Investigation of the growth of Listeria in plant-based beverages.

The objective of the present study was to evaluate whether the content of sugar, protein, fat, or fibre in commercially available and specially formulated plant-based beverages (oat, soya and pea) influences the growth rates of Listeria. Beverages were inoculated with a strain cocktail of Listeria (approximately 1 × 103 CFU/mL), and the data demonstrated that Listeria could proliferate in all tested beverages. Moreover, varying concentrations of naturally occurring or added sugar (0-3.3%), protein (3.3-5%), fat (1.1-3.5%) and added fibre (0-1.5%) did not have a statistically significant (p > 0.05) impact on the growth rates of Listeria in the tested plant-based beverages. These data suggest that the wide variety of commercial plant-based beverages serve as an ideal medium for the growth of Listeria irrespective of product composition. All the various products tested provided sufficient nutrients to support at least a 2.6-log increase of Listeria within 16 h at room temperature, with some beverages supporting a 3-log increase. Therefore, these data highlight the importance of careful storage and handling of these increasingly varied and popular products.

Association of Virulence, Biofilm, and Antimicrobial Resistance Genes with Specific Clonal Complex Types of Listeria monocytogenes.

Precise classification of foodborne pathogen Listeria monocytogenes is a necessity in efficient foodborne disease surveillance, outbreak detection, and source tracking throughout the food chain. In this study, a total of 150 L. monocytogenes isolates from various food products, food processing environments, and clinical sources were investigated for variations in virulence, biofilm formation, and the presence of antimicrobial resistance genes based on their Whole-Genome Sequences. Clonal complex (CC) determination based on Multi-Locus Sequence Typing (MLST) revealed twenty-eight CC-types including eight isolates representing novel CC-types. The eight isolates comprising the novel CC-types share the majority of the known (cold and acid) stress tolerance genes and are all genetic lineage II, serogroup 1/2a-3a. Pan-genome-wide association analysis by Scoary using Fisher's exact test identified eleven genes specifically associated with clinical isolates. Screening for the presence of antimicrobial and virulence genes using the ABRicate tool uncovered variations in the presence of Listeria Pathogenicity Islands (LIPIs) and other known virulence genes. Specifically, the distributions of actA, ecbA, inlF, inlJ, lapB, LIPI-3, and vip genes across isolates were found to be significantly CC-dependent while the presence of ami, inlF, inlJ, and LIPI-3 was associated with clinical isolates specifically. In addition, Roary-derived phylogenetic grouping based on Antimicrobial-Resistant Genes (AMRs) revealed that the thiol transferase (FosX) gene was present in all lineage I isolates, and the presence of the lincomycin resistance ABC-F-type ribosomal protection protein (lmo0919_fam) was also genetic-lineage-dependent. More importantly, the genes found to be specific to CC-type were consistent when a validation analysis was performed with fully assembled, high-quality complete L. monocytogenes genome sequences (n = 247) extracted from the National Centre for Biotechnology Information (NCBI) microbial genomes database. This work highlights the usefulness of MLST-based CC typing using the Whole-Genome Sequence as a tool in classifying isolates.

Membrane filtration processing of infant milk formula alters protein digestion in young pigs.

Reducing heat treatment (HT) during processing of infant milk formula (IMF) is desirable to produce a product that more closely resembles breast milk. By employing membrane filtration (MEM), we produced an IMF (60:40 whey to casein ratio) at pilot scale (250 kg). MEM-IMF had a significantly higher content of native whey (59.9 %) compared to HT-IMF (4.5 %) (p < 0.001). Pigs, at 28 days old, were blocked by sex, weight and litter origin and assigned to one of two treatments (n = 14/treatment): (1) starter diet containing 35 % of HT-IMF powder or (2) starter diet containing 35 % of MEM-IMF powder for 28 days. Body weight and feed intake were recorded weekly. Pigs at day 28 post weaning were sacrificed 180 min after their final feeding, for the collection of gastric, duodenal, jejunum and ileal contents (n = 10/treatment). MEM-IMF diet resulted in more water-soluble proteins and higher levels of protein hydrolysis in the digesta at various gut locations compared to HT-IMF (p < 0.05). In the jejunal digesta, a higher concentration of free amino acids were present post MEM-IMF consumption (247 ± 15 µmol g-1 of protein in digesta) compared to HT-IMF (205 ± 21 µmol g-1 of protein). Overall, average daily weight gain, average dairy feed intake and feed conversion efficiency were similar for pigs fed either MEM-IMF or HT-IMF diets, but differences and trends to difference of these indicators were determined in particular intervention periods. In conclusion, reducing heat treatment during processing of IMF influenced protein digestion and revealed minor effects on growth parameters providing in vivo evidence that babies who are fed with IMF processed by MEM are likely to have different protein digestion kinetics but minimal effect on overall growth trajectories as babies fed IMF processed by traditional thermal processing.

Growth of food-borne pathogens Listeria and Salmonella and spore-forming Paenibacillus and Bacillus in commercial plant-based milk alternatives.

The objective of the present study was to compare the growth of food-pathogens Listeria monocytogenes, Salmonella enterica, food spoilage Bacillus subtilis, an industrial milk product isolate, and spore-forming Paenibacillus in commercially available ultrahigh temperature processed (UHT) bovine milk and non-dairy, plant-based beverages (coconut, almond, cashew) stored at chilled and ambient temperatures (4 °C, 8 °C or 20 °C). Beverage samples were inoculated with a strain cocktail or individual strains of either Listeria or Salmonella, or Paenibacillus or Bacillus, respectively (approximately 1 × 103 CFU/mL). The findings indicate that the bacterial strains used in the study were capable of proliferating in plant-based beverages at higher rates than in bovine milk at 8 °C and 20 °C for Listeria and 20 °C for Salmonella and Paenibacillus, respectively. Bacillus subtilis grew equally fast in bovine milk and plant-based almond drink at 20 °C. No statistically significant difference (p > 0.05) in growth rates between different types of tested beverages was observed at 4 °C and at 8 °C for Listeria and Salmonella cocktails, respectively. These data suggest that plant-based beverages may present a significant risk for listeriosis and salmonellosis and post-opening recommendations should be carefully considered.

Comparison of conventional heat-treated and membrane filtered infant formulas using an in vitro semi-dynamic digestion method.

Introducing membrane filtration steps into infant milk formula (IMF) manufacture can partly preserve native whey proteins in the final products. In this study, the IMF produced by membrane filtration (MEM-IMF) and conventional heat treatment (HT-IMF) were compared by using a novel semi-dynamic infant in vitro digestion method. MEM-IMF exhibited a fragmented curd during gastric digestion, and confocal laser light microscopy showed that protein aggregates had disassociated from the fat droplets within 93 min in the MEM-IMF digesta. In contrast, the digesta of HT-IMF showed a more extensive curd formation and denser protein aggregates, which remained intact until the end of gastric digestion. Molecular weight profiles and the primary amine assay suggested that protein degradation and peptide release were faster in the MEM-IMF. In conclusion, the presence of native whey protein in the IMF altered the gastric digestion kinetics by changing coagulation and formation of aggregates, potentially accelerating the rate of gastric emptying in vivo.

Correlation of organic acid tolerance and genotypic characteristics of Listeria monocytogenes food and clinical isolates.

A collection of Listeria monocytogenes isolates from various food products, food processing environments and clinical sources (n = 153) were evaluated for their tolerance to acetic, lactic and propionic acids. A large variation in tolerance was observed amongst isolates under mildly acidic conditions (pH 5.3) for acetic (5-20 mM undissociated acid) and propionic acid (2-10 mM undissociated acid) but there was less variation for lactic acid (3-6 mM undissociated acid). Analysis of the isolate genome sequences for a complement of genes previously shown to have a role in acid tolerance revealed that thiT, gadT2, gadD2 and gadD3 genes were linked to higher acetic acid tolerance (P < 0.05) while lisRK was linked to higher tolerance to propionic acid (P = 1 × 10-11). An absence of plasmid genes was also linked with isolates showing higher tolerance for all acids. Scoary GWAS analysis revealed that a total of 333, 207, and 333 genes were associated with acid tolerance for acetic, lactic, and propionic acid, respectively (P < 0.05). However, the p-value adjusted with Bonferroni's method for multiple comparisons did not reveal any significant associations. Isolates were grouped into clonal complexes (CC) using Multi Locus Sequence Typing (MLST) and MIC values for the three acids were determined for representative strains. One complex, CC18, showed significantly higher (P ≤ 0.05) acetic and propionic acid MIC values than other groups, whereas only CC7 type isolates revealed significantly higher (P ≤ 0.001) lactic acid MIC values. The results demonstrate that MLST typing could be linked to acid tolerance phenotypic traits which is important in predicting the behaviour of L. monocytogenes in food products.

Variability in Cold Tolerance of Food and Clinical Listeria monocytogenes Isolates.

The aim of this study was to investigate the level of strain variability amongst food and clinical Listeria monocytogenes isolates growing at low temperatures (4 and 7 °C) in both laboratory media and real food matrices. Isolates (n = 150) grown in laboratory media demonstrated a large variation in growth profiles measured using optical density. Overall, it was noted that clinical isolates exhibited a significantly higher growth rate (p ≤ 0.05) at 7 °C than the other isolates. Analysis of variance (ANOVA) tests of isolates grouped using Multi Locus Sequence Typing (MLST) revealed that clonal complex 18 (CC18) isolates were significantly (p ≤ 0.05) faster growing at 4 °C than other CC-type isolates while CC101, CC18, CC8, CC37 and CC14 were faster growing than other CC types at 7 °C. Euclidean distance and Ward method-based hierarchical clustering of mean growth rates classified 33.33% of isolates as faster growing. Fast and slow growing representative isolates were selected from the cluster analysis and growth rates were determined using plate count data in laboratory media and model food matrices. In agreement with the optical density experiments, CC18 isolates were faster and CC121 isolates were slower than other CC types in laboratory media, UHT milk and fish pie. The same trend was observed in chocolate milk but the differences were not statistically significant. Moreover, pan-genome analysis (Scoary) of isolate genome sequences only identified six genes of unknown function associated with increased cold tolerance while failing to identify any known cold tolerance genes. Overall, an association that was consistent in laboratory media and real food matrices was demonstrated between isolate CC type and increased cold tolerance.

Investigation of combinations of rationally selected bioengineered nisin derivatives for their ability to inhibit Listeria in broth and model food systems.

In this study, we examined the ability of nisin A and a rationally assembled bank of 36 nisin derivative producing Lactococcus lactis strains to inhibit Listeria. A broth-based bioluminescence assay for screening single and combinations of bioengineered nisin derivatives using cell-free supernatants (CFS) from nisin derivative producing strains was developed. In this way, we screened 630 combinations of nisin derivative producing strains, identifying two (CFS from M17Q + N20P and M17Q + S29E) which exhibited enhanced anti-listerial activity when used together compared to when used alone, or to the nisin A producing strain. Minimal inhibitory concentration assays performed with purified peptides revealed than when used singly, the specific activities of M17Q, N20P and S29E (3.75-7.5 µM) against L. innocua were equal to, or less than that of nisin A (MIC of 3.75 µM). Broth-based growth curve assays using purified peptides demonstrated that use of the double peptide combinations and a triple peptide combination (M17Q + N20P + S29E) resulted in an extended lag phase of L. innocua, while kill curve assays confirmed the enhanced bactericidal activity of the combinations in comparison to the single derivative peptides or nisin A. Furthermore, the enhanced activity of the M17Q + N20P combination was maintained in a model food system (frankfurter homogenate) at both chill (4 °C) and abusive (20 °C) temperature conditions, with final cell numbers significantly less (1-2 log10 CFU/ml) than those observed with the derivative peptides alone, or nisin A. To our knowledge, this study is the first investigation that combines bioengineered bacteriocins with the aim of discovering a combination with enhanced antimicrobial activity.

Investigating the Use of Ultraviolet Light Emitting Diodes (UV-LEDs) for the Inactivation of Bacteria in Powdered Food Ingredients.

The addition of contaminated powdered spices and seasonings to finished products which do not undergo further processing represents a significant concern for food manufacturers. To reduce the incidence of bacterial contamination, seasoning ingredients should be subjected to a decontamination process. Ultraviolet light emitting diodes (UV-LEDs) have been suggested as an alternative to UV lamps for reducing the microbial load of foods, due to their increasing efficiency, robustness and decreasing cost. In this study, we investigated the efficacy of UV-LED devices for the inactivation of four bacteria (Listeria monocytogenes, Escherichia coli, Bacillus subtilis and Salmonella Typhimurium) on a plastic surface and in four powdered seasoning ingredients (onion powder, garlic powder, cheese and onion powder and chilli powder). Surface inactivation experiments with UV mercury lamps, UVC-LEDs and UVA-LEDs emitting at wavelengths of 254 nm, 270 nm and 365 nm, respectively, revealed that treatment with UVC-LEDs were comparable to, or better than those observed using the mercury lamp. Bacterial reductions in the seasoning powders with UVC-LEDs were less than in the surface inactivation experiments, but significant reductions of 0.75-3 log10 colony forming units (CFU) were obtained following longer (40 s) UVC-LED exposure times. Inactivation kinetics were generally nonlinear, and a comparison of the predictive models highlighted that microbial inactivation was dependent on the combination of powder and microorganism. This study is the first to report on the efficacy of UV-LEDs for the inactivation of several different bacterial species in a variety of powdered ingredients, highlighting the potential of the technology as an alternative to the traditional UV lamps used in the food industry.

Thermal or membrane processing for Infant Milk Formula: Effects on protein digestion and integrity of the intestinal barrier.

Infant Milk Formula (IMF) is designed as a breastmilk substitute to satisfy the nutritional requirements during the first months of life. This study investigates the effects of two IMF processing technologies on cow milk protein digestion using an infant static in vitro gastrointestinal model. The degree of protein hydrolysis at the end of the gastric phase was 3.7-fold higher for IMF produced by high temperature (IMF-HT), compared to IMF produced by cascade membrane filtration (IMF-CMF), as assessed by free N-terminal group analysis. The processing type also influenced the panel of bioavailable peptides detected in basolateral compartments of Caco-2 monolayers exposed to gastrointestinal digested IMFs. In addition, IMF-CMF significantly increased tight junction protein, claudin 1, whilst IMF-HT significantly reduced tight junction integrity. In conclusion, producing IMF by CMF may preserve intestinal barrier integrity and can deliver its own unique inventory of bioavailable peptides with potential bioactivity.

Comparison of predicted and impedance determined growth of Listeria innocua in complex food matrices.

Indirect impedance has been used for the detection and enumeration of bacteria, however there is limited data regarding the ability of the method to measure growth and inhibition of microorganisms in food in response to preservatives. The aim of this study was to evaluate the suitability of the technique to determine maximum growth rates of Listeria innocua (used as a surrogate for Listeria monocytogenes) in complex food matrices to which multiple preservative factors had been applied and assess the suitability of the data for use in predictive microbiology. Growth of L. innocua in laboratory medium (BHI broth) and two food matrices (zucchini purée and béarnaise sauce) under varying conditions of pH (5 & 5.3), water activity (0.93, 0.96 & 0.98) and acetic and propionic acid concentration (0, 1 & 2 mM) was monitored by the conductimetric Rapid Automated Bacterial Impedance Technology (R.A.B.I.T) system by means of CO2 emission for up to 120 h. Growth rates of L. innocua were determined for several conditions across the three test matrices and a good correlation between detection times and initial inoculum level was observed in most cases (R2 ≥ 0.82). However, growth of L. innocua was not detected in a large number of conditions and comparison of growth rates determined by indirect impedance to those determined by plate counts indicated that in general, the R.A.B.I.T. system under-estimated growth. This study demonstrates that there are limitations associated with the technology, and as a result the system may be unsuitable for measuring microbial growth rates in complex food matrices under the environmental conditions tested and within the time duration of the study.

Superior esterolytic activity in environmental Lactococcus lactis strains is linked to the presence of the SGNH hydrolase family of esterases.

Lactococcus lactis strains are widely used in the dairy industry in fermentation processes for production of cheese and fermented milks. However, the esterolytic activity of L. lactis is not generally considered high. For this reason, purified microbial lipases and esterases are often added in certain dairy processes to generate specific flavors in the final food product. This work demonstrates the superior esterolytic activity of a collection of L. lactis strains isolated from different environmental sources compared with that of dairy-derived strains. It provides further evidence of the more diverse metabolic capabilities displayed by L. lactis strains from environmental sources compared to their domesticated dairy counterparts. Furthermore, the presence of a 1,287-bp gene encoding a 428-amino acid SGNH hydrolase in the high-esterolytic environmental strains suggests a possible link between superior esterolytic activity and the presence of the esterase from the SGNH hydrolase family.

Whey proteins: targets of oxidation, or mediators of redox protection.

Bovine whey proteins are highly valued dairy ingredients. This is primarily due to their amino acid content, digestibility, bioactivities and their processing characteristics. One of the reported bioactivities of whey proteins is antioxidant activity. Numerous dietary intervention trials with humans and animals indicate that consumption of whey products can modulate redox biomarkers to reduce oxidative stress. This bioactivity has in part been assigned to whey peptides using a range of biochemical or cellular assays in vitro. Superimposing whey peptide sequences from gastrointestinal samples, with whey peptides proven to be antioxidant in vitro, allows us to propose peptides from whey likely to exhibit antioxidant activity in the diet. However, whey proteins themselves are targets of oxidation during processing particularly when exposed to high thermal loads and/or extensive processing (e.g. infant formula manufacture). Oxidative damage of whey proteins can be selective with regard to the residues that are modified and are associated with the degree of protein unfolding, with α-Lactalbumin more susceptible than ß-Lactoglobulin. Such oxidative damage may have adverse effects on human health. This review summarises how whey proteins can modulate cellular redox pathways and conversely how whey proteins can be oxidised during processing. Given the extensive processing steps that whey proteins are often subjected to, we conclude that oxidation during processing is likely to compromise the positive health attributes associated with whey proteins.

Draft Genome Sequences of Four Lactococcus lactis Strains Isolated from Diverse Niches, Including Dairy Products, Grass, and Green Peas.

Lactococcus lactis has been used for millennia as a starter organism in the production of many fermented dairy products. This announcement includes the draft genome sequences of four strains of Lactococcus lactis, two of dairy origin and two from nondairy sources.

Exploring the Impacts of Postharvest Processing on the Microbiota and Metabolite Profiles during Green Coffee Bean Production.

The postharvest treatment and processing of fresh coffee cherries can impact the quality of the unroasted green coffee beans. In the present case study, freshly harvested Arabica coffee cherries were processed through two different wet and dry methods to monitor differences in the microbial community structure and in substrate and metabolite profiles. The changes were followed throughout the postharvest processing chain, from harvest to drying, by implementing up-to-date techniques, encompassing multiple-step metagenomic DNA extraction, high-throughput sequencing, and multiphasic metabolite target analysis. During wet processing, a cohort of lactic acid bacteria (i.e., Leuconostoc, Lactococcus, and Lactobacillus) was the most commonly identified microbial group, along with enterobacteria and yeasts (Pichia and Starmerella). Several of the metabolites associated with lactic acid bacterial metabolism (e.g., lactic acid, acetic acid, and mannitol) produced in the mucilage were also found in the endosperm. During dry processing, acetic acid bacteria (i.e., Acetobacter and Gluconobacter) were most abundant, along with Pichia and non-Pichia (Candida, Starmerella, and Saccharomycopsis) yeasts. Accumulation of associated metabolites (e.g., gluconic acid and sugar alcohols) took place in the drying outer layers of the coffee cherries. Consequently, both wet and dry processing methods significantly influenced the microbial community structures and hence the composition of the final green coffee beans. This systematic approach to dissecting the coffee ecosystem contributes to a deeper understanding of coffee processing and might constitute a state-of-the-art framework for the further analysis and subsequent control of this complex biotechnological process. IMPORTANCE: Coffee production is a long process, starting with the harvest of coffee cherries and the on-farm drying of their beans. In a later stage, the dried green coffee beans are roasted and ground in order to brew a cup of coffee. The on-farm, postharvest processing method applied can impact the quality of the green coffee beans. In the present case study, freshly harvested Arabica coffee cherries were processed through wet and dry processing in four distinct variations. The microorganisms present and the chemical profiles of the coffee beans were analyzed throughout the postharvest processing chain. The up-to-date techniques implemented facilitated the investigation of differences related to the method applied. For instance, different microbial groups were associated with wet and dry processing methods. Additionally, metabolites associated with the respective microorganisms accumulated on the final green coffee beans.

Complete Genome Sequence of vB_EcoM_112, a T-Even-Type Bacteriophage Specific for Escherichia coli O157:H7.

Bacteriophage vB_EcoM_112 (formerly e11/2) is an Escherichia coli phage with specificity for the O157:H7 serotype. The vB_EcoM_112 genome sequence shares high degrees of similarity with the phage T4 genome sequence.

In situ investigation of Geobacillus stearothermophilus spore germination and inactivation mechanisms under moderate high pressure.

Bacterial spores are a major concern for food safety due to their high resistance to conventional preservation hurdles. Innovative hurdles can trigger bacterial spore germination or inactivate them. In this work, Geobacillus stearothermophilus spore high pressure (HP) germination and inactivation mechanisms were investigated by in situ infrared spectroscopy (FT-IR) and fluorometry. G. stearothermophilus spores' inner membrane (IM) was stained with Laurdan fluorescent dye. Time-dependent FT-IR and fluorescence spectra were recorded in situ under pressure at different temperatures. The Laurdan spectrum is affected by the lipid packing and level of hydration, and provided information on the IM state through the Laurdan generalized polarization. Changes in the -CH2 and -CH3 asymmetric stretching bands, characteristic of lipids, and in the amide I' band region, characteristic of proteins' secondary structure elements, enabled evaluation of the impact of HP on endospores lipid and protein structures. These studies were complemented by ex situ analyses (plate counts and microscopy). The methods applied showed high potential to identify germination mechanisms, particularly associated to the IM. Germination up to 3 log10 was achieved at 200 MPa and 55 °C. A molecular-level understanding of these mechanisms is important for the development and validation of multi-hurdle approaches to achieve commercial sterility.

(Ultra) high pressure homogenization for continuous high pressure sterilization of pumpable foods - a review.

Bacterial spores have a strong resistance to both chemical and physical hurdles and create a risk for the food industry, which has been tackled by applying high thermal intensity treatments to sterilize food. These strong thermal treatments lead to a reduction of the organoleptic and nutritional properties of food and alternatives are actively searched for. Innovative hurdles offer an alternative to inactivate bacterial spores. In particular, recent technological developments have enabled a new generation of high pressure homogenizer working at pressures up to 400 MPa and thus, opening new opportunities for high pressure sterilization of foods. In this short review, we summarize the work conducted on (ultra) high pressure homogenization (U)HPH to inactivate endospores in model and food systems. Specific attention is given to process parameters (pressure, inlet, and valve temperatures). This review gathers the current state of the art and underlines the potential of UHPH sterilization of pumpable foods while highlighting the needs for future work.

Association of bovine leptin polymorphisms with energy output and energy storage traits in progeny tested Holstein-Friesian dairy cattle sires.

BACKGROUND: Leptin modulates appetite, energy expenditure and the reproductive axis by signalling via its receptor the status of body energy stores to the brain. The present study aimed to quantify the associations between 10 novel and known single nucleotide polymorphisms in genes coding for leptin and leptin receptor with performance traits in 848 Holstein-Friesian sires, estimated from performance of up to 43,117 daughter-parity records per sire. RESULTS: All single nucleotide polymorphisms were segregating in this sample population and none deviated (P > 0.05) from Hardy-Weinberg equilibrium. Complete linkage disequilibrium existed between the novel polymorphism LEP-1609, and the previously identified polymorphisms LEP-1457 and LEP-580. LEP-2470 associated (P < 0.05) with milk protein concentration and calf perinatal mortality. It had a tendency to associate with milk yield (P < 0.1). The G allele of LEP-1238 was associated (P < 0.05) with reduced milk fat concentration, reduced milk protein concentration, longer gestation length and tended to associate (P < 0.1) with an increase in calving difficulty, calf perinatal mortality and somatic cells in the milk. LEP-963 exhibited an association (P < 0.05) with milk fat concentration, milk protein concentration, calving difficulty and gestation length. It also tended to associate with milk yield (P < 0.1). The R25C SNP associated (P < 0.05) with milk fat concentration, milk protein concentration, calving difficulty and length of gestation. The T allele of the Y7F SNP significantly associated with reduced angularity (P < 0.01) and reduced milk protein yield (P < 0.05). There was also a tendency (P < 0.1) for Y7F to associate with increased body condition score, reduced milk yield and shorter gestation (P < 0.1). A80V associated with reduced survival in the herd (P < 0.05). CONCLUSIONS: Several leptin polymorphisms (LEP-2470, LEP-1238, LEP-963, Y7F and R25C) associated with the energetically expensive process of lactogenesis. Only SNP Y7F associated with energy storage. Associations were also observed between leptin polymorphisms and calving difficulty, gestation length and calf perinatal mortality. The lack of an association between the leptin variants investigated with calving interval in this large data set would question the potential importance of these leptin variants, or indeed leptin, in selection for improved fertility in the Holstein-Friesian dairy cow.

Exploitation of the diverse insertion sequence element content of dairy Lactobacillus helveticus starters as a rapid method to identify different strains.

The species Lactobacillus helveticus is a commonly used thermophilic starter and/or adjunct culture for Swiss and Cheddar cheese manufacture. Its use is normally associated with flavour improvement which is known to be associated with culture traits such as rapid autolysis and high proteolytic activity. The genome of the commercial strain, DPC4571, was recently sequenced and found to have an abundance of IS sequences in terms of both abundance (213 intact) and diversity (21 types). Given this unique diversity for a lactic acid bacterium, we investigated whether PCR-based IS fingerprinting could be used as a discriminatory tool to distinguish between different strains of Lb. helveticus. A set of ten primers targeting five of the most numerous groups (ISL1201, ISLhe65, ISLhe2, ISLhe15 and ISL2) of IS elements was designed. Multiplex-PCR with all primers resulted in 1-12 discreet amplicons for each strain tested. The resultant fingerprints (in the 0.5 kb-3 kb range) were found to be strain specific and reproducible. This approach thus provides a valuable method to distinguish between Lb. helveticus strains while giving some indication of the relative abundance of IS sequences in each strain.