Finding a correct species assignment for a Metschnikowia strain: insights from the genome sequencing of strain DBT012.

Metschnikowia pulcherrima is an important yeast species that is attracting increased interest thanks to its biotechnological potential, especially in agri-food applications. Phylogenetically related species of the so-called 'pulcherrima clade' were first described and then reclassified in one single species, which makes the identification an intriguing issue. Starting from the whole-genome sequencing of the protechnological strain Metschnikowia sp. DBT012, this study applied comparative genomics to calculate similarity with the M. pulcherrima clade publicly available genomes with the aim to verify if novel single-copy putative phylogenetic markers could be selected, in comparison with the commonly used primary and secondary barcodes. The genome-based bioinformatic analysis allowed the identification of 85 consensus single-copy orthologs, which were reduced to three after split decomposition analysis. However, wet-lab amplification of these three genes in nonsequenced type strains revealed the presence of multiple copies, which made them unsuitable as phylogenetic markers. Finally, average nucleotide identity (ANI) was calculated between strain DBT012 and available genome sequences of the M. pulcherrima clade, although the genome dataset is still rather limited. Presence of multiple copies of phylogenetic markers as well as ANI values were compatible with the recent reclassification of the clade, allowing the identification of strain DBT012 as M. pulcherrima.

Fate of Grape-Derived Terpenoids in Model Systems Containing Active Yeast Cells.

Terpenes are important contributors to wine aroma. Free and glycosidically bound terpenes are primarily formed in grapes. During fermentation, they undergo important transformation catalyzed by yeast, so that the terpene profile of grape is substantially different from that of the corresponding wine. The present paper assessed the ability of a Saccharomyces cerevisiae strain to transform 17 different terpenes. Biotransformation was performed by placing target compounds in incubation with resting cells. Volatile compounds produced were extracted by solid-phase extraction and analyzed by gas chromatography-mass spectrometry. Geranyl acetate, neryl acetate, citronellyl acetate, and menthyl acetate were formed from the corresponding terpene alcohols. ß-Citronellol was the main product of geraniol transformation; geranial, an intermediate of this pathway, has also been detected. Limonene was hydroxylated by yeast to form carveol, trans-2,8-menthadien-1-ol, and cis-2,8-menthadien-1-ol. Moreover, yeast cells were found to be able to adsorb a significant portion of the terpenes present in the reaction batches, with the extent of this phenomenon being linked to terpene hydrophobicity.

Antibiotic-Resistant Bacteria in Greywater and Greywater-Irrigated Soils.

This study represents the first systematic attempt to evaluate antibiotic-resistant bacteria (ARB) occurrence in treated greywater and the potential spread of these bacteria from the greywater to greywater-irrigated soil. Treated greywater from three recirculating vertical flow constructed wetlands, each located in a household in the central Negev Desert, Israel, was surveyed. The presence of antibiotic-resistant bacteria in raw and treated greywater was investigated with culture and molecular methods, as well as their presence in the corresponding treated-greywater-irrigated soils. Additionally, the effectiveness of chlorination to prevent the spread of ARB was tested. The total count of tetracycline-resistant bacteria significantly increased in the treated greywater, likely due to their concentration on the filter matrix of the treatment systems. Twenty-four strains of tetracycline-resistant bacteria were isolated and identified at the genus level by 16Sr RNA gene sequencing. All the tetracycline-resistant bacteria showed high resistance traits, and some of them presented multiple antibiotic resistances. Six tetracycline resistance genes (coding for efflux and ribosomal resistance mechanisms) and five ß-lactamase genes were detected. In 14 of the isolated strains, the gene tet39, which is phylogenetically related to both environmental and clinical strains, was identified. All the tet39 resistant bacteria were positive to at least one of the ß-lactamase genes tested. Chlorination was found to be an efficient method to reduce ARB in treated greywater. We concluded that disinfection of treated greywater may reduce the risks not only from the potential presence of pathogens but also from the presence of ARB and antibiotic resistance genes.

Impact of the reusing of food manufacturing wastewater for irrigation in a closed system on the microbiological quality of the food crops.

In order to evaluate if the reuse of food industry treated wastewater is compatible for irrigation of food crops, without increased health risk, in the present study a cropping system, in which ground water and treated wastewater were used for irrigation of tomato and broccoli, during consecutive crop seasons was monitored. Water, crop environment and final products were monitored for microbial indicators and pathogenic bacteria, by conventional and molecular methods. The microbial quality of the irrigation waters influenced sporadically the presence of microbial indicators in soil. No water sample was found positive for pathogenic bacteria, independently from the source. Salmonella spp. and Listeria monocytogenes were detected in soil samples, independently from the irrigation water source. No pathogen was found to contaminate tomato plants, while Listeria monocytogenes and E. coli O157:H7 were detected on broccoli plant, but when final produce were harvested, no pathogen was detected on edible part. The level of microbial indicators and detection of pathogenic bacteria in field and plant was not dependent upon wastewater used. Our results, suggest that reuse of food industry wastewater for irrigation of agricultural crop can be applied without significant increase of potential health risk related to microbial quality.