In this research communication we report on the diversity of yeast and mould species in 69 samples of milk and different dairy products from three plants located in Umbria, central Italy. Isolates were characterised both macroscopically and microscopically and then identified by PCR and genome sequencing of the ITS region and the D1–D2 domain of the large-subunit rRNA gene for filamentous fungi and yeasts, respectively. Out of the 69 samples analysed, 51 (73.9%) tested positive for the presence of yeasts, whereas moulds were detected in 25 (36.2%) samples. A total of 9 yeast species belonging to 8 different genera and 13 mould species belonging to 6 different genera were isolated. The most common genera isolated were Debaryomyces and Kluyveromyces among the yeasts and Penicillium and Galactomyces among the moulds. Microbiota play a key role in the formation of flavour, aroma, texture and appearance of dairy products. This complex microbial ecosystem includes both cultured and external bacteria, yeasts and moulds. Some of them have an important role in the production of cheeses, whereas others are responsible for dairy product spoilage, resulting in significant food waste and economic losses. Some species can produce mycotoxins, representing a potential hazard for the consumer's safety. This study provides interesting information on the diversity of fungi species in dairy products from central Italy that can be of major importance to identify these products and to develop adequate strategies for fungal spoilage control and consumer safety.

The lichenicolous ‘heterobasidiomycetes’ belong in the Tremellomycetes (Agaricomycotina) and in the Pucciniomycotina. In this paper, we provide an introduction and review of these lichenicolous taxa, focusing on recent studies and novelties of their classification, phylogeny and evolution. Lichen-inhabiting fungi in the Pucciniomycotina are represented by only a small number of species included in the genera Chionosphaera, Cyphobasidium and Lichenozyma. The phylogenetic position of the lichenicolous representatives of Chionosphaera has, however, never been investigated by molecular methods. Phylogenetic analyses using the nuclear SSU, ITS, and LSU ribosomal DNA markers reveal that the lichenicolous members of Chionosphaera form a monophyletic group in the Pucciniomycotina, distinct from Chionosphaera and outside the Chionosphaeraceae. The new genus Crittendenia is described to accommodate these lichen-inhabiting species. Crittendenia is characterized by minute synnemata-like basidiomata, the presence of clamp connections and aseptate tubular basidia from which 4–7 spores discharge passively, often in groups. Crittendenia, Cyphobasidium and Lichenozyma are the only lichenicolous lineages known so far in the Pucciniomycotina, whereas Chionosphaera does not include any lichenicolous taxa.

This study aimed to explore the potential associations of adherence to the Mediterranean diet with gut microbiota characteristics and gastrointestinal symptomatology in an adult population. Other long-term dietary habits (e.g. consumption of snacks and junk food or stimulant intake) were also evaluated in terms of the gut microbiota profile. Participants (n 120) underwent anthropometric, dietary, physical activity and lifestyle evaluation. Adherence to the Mediterranean diet was assessed using a Mediterranean diet score, the MedDietScore, and subjects were classified into three tertiles according to individual adherence scoring. Gut microbiota composition was determined using quantitative PCR and plate-count techniques, and faecal SCFA were analysed using GC. Gastrointestinal symptoms were also evaluated. Participants with a high adherence to the Mediterranean diet had lower Escherichia coli counts (P=0·022), a higher bifidobacteria:E. coli ratio (P=0·025), increased levels and prevalence of Candida albicans (P=0·039 and P=0·050, respectively), greater molar ratio of acetate (P=0·009), higher defaecation frequency (P=0·028) and a more pronounced gastrointestinal symptomatology compared with those reporting low adherence. A lower molar ratio of valerate was also observed in the case of high adherence to the Mediterranean diet compared with the other two tertiles (Pfor trend=0·005). Positive correlations of MedDietScore with gastrointestinal symptoms, faecal moisture, total bacteria, bifidobacteria:E. coli ratio, relative share of Bacteroides, C. albicans and total SCFA, as well as negative associations with cultivable E. coli levels and valerate were indicated. Fast food consumption was characterised by suppressed representation of lactobacilli and butyrate-producing bacteria. In conclusion, our findings support a link between adherence to the Mediterranean diet and gut microbiota characteristics.

This work applies culture-independent methods for the characterization of fungal populations (yeasts and moulds) naturally occurring in Sardinian ewe's milk sampled in the Italian areas with the largest dairy production (Sardinia and Lazio regions). Sequences of the D1/D2 variable domains at the 5′ end of the 26S rRNA gene were obtained by amplification of DNA directly isolated from milk, and this allowed identification of a total of 6 genera and 15 species of fungi. Among the 6 identified genera Geotrichum spp., Candida spp., Phaeosphaeriopsis spp., Pestalotiopsis spp. and Cladosporium spp. belong to the phylum of Ascomycota, while Cryptococcus spp. is part of the phylum of Basidiomycota. In particular, two genera (Pestalotiopsis and Phaeosphaeriopsis) and two species (Plectosphaerella cucumerina and Pryceomyces carsonii) have never been reported in dairy ecosystems before. Results provide evidence that several moulds and yeasts, previously described only in ovine cheeses, are transferred directly from raw milk. The knowledge of fungal consortia inhabiting sheep raw milk is a particularly relevant issue because several species are directly involved in cheese making and ripening, determining the typical aroma. On the other hand, spoilage yeasts and moulds are involved in anomalous fermentation of cheese and may be responsible for considerable economic losses and serious risks for consumers' health.

PCR primers were developed for the specific detection of Clavispora lusitaniae, Debaryomyces hansenii var hansenii, Geotrichum candidum, Kluyveromyces lactis and K. marxianus and Yarrowia lipolytica, yeast species commonly found on the surface of smear cheese. Forty eight representative strains frequently found in smear cheeses or taxonomically related to the target yeasts were used as templates, to validate the designed primers. The specific and selective detection of these yeasts was effective in situ, in Livarot smear, without yeast isolation and culture and was comparable with data obtained with a conventional method. The primers described here have thus potential for PCR studies applied to cheese. It should also be possible to use some of these primers with other substrates.

Six ruminal cannulated Texel sheep were used to assess the dose response and the effect of live yeasts (Levucell® SC, Saccharomyces cerevisiae CNCM I-1077) on the prevention of induced ruminal latent acidosis. The sheep received, in a replicated 3×3 Latin-square design, an acidotic diet (wheat +lucerne hay, 60:40 (dry matter (DM) basis); starch: 410 g/kg DM) without yeast (control group; L0 treatment), supplemented with 0·2 g/day yeast (4×109 colony-forming units (c.f.u.) per day corresponding to producer recommendations; L1 treatment) or with 2 g/day yeast (4×1010 c.f.u. per day; L10 treatment). The following measurements were carried out: food intake, ruminal pH, ruminal volatile fatty acids (VFA), lactate and ammonia (NH3) concentrations, protozoal and lactate-utilizing bacterial counts, relative proportions of two main bacteria implicated in lactate metabolism (a lactate-producing species, Streptococcus bovis, and a lactate-utilizing species, Megasphaera elsdenii) using specific 16S-rRNA-targeting oligonucleotide probes, activities of lactate dehydrogenase (LDH) and of polysaccharidases involved in plant cell wall (xylanase, carboxymethylcellulase) and starch (amylase) degradation. The acidotic diet (L0) induced a butyric (12 mol per 100 mol total VFA) rather than lactic (<4 mmol/l) ruminal latent acidosis. Ruminal pH, fermentative patterns, lactate-metabolizing bacteria (concentration, LDH activity) and polysaccharidase activities were similar between treatments (P>0·1). Yeast supplementation tended to increase ruminal protozoal population (P<0·1) and intake of animals (P<0·1). The recommended yeast concentration (L1) was sufficient to ensure these effects. The yeast's action may differ according to the nature and function of the micro-organisms involved and the type of fermentative pattern favoured (protozoa/butyrate v. lactate-metabolizing bacteria/lactate and propionate) during ruminal acidosis in sheep.

The aim of this study was to evaluate the effects of two dietary supplements (monensin and a live yeast culture) on acid-base balance in steers maintained in a commercial feedlot system, considering effects over the finishing productive cycle. Steers (no. =42) were allotted randomly to one of the three study groups: (1) control group (no supplementation, C), (2) monensin supplementation (MON), and (3) live Saccharomyces cerevisiae strain 47 supplementation (SACC). Venous blood samples were collected for the measurement of acid-base parameters and l-lactate. Production parameters were also used as a complementary tool for understanding the internal changes associated with supplementation. During the finishing period, MON steers tended to gain more efficiently than C and SACC steers. In the C group, the finishing-period diet caused a progressive decline in blood bases, in line with the high-grain diet consumption. In contrast, supplemented animals did not show this trend, although lower HCO3− and base excess values were registered in SACC steers than in MON, indicating that ionophore supplementation is less effective for reducing blood base consumption than yeast supplementation. In our study, the lack of the expected response to yeast supplementation may be attributable to the high crude protein content of the ration, a common feature of commercial feedlot industries.

Volatile sulphur compounds are major flavouring compounds in many traditional fermented foods including cheeses. These compounds are products of the catabolism of L-methionine by cheese-ripening microorganisms. The diversity of L-methionine degradation by such microorganisms, however, remains to be characterized. The objective of this work was to compare the capacities to produce volatile sulphur compounds by five yeasts, Geotrichum candidum, Yarrowia lipolytica, Kluyveromyces lactis, Debaryomyces hansenii, Saccharomyces cerevisiae and five bacteria, Brevibacterium linens, Corynebacterium glutamicum, Arthrobacter sp., Micrococcus luteus and Staphylococcus equorum of technological interest for cheese-ripening. The ability of whole cells of these microorganisms to generate volatile sulphur compounds from L-methionine was compared. The microorganisms produced a wide spectrum of sulphur compounds including methanethiol, dimethylsulfide, dimethyldisulfide, dimethyltrisulfide and also S-methylthioesters, which varied in amount and type according to strain. Most of the yeasts produced methanethiol, dimethylsulfide, dimethyldisulfide and dimethyltrisulfide but did not produce S-methylthioesters, apart from G. candidum that produced S-methyl thioacetate. Bacteria, especially Arth. sp. and Brevi. linens, produced the highest amounts and the greatest variety of volatile sulphur compounds including methanethiol, sulfides and S-methylthioesters, e.g. S-methyl thioacetate, S-methyl thiobutyrate, S-methyl thiopropionate and S-methyl thioisovalerate. Cell-free extracts of all the yeasts and bacteria were examined for the activity of enzymes possibly involved in L-methionine catabolism, i.e. L-methionine demethiolase, L-methionine aminotransferase and L-methionine deaminase. They all possessed L-methionine demethiolase activity, while some (K. lactis, Deb. hansenii, Arth. sp., Staph. equorum) were deficient in L-methionine aminotransferase, and none produced L-methionine deaminase. The catabolism of L-methionine in these microorganisms is discussed.

Introduction. Black plum (Vitex doniana sweet) is a plant widely used by various communities in Nigeria for many purposes, including production of wine and jam. Ripe mature black plum fruit for food use commonly collected from the ground instead of plucked which is more hygienic. An investigation was conducted to find out the mycoflora on the surface and within the fruits of black plum fruits. Materials and methods. Mycoflora associated with the surface, skin, pulp and juice of healthy intact fruits of black plum was isolated in malt extract agar and potato dextrose agar and identified. Pathogenicity test was carried out with three pathogens of the fruit. Results and discussion. Fungi with incidence of 20-100% of occurrence were classified as major components and those below 20% of occurrence were regarded as minor components. Yeasts making up the major components in the ripe fruits were Saccharomyces cerevisiae and Candida albicans, while the minor yeast components were Rhodotrula sp., S. coreanus and Pichia sp. Filamentous fungi constituting major components were Aspergillus niger and Penicillium sp. whereas minor components were Botryodiplodia theobromae and Penicillium chrysogenum. Spoilage of fruits was caused by B. theobromae, Aspergillus sp. and A. niger. Conclusion. Yeasts and, occasionally, filamentous fungi occur within apparently healthy fruits of Vitex doniana sweet. Stages of development when the yeasts enter the pulp of the fruit need to be thoroughly investigated.