How Can We Advance Integrative Biology Research in Animal Science in 21st Century? Experience at University of Ljubljana from 2002 to 2022.

In this perspective analysis, we strive to answer the following question: how can we advance integrative biology research in the 21st century with lessons from animal science? At the University of Ljubljana, Biotechnical Faculty, Department of Animal Science, we share here our three lessons learned in the two decades from 2002 to 2022 that we believe could inform integrative biology, systems science, and animal science scholarship in other countries and geographies. Cultivating multiomics knowledge through a conceptual lens of integrative biology is crucial for life sciences research that can stand the test of diverse biological, clinical, and ecological contexts. Moreover, in an era of the current COVID-19 pandemic, animal nutrition and animal science, and the study of their interactions with human health (and vice versa) through integrative biology approaches hold enormous prospects and significance for systems medicine and ecosystem health.

Evaluation of different primers for PCR-DGGE analysis of cheese-associated enterococci.

Enterococci represent an important part of bacterial microbiota in different types of artisanal cheeses, made from either raw or pasteurized milk. Polymerase chain reaction denaturing gradient gel electrophoresis (PCR-DGGE) of ribosomal DNA is currently one of the most frequently used fingerprinting method to study diversity and dynamics of microbial communities and also a tool for microbial identification. Among several primer pairs for DGGE analysis published so far, six primer pairs amplifying different variable regions of 16S rDNA were selected and applied in our DGGE analysis of 12 species belonging to genus Enterococcus and eight other bacterial species often found in cheeses (seven lactobacilli and one Lactoccocus lactis). When DGGE procedures were optimized, the same set of primers was used for DGGE analysis of five cheese samples. Our study demonstrates that the use of different primer pairs generate significant differences in DGGE analysis of enterococcal population, consequently, appropriate primers regarding the purpose of analysis can be selected. For differentiation and identification of pure enterococcal isolates, primer pair P1V1/P2V1 showed the most promising results since all 12 enterococcal isolates gave distinctive DGGE fingerprints, but with multiple bands patterns; therefore, these primers do not seem to be appropriate for identification of enterococcal species in mixed cultures. Use of primer pairs HDA1/HDA2 and V3f/V3r amplifying V3 region showed better potential for detection and identification of enterococci in mixed communities, but since some bacterial species showed the same fingerprint, for clear identification combination of DGGE and some other method (e.g. species specific PCR) or combined DGGE analysis using two primer pairs generating distinctive results should be used.

Expression of the sweet-tasting plant protein brazzein in Escherichia coli and Lactococcus lactis: a path toward sweet lactic acid bacteria.

Brazzein is an intensely sweet-tasting plant protein with good stability, which makes it an attractive alternative to sucrose. A brazzein gene has been designed, synthesized, and expressed in Escherichia coli at 30 degrees C to yield brazzein in a soluble form and in considerable quantity. Antibodies have been produced using brazzein fused to His-tag. Brazzein without the tag was sweet and resembled closely the taste of its native counterpart. The brazzein gene was also expressed in Lactococcus lactis, using a nisin-controlled expression system, to produce sweet-tasting lactic acid bacteria. The low level of expression was detected with anti-brazzein antibodies. Secretion of brazzein into the medium has not led to significant yield increase. Surprisingly, optimizing the codon usage for Lactococcus lactis led to a decrease in the yield of brazzein.