Mitochondrial DNA D-Loop Amplification and Sequencing for Species Differentiation in Milk.

Adulteration of dairy products, mainly through the substitution of high-quality milk for lower-quality milk, results in the production of low-value products, raising health, social, and economic concerns. As such, the development of methods to ensure dairy products' safety and quality is of great concern for governments and consumers. Although several methods have been developed for species differentiation in dairy products, their application and the establishment of reliable molecular markers for authentication purposes still need to be improved. In this chapter, we describe a low-cost, sensitive, fast, and reliable PCR-based method for mitochondrial D-loop DNA amplification for efficient detection of cattle milk in binary mixtures with sheep milk, thereby allowing the authentication of processed dairy products.

Kluyveromyces marxianus as a microbial cell factory for lignocellulosic biomass valorisation.

The non-conventional yeast Kluyveromyces marxianus is widely used for several biotechnological applications, mainly due to its thermotolerance, high growth rate, and ability to metabolise a wide range of sugars. These cell traits are strategic for lignocellulosic biomass valorisation and strain diversity prompts the development of robust chassis, either with improved tolerance to lignocellulosic inhibitors or ethanol. This review summarises bioethanol and value-added chemicals production by K. marxianus from different lignocellulosic biomasses. Moreover, metabolic engineering and process optimization strategies developed to expand K. marxianus potential are also compiled, as well as studies reporting cell mechanisms to cope with lignocellulosic-derived inhibitors. The main lignocellulosic-based products are bioethanol, representing 71% of the reports, and xylitol, representing 17% of the reports. K. marxianus also proved to be a good chassis for lactic acid and volatile compounds production from lignocellulosic biomass, although the literature on this matter is still scarce. The increasing advances in genome editing tools and process optimization strategies will widen the K. marxianus-based portfolio products.

Establishment of Kluyveromyces marxianus as a Microbial Cell Factory for Lignocellulosic Processes: Production of High Value Furan Derivatives.

The establishment of lignocellulosic biorefineries is dependent on microorganisms being able to cope with the stressful conditions resulting from the release of inhibitory compounds during biomass processing. The yeast Kluyveromyces marxianus has been explored as an alternative microbial factory due to its thermotolerance and ability to natively metabolize xylose. The lignocellulose-derived inhibitors furfural and 5-hydroxymethylfurfural (HMF) are considered promising building-block platforms that can be converted into a wide variety of high-value derivatives. Here, several K. marxianus strains, isolated from cocoa fermentation, were evaluated for xylose consumption and tolerance towards acetic acid, furfural, and HMF. The potential of this yeast to reduce furfural and HMF at high inhibitory loads was disclosed and characterized. Our results associated HMF reduction with NADPH while furfural-reducing activity was higher with NADH. In addition, furans' inhibitory effect was higher when combined with xylose consumption. The furan derivatives produced by K. marxianus in different conditions were identified. Furthermore, one selected isolate was efficiently used as a whole-cell biocatalyst to convert furfural and HMF into their derivatives, furfuryl alcohol and 2,5-bis(hydroxymethyl)furan (BHMF), with high yields and productivities. These results validate K. marxianus as a promising microbial platform in lignocellulosic biorefineries.