Isolation of Hanseniaspora uvarum (Kloeckera apiculata) in humans

Isolation of Hanseniaspora uvarum, a yeast of the ascomycetes group, whose anamorph corresponds to Kloeckera apiculata, obtained from stool and two ungual specimens from three patients, is reported. This yeast has been found in soil, water, various fruits, bivalve molluscs, crabs, prawns and fruit flies; in Spain, it has been described in the fermentation processes of some wines. In our region, it has also been found in the intestine of mackerel ( Scomber scombrus). Its finding in humans constitutes a clinical rarity. This revised version was published online in August 2006 with corrections to the Cover Date.     

Volatile metabolites produced in wine by mixed and sequential cultures of Hanseniaspora guilliermondii or Kloeckera apiculata and Saccharomyces cerevisiae

Summary Secondary products in wines obtained by pure, mixed and sequential cultures of Saccharomyces cerevisiae, Hanseniaspora guilliermondii or Kloeckera apiculata were studied. Consistent differences in the composition were determined in wines fermented by sequential cultures. When S. cerevisiae was added to musts partially fermented by apiculate yeasts, its metabolism was significantly affected. In particular it synthesized high amounts of n-propanol and metabolized high quantities of acetoin, produced by apiculate yeasts     

Kloeckera apis st. nov.; the imperfect state of Hanseniaspora guilliermondii Pijper

K. apiculata var. apis (nom. nud.) was found to be the imperfect state of H. guilliermondii Pijper by a high degree of DNA reassociation. The name is validated and raised to species rank, K. apis Lavie ex Smith, Simione and Meyer. K. apis and H. guilliermondii could be distinguished from H. uvarum and H. valbyensis by a low DNA reassociation and by growth at 37 C.DNA studies performed at the ATCC were supported in part by Public Health Service grant GM 19240-05.     

Systematics of Hanseniaspora Zikes and Kloeckera Janke

The physiological and morphological characteristics of eighty-two strains of Hanseniaspora and Kloeckera, representing twenty-nine described species, were examined. These results along with DNA base composition and DNA/DNA reassociation experiments revealed that the genus Hanseniaspora comprises six distinct species, viz. H. valbyensis, H. uvarum, H. guilliermondii, H. occidentalis, H. osmophila and H. vineae, with K. japonica, K. apiculata, K. apis, K. javanica, K. corticis and K. africana, respectively, as their imperfect states.     

Killer character in Kluyveromyces yeasts: Activity on Kloeckera apiculata

The screening for ‘killer’ character among 23 strains of the genus Kluyveromyces showed the presence of three ‘killer’ strains (K. phaffii; K. vanudenii and K. wikenii). Their ‘killer’ activity against yeasts of other genera is in agreement with the results of others studies. Particularly K. phaffii inhibited many strains of Kloeckera apiculata.     

Flocculation and cell surface characterization of Kloeckera apiculata from wine

AIMS: To characterize and analyze the flocculation phenomenon of Kloeckera apiculata mc1 from Argentinian wine to understand the cell-cell interaction pattern. METHODS AND RESULTS: Kloeckera apiculata mc1 possess intense cell-cell interactions in MYPG medium (0.5% malt extract, 1% yeast extract, 2% glucose, 2% peptone), pH 5.5 by shaking at 25 degrees C. Optimum flocculation is observed at pH 4.5 in the presence of 3 mmol l-1 Ca2+. The flocculation is induced by peptone and malt extract and not by yeast extract and is reversed by 50 mmol l-1 galactose or lactose. The flocculation is highly susceptible to pronase, chymotrypsine and proteases types IV and XXVII and is partially resistant to trypsin. The electronic microscopy shows that the cells are attached to each other along their sides by fine hair-like threads. CONCLUSIONS: The mechanism of flocculation of K. apiculata mc1 is mediated by protein-carbohydrate interaction, stabilized by Ca2+. SIGNIFICANCE AND IMPACT OF THE STUDY: The use of selected pure yeast inocula of known ability is preferred to wine elaboration, therefore the indigenous flora must be avoided and the flocculation of K. apiculata could be an economic method to do it.     

Typing of Saccharomyces cerevisiae and Kloeckera apiculata strains from Aglianico wine

AIMS: Kloeckera apiculata and Saccharomyces cerevisiae yeast species are dominant, respectively, at the early and at the following stages of wine fermentation. In the present study, PCR fingerprinting and NTS region amplification and restriction were applied as techniques for monitoring yeast population performing Aglianico of Vulture grape must fermentation. METHODS AND RESULTS: Thirty S. cerevisiae and 30 K. apiculata strains were typed by PCR fingerprinting with (GAC)5 and (GTG)5 primers and by complete NTS region amplification followed by restriction with HaeIII and MspI enzymes. S. cerevisiae strains generated two patterns with (GAC)5 primer, while (GTG)5 primer yielded a higher genetic polymorphism. Conversely, in K. apiculata Aglianico wine strains (GAC)5 and (GTG)5 primers generated the same profile for all strains. Restriction analysis of the amplified NTS region gave the same profile for all strains within the same species, except for one strain of S. cerevisiae. CONCLUSIONS: The PCR fingerprinting technique was useful in discriminating at strain level S. cerevisiae, particularly with the primer (GTG)5. RFLP patterns generated from the NTS region of the two species can be more easily compared than the patterns resulting from PCR fingerprinting, thus RFLP is more suitable for the rapid monitoring of the species involved in different stages of fermentation. SIGNIFICANCE AND IMPACT OF THE STUDY: The molecular techniques used allow discrimination of S. cerevisiae at strain level and monitoring of the ratio of S. cerevisiae/K. apiculata during the fermentation process. Thus, their application can assure technological adjustments in a suitable time.     

Physiological behaviour of Hanseniaspora guilliermondii in aerobic glucose-limited continuous cultures

The physiology of Hanseniaspora guilliermondii was studied under aerobic glucose-limited conditions using the accelerostat procedure (continuous acceleration of dilution rate) and classical chemostat cultures. By both cultivation techniques this yeast was found to be Crabtree-positive. Up to a dilution rate of 0.25 h(-1), glucose was completely metabolised into biomass, glycerol and carbon dioxide. Above this value, an increase in the dilution rate was accompanied by the production of other metabolites like ethanol, acetic and malic acids. Biomass yield during the purely oxidative growth was 0.49 g g(-1) and decreased to 0.26 g g(-1) for D=0.34 h(-1). A maximal specific ethanol production rate of 1.36 mmol g(-1) h(-1) and a maximal ethanol yield of 0.05 g g(-1) were achieved at D=0.34 h(-1).     

Metabolic characterization of Kloeckera apiculata strains from star fruit fermentation

A total of 37 strains of Kloeckera apiculata was isolated during the spontaneous fermentation of star fruit must. Each strain was differentiated from the others on the basis of its capacity to produce acetaldehyde, ethyl acetate, higher alcohols, acetoin and acetic acid. All the strains were characterized by the low production of higher alcohols and the high production of ethyl acetate, whereas consistent differences in the production of acetaldehyde, acetoin and acetic acid served to differentiate star fruit apiculate strains into six different phenotypes, present at different stages of the fermentation process. The metabolic strain diversity found can be interpreted as a natural consequence of environmental conditions, which influenced the frequency and selection of specific apiculate strains. From the biotechnological point of view the different metabolic biotypes represent an important source of strains for potential use as starter cultures for star fruit fermentation.     

Characterization of Kloeckera apiculata strains from the Friuli regionin Northern Italy

Forty-nine strains of Kloeckera apiculata, isolated from the Friuli region in Italy, were differentiated on the basis of fermentation behaviour and production of secondary compounds in two different grape musts at 18 °C. The isolates exhibited a controlled production of acetic acid, only in a few cases more that 1 g/l. In Moscato grape must the strains exhibited a more uniform behaviour for the production of higher alcohols, ethyl acetate and acetoin than in red grapes. In general, higher levels of ethanol, glycerol and acetic acid were produced in red grape must fermentation. Apiculate strains behaved differently in the two musts, with different metabolic phenotypes dominating the fermentation process. The existence of different metabolic phenotypes correlated with the must composition underlines the need to perform a selection of indigenous apiculate yeasts to obtain the desired consistent products.     

The flocculation of wine yeasts: biochemical and morphological characteristics in Kloeckera apiculata

The floc-forming ability of flocculent strains of Kloeckera apiculata, isolated from musts, was tested for susceptibility to proteinase and sugar treatments. Three different flocculation phenotypes were discriminated by protease digestion, whereas the inhibition of flocculation by sugars distinguished two definite patterns: one mechanism of flocculation involved a galactose-specific protein and the other a broad-specificity lectin. SEM and TEM observation of the cell surface of two different Kloeckera strains revealed fine fibrils and a diffuse structure at the point of contact in one strain, and thick masses of mucus on the cell wall of the other strain.     

Molecular identification and genetic diversity within species of the genera Hanseniaspora and Kloeckera

Three molecular methods, RAPD-PCR analysis, electrophoretic karyotyping and RFLP of the PCR-amplified ITS regions (ITS1, ITS2 and the intervening 5.8S rDNA), were studied for accurate identification of Hanseniaspora and Kloeckera species as well as for determining inter- and intraspecific relationships of 74 strains isolated from different sources and/or geographically distinct regions. Of these three methods, PCR-RFLP analysis of ITS regions with restriction enzymes DdeI and HinfI is proposed as a rapid identification method to discriminate unambiguously between all six Hanseniaspora species and the single non-ascospore-forming apiculate yeast species Kloeckera lindneri. Electrophoretic karyotyping produced chromosomal profiles by which the seven species could be divided into four groups sharing similar karyotypes. Although most of the 60 strains examined exhibited a common species-specific pattern, a different degree of chromosomal-length polymorphism and a variable number of chromosomal DNA fragments were observed within species. Cluster analysis of the combined RAPD-PCR fingerprints obtained with one 10-mer primer, two microsatellite primers and one minisatellite primer generated clusters which with a few exceptions are in agreement with the groups as earlier recognized in DNA-DNA homology studies.     

柠檬形克勒克酵母对温州蜜柑“国庆一号”采后贮藏的防腐效果

Citrus unshiu “Guoqing No.1” was used as material for elucidating the effect of Kloeckera apiculata suspension post-harvest treatment on preventing fruit decay during storage period. The result showed that K. apiculata preparation of 2×106CFU/mL (CFU, cloning forming unit) significantly reduced the development of decay caused by green mold, blue mold and stem rot, showing the same effect of carbendazim treatment after storage period of 100 days, and the disease incidence was controlled under 1%. The antagonist population was between 3.5×105CFU/mL and 1.7×107CFU/mL in citrus fruit, and increased 1.7×107CFU/mL after 40 days storage, and then stabilized for the remaining storage period. After storage at 5℃ for 100 days, K. apiculata strain 34-9 did not alter any quality parameters of fruit. During storage, the change of relative electrical conductivity (REC) showed a coordinate tendency with malondlaldehyde (MDA) content, which kept continuously growing from the beginning to the end of the storage. In the storage period of 25 days, the REC value and MDA content of the treatments were all significantly lower than those of water treatment, but had no obvious difference from those of carbendazim treatment. K. apiculata strain 34-9 had no effect on the enzyme activity of super oxide dismutase (SOD), but significantly improved peroxidase (POD) activity after 100 day storage.     

柠檬形克勒克酵母对温州蜜柑“国庆一号”采后贮藏的防腐效果

本实验以温州蜜柑"国庆一号"为试材,研究了柠檬形克勒克酵母Kloeckera apiculata悬浮液处理对柑橘果实采后防腐保鲜效果及相关酶的影响.结果表明,贮藏100d后,2×106CFU/mL(CFC表示菌落形成单位)的酵母菌液能有效地抑制青霉、绿霉和蒂腐病等贮藏期病害的发生,柑橘果实的腐烂率控制在1%以下,效果显著高于清水对照,与多菌灵相当;柠檬形克勒克酵母在柑橘果皮上的数量在3.5×105CFU/mL-1.7×107CFU/mL之间,在贮藏40d后最大值达到1.7×107CFU/mL,随后保持在106CFU/mL左右,100d贮藏结束后,品质分析表明,柠檬形克勒克酵母处理对柑橘品质没有不良影响.在整个贮藏期细胞膜透性与丙二醛(MDA)含量变化相对应,均呈上升趋势,在贮藏25d左右时酵母菌液处理和多菌灵处理的细胞膜透性与MDA含量均显著低于清水对照,而前二者之间无差异.酵母菌液对超氧化物歧化酶(SOD)活性没有诱导作用,而在贮藏后100d左右对过氧化物酶(POD)活性有诱导作用.     

The effects of Kloeckera apiculata on the cuticular waxes of navel orange fruit

Kloeckera apiculata 34-9 was selected from the rhizosphere soil for its high efficacy in controlling citrus green and blue mold. In this study, the effect of the antagonistic yeast K. apiculata on citrus cuticular wax was investigated in ripening Newhall navel orange (Citrus sinensis L. Osbeck). Our results show that K. apiculata triggers the production of cuticular waxes and surface wax morphology changes in the fruit surface. 15 K. apiculata-responsive differentially expressed genes (DEGs) were identified for wax metabolism by using the Affymetrix citrus genome GeneChip. Using GC–MS, 46 wax compounds were found in the Newhall fruit surface. On one hand, esters including docosanoic acid, 1,2,3-propanetriyl ester and 9-hexadecenoic acid, 9-octadecenyl ester were up-regulated approximately twofold in the treatment condition compared with the control; and on the other hand, the fatty acids and fatty alcohols decreased by 74.4% and 72.3%, respectively, in the treatment condition. The new wax production and increased hydrophobicity of the Newhall surface resulting from the treatment may influence spore adhesion and germination. Furthermore, the response of the fruit waxes to the K. apiculata stimulus is likely to be regulated by intra-cellular H₂O₂ signaling. This study demonstrated the response fruit waxes to K. apiculata in Newhall navel oranges, thus providing new clues that aid our understanding of the mechanisms of action of antagonistic yeasts in postharvest fruits.     

Physiological variants of Saccharomyces cerevisiae and Kloeckera apiculata from palm wine and cashew juice

Physiological variants of Saccharomyces cerevisiae and Kloeckera apiculata have been identified in oil palm wine and cashew juice from Nigeria. Genomic DNA from the four S. cerevisiae variants had a % G + C of 36-41% while that of K. apiculata was 32.2%. Fermentation of cashew juice produced wine of alcoholic contents of 10% with S. cerevisiae, 8% with K. apiculata and 9.3% with both yeasts simultaneously.     

Physiological variants of Saccharomyces cerevisiae and Kloeckera apiculata from palm wine and cashew juice

Physiological variants of Saccharomyces cerevisiae and Kloeckera apiculata have been identified in oil palm wine and cashew juice from Nigeria. Genomic DNA from the four S. cerevisiae variants had a % G + C of 36–41% while that of K. apiculata was 32.2%. Fermentation of cashew juice produced wine of alcoholic contents of 10% with S. cerevisiae , 8% with K. apiculata and 9.3% with both yeasts simultaneously. and accepted 31 August 1989     

The conversion of 3-Hydroxy-2,4,5-trihydroxymethylpyridine into pyridoxine by Kloeckera apiculata.

Kloeckera apiculata, a vitamin B-6-dependent yeast, grows in the presence of 3-hydroxy-2,4,5-trihydroxymethylpyridine in vitamin B-6-free media. On a molar basis the growth-promoting activity of this compound is approximately one-tenth that of other forms of vitamin B-6. [G-3H]3-Hydroxy-2,4,5-trihydroxymethylpyridine is converted into radioactive vitamin B-6 compounds of the same specific radioactivity by growing cultures of K. apiculata.     

微波诱变选育柑橘采后病害拮抗菌柠檬形克勒克酵母研究

采用微波辐照,对柠檬形克勒克酵母(Kloeckera apiculata)进行诱变处理,确立了辐照的最佳时间为 50 s.选育到1株拮抗柑橘青霉病抑菌活性提高16.90%、生物量提高6.96%的菌株W50-56;选育到1株拮抗柑橘绿霉病抑菌活性提高17.38%、生物量提高5.86%的菌株W50-47.所选育的菌株经多次传代,遗传性状十分稳定.