Optimisation of the food dairy coop supply chain

In the European Union under the Common Agricultural Policy (CAP) milk production was restricted by milk quotas since 1984. However, due to recent changes in the Common Agricultural Policy (CAP), milk quotas will be abolished by 2015. Therefore, the European dairy sector will soon face an opportunity, for the first time in a generation, to expand. Numerous studies have shown that milk production in Ireland will increase significantly post quotas (Laepple and Hennessy (2010), Donnellan and Hennessy (2007) and Lips and Reider (2005)). The research in this thesis explored milk transport and dairy product processing in the Irish dairy processing sector in the context of milk quota removal and expansion by 2020. In this study a national milk transport model was developed for the Irish dairy industry, the model was used to examine different efficiency factors in milk transport and to estimate milk transport costs post milk quota abolition. Secondly, the impact of different milk supply profiles on milk transport costs was investigated using the milk transport model. Current processing capacity in Ireland was compared against future supply, it was concluded that additional milk processing capacity would not be sufficient to process the additional milk. Thirdly, the milk transport model was used to identify the least cost locations (based on transport costs) to process the additional milk supply in 2020. Finally, an optimisation model was developed to identify the optimum configuration for the Irish dairy processing sector in 2020 taking cognisance of increasing transport costs and decreasing processing costs.

Investigation of polyhydroxyalkanoate production by an activated sludge microbial consortium treating artificial dairy wastewater

Petrochemical plastics/polymers are a common feature of day to day living as they occur in packaging, furniture, mobile phones, computers, construction equipment etc. However, these materials are produced from non-renewable materials and are resistant to microbial degradation in the environment. Considerable research has therefore been carried out into the production of sustainable, biodegradable polymers, amenable to microbial catabolism to CO2 and H2O. A key group of microbial polyesters, widely considered as optimal replacement polymers, are the Polyhydroxyalkaonates (PHAs). Primary research in this area has focused on using recombinant pure cultures to optimise PHA yields, however, despite considerable success, the high costs of pure culture fermentation have thus far hindered the commercial viability of PHAs thus produced. In more recent years work has begun to focus on mixed cultures for the optimisation of PHA production, with waste incorporations offering optimal production cost reductions. The scale of dairy processing in Ireland, and the high organic load wastewaters generated, represent an excellent potential substrate for bioconversion to PHAs in a mixed culture system. The current study sought to investigate the potential for such bioconversion in a laboratory scale biological system and to establish key operational and microbial characteristics of same. Two sequencing batch reactors were set up and operated along the lines of an enhanced biological phosphate removal (EBPR) system, which has PHA accumulation as a key step within repeated rounds of anaerobic/aerobic cycling. Influents to the reactors varied only in the carbon sources provided. Reactor 1 received artificial wastewater with acetate alone, which is known to be readily converted to PHA in the anaerobic step of EBPR. Reactor 2 wastewater influent contained acetate and skim milk to imitate a dairy processing effluent. Chemical monitoring of nutrient remediation within the reactors as continuously applied and EBPR consistent performances observed. Qualitative analysis of the sludge was carried out using fluorescence microscopy with Nile Blue A lipophillic stain and PHA production was confirmed in both reactors. Quantitative analysis via HPLC detection of crotonic acid derivatives revealed the fluorescence to be short chain length Polyhydroxybutyrate, with biomass dry weight accumulations of 11% and 13% being observed in reactors 1 and 2, respectively. Gas Chromatography-Mass Spectrometry for medium chain length methyl ester derivatives revealed the presence of hydroxyoctanoic, -decanoic and -dodecanoic acids in reactor 1. Similar analyses in reactor 2 revealed monomers of 3-hydroxydodecenoic and 3-hydroxytetradecanoic acids. Investigation of the microbial ecology of both reactors as conducted in an attempt to identify key species potentially contributing to reactor performance. Culture dependent investigations indicated that quite different communities were present in both reactors. Reactor 1 isolates demonstrated the following species distributions Pseudomonas (82%), Delftia acidovorans (3%), Acinetobacter sp. (5%) Aminobacter sp., (3%) Bacillus sp. (3%), Thauera sp., (3%) and Cytophaga sp. (3%). Relative species distributions among reactor 2 profiled isolates were more evenly distributed between Pseudoxanthomonas (32%), Thauera sp (24%), Acinetobacter (24%), Citrobacter sp (8%), Lactococcus lactis (5%), Lysinibacillus (5%) and Elizabethkingia (2%). In both reactors Gammaproteobacteria dominated the cultured isolates. Culture independent 16S rRNA gene analyses revealed differing profiles for both reactors. Reactor 1 clone distribution was as follows; Zooglea resiniphila (83%), Zooglea oryzae (2%), Pedobacter composti (5%), Neissericeae sp. (2%) Rhodobacter sp. (2%), Runella defluvii (3%) and Streptococcus sp. (3%). RFLP based species distribution among the reactor 2 clones was as follows; Runella defluvii (50%), Zoogloea oryzae (20%), Flavobacterium sp. (9%), Simplicispira sp. (6%), Uncultured Sphingobacteria sp. (6%), Arcicella (6%) and Leadbetterella bysophila (3%). Betaproteobacteria dominated the 16S rRNA gene clones identified in both reactors. FISH analysis with Nile Blue dual staining resolved these divergent findings, identifying the Betaproteobacteria as dominant PHA accumulators within the reactor sludges, although species/strain specific allocations could not be made. GC analysis of the sludge had indicated the presence of both medium chain length as well short chain length PHAs accumulating in both reactors. In addition the cultured isolates from the reactors had been identified previously as mcl and scl PHA producers, respectively. Characterisations of the PHA monomer profiles of the individual isolates were therefore performed to screen for potential novel scl-mcl PHAs. Nitrogen limitation driven PHA accumulation in E2 minimal media revealed a greater propensity among isoates for mcl-pHA production. HPLC analysis indicated that PHB production was not a major feature of the reactor isolates and this was supported by the low presence of scl phaC1 genes among PCR screened isolates. A high percentage distribution of phaC2 mcl-PHA synthase genes was recorded, with the majority sharing high percentage homology with class II synthases from Pseudomonas sp. The common presence of a phaC2 homologue was not reflected in the production of a common polymer. Considerable variation was noted in both the monomer composition and ratios following GC analysis. While co-polymer production could not be demonstrated, potentially novel synthase substrate specificities were noted which could be exploited further in the future.

Dairy-Met: compositional metagenomic analysis of milk and cheeses

Unpasteurised milk and many cheeses contain a diverse microbiological population. These microorganisms play important roles in dairy foods and can, for example, contribute to the development of flavours and aromas, determine safety, cause spoilage or enhance the health of the consumer. It is thus important to understand thoroughly the microorganisms present in these food types. Traditional culture dependent and culture-independent methods have provided much detail regarding the microbial content of dairy foods. However, the development of next-generation DNA sequencing technologies has revolutionised our knowledge of complex microbial environments. Throughout this thesis we observe the benefits of applying these technologies to provide a detailed understanding of the bacterial content of dairy foods, including those present in milk pre- and post-pasteurisation, Irish farmhouse cheeses and commercially produced cheeses which encounter a discolouration defect, as well as to study genomic changes in microbes associated with dairy foods. Through the application of these state-of-the-art technologies we identified the presence of microorganisms not previously associated with dairy foods.

Exploring alternatives for milk quality improvement and more efficient dairy production in a smallholder farming context – case study: Mantaro Valley (Peru)

Consumer demand is revolutionizing the way products are being produced, distributed and marketed. In relation to the dairy sector in developing countries, aspects of milk quality are receiving more attention from both society and the government. However, milk quality management needs to be better addressed in dairy production systems to guarantee the access of stakeholders, mainly small-holders, into dairy markets. The present study is focused on an analysis of the interaction of the upstream part of the dairy supply chain (farmers and dairies) in the Mantaro Valley (Peruvian central Andes), in order to understand possible constraints both stakeholders face implementing milk quality controls and practices; and evaluate “ex-ante” how different strategies suggested to improve milk quality could affect farmers and processors’ profits. The analysis is based on three complementary field studies conducted between 2012 and 2013. Our work has shown that the presence of a dual supply chain combining both formal and informal markets has a direct impact on dairy production at the technical and organizational levels, affecting small formal dairy processors’ possibilities to implement contracts, including agreements on milk quality standards. The analysis of milk quality management from farms to dairy plants highlighted the poor hygiene in the study area, even when average values of milk composition were usually high. Some husbandry practices evaluated at farm level demonstrated cost effectiveness and a big impact on hygienic quality; however, regular application of these practices was limited, since small-scale farmers do not receive a bonus for producing hygienic milk. On the basis of these two results, we co-designed with formal small-scale dairy processors a simulation tool to show prospective scenarios, in which they could select their best product portfolio but also design milk payment systems to reward farmers’ with high milk quality performances. This type of approach allowed dairy processors to realize the importance of including milk quality management in their collection and manufacturing processes, especially in a context of high competition for milk supply. We concluded that the improvement of milk quality in a smallholder farming context requires a more coordinated effort among stakeholders. Successful implementation of strategies will depend on the willingness of small-scale dairy processors to reward farmers producing high milk quality; but also on the support from the State to provide incentives to the stakeholders in the formal sector.

Bioengineering of nisin to enhance functionality against dairy pathogens

The bacteriocin class of antimicrobial peptides have emerged as a viable alternative to at least partially fill the void created by the end of the golden age of antibiotic discovery. Along with this potential use in a clinical setting, bacteriocins also play an important role as bio-preservatives in the food industry. This thesis focuses on a specific bacteriocin group, the lantibiotics (Lanthionine-containing antibiotics). Their numerous methods of appliance in a food setting and how their gene-encoded nature can be modified to improve on overall bioactivity and functionality are explored here. The use of a lantibiotic (lacticin 3147) producing starter culture to control the Crohn’s disease-linked pathogen Mycobacterium paratuberculosis was assessed in a raw milk cheese. Although lacticin 3147 production did not effectively control the pathogen, the study provided an impetus to employ a variety of PCR-based mutagenesis techniques with a view to the creation of enhanced lantibiotic derivatives. Through the use of these techniques, a number of enhanced derivatives were generated from the ‘hinge’ region of the nisin peptide. Furthermore, a derivative in which the three hinge amino acids were replaced with three alanines represents the first enhanced derivative of nisin to have been designed through a rational process. This derivative also formed the backbone for the creation of an active, trypsin resistant, variant. Through the employment of further mutagenesis methods a derivative was created with potential use as an oral anti-bacterial in the future. Finally a number of lead nisin derivatives were investigated to assess their anti- Streptococcus agalactiae ability, a mastitis associated pathogen. Also a system was developed to facilitate the large scale production of these candidates, or other nisin derivatives, from dairy substrates.

The effects of formulation and processing on surface characteristics and functional properties of dairy powders

The objectives of this thesis were to (i) study the effect of increasing protein concentration in milk protein concentrate (MPC) powders on surface composition and sorption properties; (ii) examine the effect of increasing protein content on the rehydration properties of MPC; (iii) study the physicochemical properties of spraydried emulsion-containing powders having different water and oil contents; (iv) analyse the effect of protein type on water sorption and diffusivity properties in a protein/lactose dispersion, and; (v) characterise lactose crystallisation and emulsion stability of model infant formula containing intact or hydrolysed whey proteins. Surface composition of MPC powders (protein contents 35 - 86 g / 100 g) indicated that fat and protein were preferentially located on the surface of powders. Low protein powder (35 g / 100 g) exhibited lactose crystallisation, whereas powders with higher protein contents did not, due to their high protein: lactose ratio. Insolubility was evident in high protein MPCs and was primarily related to insolubility of the casein fraction. High temperature (50 °C) was required for dissolution of high protein MPCs (protein content > 60 g / 100 g). The effect of different oil types and spray-drying outlet temperature on the physicochemical properties of the resultant fat-filled powders was investigated and showed that increasing outlet temperature reduced water content, water activity and tapped bulk density, irrespective of oil type, and increased solvent-extractable free fat for all oil types and onset of glass transition (Tg) and crystallisation (Tcr) temperature. Powder dispersions of protein/lactose (0.21:1), containing either intact or hydrolysed whey protein (12 % degree of hydrolysis; DH), were spray-dried at pilot scale. Moisture sorption analysis at 25 °C showed that dispersions containing intact whey protein exhibited lactose crystallisation at a lower relative humidity (RH). Dispersions containing hydrolysed whey protein had significantly higher (P < 0.05) water diffusivity. Finally, a spray-dried model infant formula was produced containing hydrolysed or intact whey as the protein with sunflower oil as the fat source. Reconstituted, hydrolysed formula had a significantly (P < 0.05) higher fat globule size and lower emulsion stability than intact formula. Lactose crystallisation in powders occurred at higher RH for hydrolysed formula. In conclusion, this research has shown the effect of altering the protein type, protein composition, and oil type on the surface composition and physical properties of different dairy powders, and how these variations greatly affect their rehydration characteristics and storage stability.

From field to fermentation: characterisation and application of non-dairy cultures in dairy foods

This thesis investigates the phenotypic and genotypic diversity of non-dairy L. lactis strains and their application to dairy fermentations. A bank of non-dairy lactococci were isolated from grass, vegetables and the bovine rumen. Subsequent analysis of these L. lactis strains revealed seven strains to possess cremoris genotypes which did not correlate with their observed phenotypes. Multi-locus sequence typing (MLST) and average nucleotide identity (ANI) highlighted the genetic diversity of lactis and cremoris subspecies. The application of these non-dairy lactococci to cheese production was also assessed. In milk, non-dairy strains formed diverse volatile profiles and selected strains were used as adjuncts in a mini Gouda-type cheese system. Sensory analysis showed non-dairy strains to be strongly associated with the development of off-flavours and bitterness. However, microfluidisation appeared to reduce bitterness. A novel bacteriophage, ɸL47, was isolated using the grass isolate L. lactis ssp. cremoris DPC6860 as a host. The phage, a member of the Siphoviridae, possessed a long tail fiber, previously unseen in dairy lactococcal phages. Genome sequencing revealed ɸL47 to be the largest sequenced lactococcal phage to date and owing to the high % similarity with ɸ949, a second member of the 949 group. Finally, to identify and characterise specific genes which may be important in niche adaptation and for applications to dairy fermentations, comparative genome sequence analysis was performed on L. lactis from corn (DPC6853), the bovine rumen (DPC6853) and grass (DPC6860). This study highlights the contribution of niche specialisation to the intra-species diversity of L. lactis and the adaptation of this organism to different environments. In summary this thesis describes the genetic diversity of L. lactis strains from outside the dairy environment and their potential application in dairy fermentations.