyensis, Kloeckera apiculata, Saccharomycodes ludwigii andSaccharmyces cerevisiae were shown to be similar. Chromatography of cell wall hydrolysates of all these species demonstrated that glucose and mannose were the only sugars present (in about equal amounts) besides traces of glucosamine. The cell walls ofH. uvarum contained 78.1 per cent carbohydrates, 7 per cent protein and approximately 0.05 per cent of chitin. Fractionation of the polysaccharides lead to a recovery of 83.3 per cent of the carbohydrates present (30.4 per cent glucan and 34.9 per cent mannan). Saccharomyces cerevisiae cell walls were found to have a carbohydrate content of 82.8 per cent, 6.5 per cent protein and a trace of chitin (0.04 per cent). Nadsonia elongata contained a relatively large amount of chitin (ca. 5 per cent) and lacked mannan in its cell walls. It was concluded thatHanseniaspora andSaccharomycodes are closely related to theSaccharomyceteae but they have little in common with species ofNadsonia.     

Carbohydrate composition and taxonomy of the genusDipodascus

Carbohydrates released during acid hydrolysis of intact cells ofDipodascus were studied by gas-liquid chromatographic analysis as their trimethylsilyl derivatives. In addition, cells were characterized by pyrolysis gas-liquid chromatography and pyrolysis mass spectrometry. The data obtained support the classification ofDipodascus uninucleatus in a separate genusDipodascopsis. Glucuronic acid is present inD. uninucleatus and, therefore, a possible affinity to fungi classified in the Zygomycetes is considered.Dipodascus aggregatus andDipodascus australiensis were found to be rather different, but very close toGeotrichum candidum and related species.     

Host specificity and growth of kelp gametophytes symbiotic with filamentous red algae (Ceramiales,Rhodophyta)

Kelp gametophytes were previously observed in nature living endophytically in red algal cell walls. Here we examine the interactions of two kelp species and six red algae in culture. Gametophytes of Nereocystis luetkeana (Mertens) Postels et Ruprecht became endophytic in the cell walls of Griffithsia pacifica Kylin and Antithamnion defectum Kylin, and grew epiphytically in high abundance on G. japonica Okamura and Aglaothamnion oosumiense Itono. Alaria esculenta (Linnaeus) Greville from the Atlantic coast of Nova Scotia became endophytic in Aglaothamnion oosumiense, Antithamnion defectum, Callithamnion sp., G. japonica, G. pacifica, and Pleonosporium abysicola Gardner, all from the Pacific Ocean. Some cultures were treated with phloroglucinol before infection to thicken the cell walls. The endophytic gametophytes were smaller and grew more slowly than gametophytes epiphytic on the same host. N. luetkeana failed to become endophytic in some of the potential hosts, and this may reflect host specificity, or culture artifacts. This work improves our understanding of the process of infection of red algae by kelp gametophytes, and broadens our knowledge of host specificity in endophytic symbioses.Communicated by K. Lüning     

Localization of chitin on ultrathin sections of cysts of two ciliated protozoa,Blepharisma undulans andPseudomicrothorax dubius,using colloidal gold conjugated wheat germ agglutinin

Summary The cyst walls of the ciliatesBlepharisma undulans andPseudomicrothorax dubius were examined ultrastructurally and by postembedding labeling with wheat germ agglutinin (WGA)-gold conjugate. Different methods of fixation and embedding were performed. In all procedures, WGA-gold binds selectively to material of the cyst wall. Pretreatment of the sections with chitinase inhibits labeling. The cyst walls of both species contain 3 nm fibrils, which are supposed to be of chitinous nature. In the cyst wall ofB. undulans, several thin layers of WGA-binding fibrils are interspaced with thick layers of other material. InP. dubius, WGA-binding sites are mainly concentrated in the mesocyst, where the microfibrils appear to represent the major component. These results obtained from two phylogenetically distant species confirm that chitin synthesis is an ancestral feature of ciliated protozoa. The amount and distribution of the chitin fibrils may play an important role in the properties and functions of the wall of the resting cyst.     

Identification of Geotrichum candidum at the species and strain level: proposal for a standardized protocol

In this study, the M13 primer was used to distinguish Geotrichum candidum from the anamorphic and teleomorphic forms of other arthrospore-forming species (discriminatory power = 0.99). For intraspecific characterization, the GATA4 primer showed the highest level of discrimination for G. candidum among the 20 microsatellite primers tested. A molecular typing protocol (DNA concentration, hybridization temperature and type of PCR machine) was optimized through a series of intra- and interlaboratory trials. This protocol was validated using 75 strains of G. candidum, one strain of G. capitatum and one strain of G. fragrans, and exhibited a discrimination score of 0.87. This method could therefore be used in the agro-food industries to identify and to evaluate biodiversity and trace strains of G. candidum. The results show that the GATA4 primer might be used to differentiate strains according to their ecological niche.     

Über die Struktur und Teilung der Kerne von Geotrichum candidum und Geotrichum magnusii

Summary The nuclei in germinating spores and growing hyphae ofGeotrichum magnusii andG. candidum have been examined during life and in fixed and stained preparations.The spores and the cells of the hyphae are multinucleate. The nuclei consist of a dense Feulgen-negative nucleolus surrounded by a less dense shell of Feulgen-positive particles. No membrane was seen at the margin of either living or fixed and stained nuclei. The mass of chromatin and the nucleolus divide at the same time by elongation followed by constriction. Chromosomes could not be detected in either resting or dividing nuclei.

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Mit 5 Textabbildungen     

A comparison of the fine structure ofMicrococcus diversus,Neisseria catarrhalis andNeisseria sicca

Examination of thin sections ofMicrococcus diversus, Neisseria catarrhalis andN. sicca reveals a cytoplasmic membrane, nuclear region and ribosomes which correspond to those recognized in other bacteria. Their cell walls are multilayered and resemble those of gram-negative microorganisms. Structures suggestive of mesosomes were observed inM. diversus andN. catarrhalis. These cytological observations concur with the findings of Kocur for the fine structure ofMicrococcus denitrificans andM. halodenitrificans. The taxonomic position of the problematic species in the genusMicrococcus is discussed.This study was supported by a grant from United States Department of Agriculture (USDA) Forest Service 4000, Grant 1.