Love in the Laboratory

This chapter describes the marriage of two prodigies and how it represented a fruitful alliance of complementary research personalities: the brilliant theoretician and the skillful experimenter. Esther Zimmer and Joshua Lederberg were two of the youngest scientists to attend the 1946 summer symposium at Cold Spring Harbor. Edward Tatum arranged for his protégé, young Lederberg, to present his stupendous discovery of bacterial conjugation, showing that bacteria could mate and recombine their genes. Zimmer and Lederberg began a short romance and married five months later. The young couple moved near the campus of Yale University, where Joshua wrote up his thesis and Esther researched Neurospora genetics with Norman Giles. The following summer, Tatum negotiated with Yale to grant an accelerated PhD to Joshua. The University of Wisconsin offered him an assistant professorship, and Joshua and Esther moved to Madison in 1947. There they established the first research program in bacterial genetics.

Behind the Laboratory Doors

This chapter compares Esther Lederberg’s role with that of other notable women scientists whose achievements exhibited creative laboratory skills. Esther’s career peaked in 1956 when the Society of Illinois Bacteriologists jointly bestowed the Pasteur Medal on the Lederberg couple. Usually, Joshua Lederberg was the public face of their research program. Esther’s place was behind the laboratory doors where she managed the lab and performed the experiments. For over a hundred years, this was the typical arrangement for women and their male associates. Prestigious faculty positions and accolades were unattainable for so many women in science. For Esther and many of her female colleagues, the thrill of discovery was enough reward. Esther valued the camaraderie of the brilliant personalities that made up the circle of pioneering researchers. Stanley Falkow called her a kind of Boswell of bacterial genetics. Her extensive photographic collection is a who’s who of molecular biology, many as their younger selves.

Introduction

This chapter relates how one day in 1950, Esther Zimmer Lederberg cleverly re-purposed her compact makeup pad and invented replica plating. This whimsical experiment led to an elegant technique for duplicating many bacterial clones in one step, a clever invention that epitomized her experimental creativity. The chapter shows how the Lederbergs established the field of bacterial genetics years before the birth of molecular biology and together discovered bacterial sex (or horizontal gene transfer, HGT) the peculiar processes that enable bacteria to rapidly spread their genes, leading to antibiotic resistance and the evolution of new species. The stellar reputation of her brilliant husband and collaborator, however, diminished Esther Lederberg’s legacy. The systematic bias against giving due credit for achievements of women scientists whose work is misattributed to their scientific colleagues is known as the Matilda Effect. Esther Lederberg’s story is sadly similar to those of many exemplary women scientists.

The Anomaly of Bacterial Genetics

This chapter reviews the research that set the stage for Joshua Lederberg’s surprising discovery of bacterial conjugation. While the foundational research of Gregor Mendel and his principles of inheritance had been effectively combined with Darwinian evolution, producing the Modern Synthesis in the mid-forties, bacteria did not fit into this grand synthesis. Most biologists believed that bacteria were too primitive to have real genes. But Delbruck, Hershey and Luria organized the Phage School, leading a novel approach to discovering the molecular biology of the gene by studying bacteriophages. Microbiologists like Tracy Sonneborn and Carl Lindegren turned to alternative microorganisms—protists, fungi, and yeast—to develop new model systems that offered advantages over the classical genetics organisms of animals and plants. The research of Edward Tatum and Jacques Monod indicated that mutations seemed to explain variation in bacteria. For many years, however, bacteriologists had known that bacteria reproduced by fission. The lack of any genetic hybridization seemed to argue against using bacteria to study basic genetic processes.

Direct cobamide remodeling via additional function of cobamide biosynthesis protein CobS from Vibrio cholerae

Vitamin B12 belongs to a family of structurally-diverse cofactors with over a dozen natural analogs, collectively referred to as cobamides. Most bacteria encode cobamide-dependent enzymes, many of which can only utilize a subset of cobamide analogs. Some bacteria employ a mechanism called cobamide remodeling, a process in which cobamides are converted into other analogs, to ensure that compatible cobamides are available in the cell. Here we characterize an additional pathway for cobamide remodeling that is distinct from the previously-characterized ones. Cobamide synthase (CobS) is an enzyme required for cobamide biosynthesis that attaches the lower ligand moiety in which the base varies between analogs. In a heterologous model system, we previously showed Vibrio cholerae CobS (VcCobS) unexpectedly conferred remodeling activity, in addition to performing the known cobamide biosynthesis reaction. Here we show that additional Vibrio species perform the same remodeling reaction, and we further characterize VcCobS-mediated remodeling using bacterial genetics and in vitro assays. We demonstrate that VcCobS acts upon the cobamide pseudocobalamin directly to remodel it, a mechanism which differs from the known remodeling pathways in which cobamides are first cleaved into biosynthetic intermediates. This suggests that some CobS homologs have the additional function of cobamide remodeling, and we propose the term "direct remodeling" for this process. This characterization of yet another pathway for remodeling suggests that cobamide profiles are highly dynamic in polymicrobial environments, with remodeling pathways conferring a competitive advantage. Importance Cobamides are widespread cofactors that mediate metabolic interactions in complex microbial communities. Few studies directly examine cobamide profiles, but several have shown that mammalian gastrointestinal tracts are rich in cobamide analogs. Studies of intestinal bacteria, including beneficial commensals and pathogens, show variation in the ability to produce and utilize different cobamides. Some bacteria can convert imported cobamides into compatible analogs in a process called remodeling. Recent discoveries of additional cobamide remodeling pathways, including this work, suggest that remodeling is an important factor in cobamide dynamics. Characterization of such pathways is critical in understanding cobamide flux and nutrient cross-feeding in polymicrobial communities, and facilitates the establishment of microbiome manipulation strategies via modulation of cobamide profiles.

Acinetobacter baylyi ADP1—naturally competent for synthetic biology

Acinetobacter baylyi ADP1 is a non-pathogenic soil bacterium known for its metabolic diversity and high natural transformation and recombination efficiency. For these features, A. baylyi ADP1 has been long exploited in studying bacterial genetics and metabolism. The large pool of information generated in the fundamental studies has facilitated the development of a broad range of sophisticated and robust tools for the genome and metabolic engineering of ADP1. This mini-review outlines and describes the recent advances in ADP1 engineering and tool development, exploited in, for example, pathway and enzyme evolution, genome reduction and stabilization, and for the production of native and non-native products in both pure and rationally designed multispecies cultures. The rapidly expanding toolbox together with the unique features of A. baylyi ADP1 provide a strong base for a microbial cell factory excelling in synthetic biology applications where evolution meets rational engineering.

Genetics bacterial teaching materials development based on flipbook in microbiology subject to improve learning motivation

Microbiology is the study of microbes where one of the materials is bacterial genetics. The results of observations about microbiology learning students experienced difficulties in understanding bacterial genetics, and there was no teaching material on bacterial genetics. Flipbook-based teaching materials are an alternative solution for teaching bacterial genetics well to students. The purpose of this study was to develop flipbook-based bacterial genetics teaching materials in microbiology courses at IKIP Budi Utomo. The development research model used is 4D including define, design, develop, and disseminate. This research is limited to the development stage. The results showed that the assessment of bacterial genetics teaching materials from material experts was 90.38%, media experts were 90%, field practitioner tests were 91.67% so that it met the very worthy criteria. The readability test with most of the students answered strongly agree with the statements about bacterial genetics teaching materials. The flipbook-based bacterial genetics teaching materials in the Microbiology course can improve learning motivation.Keywords: Learning materials, bacterial genetics, flipbook maker

High-quality genome sequence assembly of R.A73 Enterococcus faecium isolated from freshwater fish mucus

Background Whole-genome sequencing using high throughput technologies has revolutionized and speeded up the scientific investigation of bacterial genetics, biochemistry, and molecular biology. Lactic acid bacteria (LABs) have been extensively used in fermentation and more recently as probiotics in food products that promote health. Genome sequencing and functional genomics investigations of LABs varieties provide rapid and important information about their diversity and their evolution, revealing a significant molecular basis. This study investigated the whole genome sequences of the Enterococcus faecium strain (HG937697), isolated from the mucus of freshwater fish in Tunisian dams. Genomic DNA was extracted using the Quick-GDNA kit and sequenced using the Illumina HiSeq2500 system. Sequences quality assessment was performed using FastQC software. The complete genome annotation was carried out with the Rapid Annotation using Subsystem Technology (RAST) web server then NCBI PGAAP. Results The Enterococcus faecium R.A73 assembled in 28 contigs consisting of 2,935,283 bps. The genome annotation revealed 2884 genes in total including 2834 coding sequences and 50 RNAs containing 3 rRNAs (one rRNA 16 s, one rRNA 23 s and one rRNA 5 s) and 47 tRNAs. Twenty-two genes implicated in bacteriocin production are identified within the Enterococcus faecium R.A73 strain. Conclusion Data obtained provide insights to further investigate the effective strategy for testing this Enterococcus faecium R.A73 strain in the industrial manufacturing process. Studying their metabolism with bioinformatics tools represents the future challenge and contribution to improving the utilization of the multi-purpose bacteria in food.

High-Quality Genome Sequence Assembly of R.A73 Enterococcus faecium Isolated from Freshwater fish mucus

Background: Whole-genome sequencing using high throughput technologies has revolutionized and speeded up the scientific investigation of bacterial genetics, biochemistry, and molecular biology. Lactic acid bacteria (LABs) have been extensively used in fermentation and more recently as probiotics in food products that promote health. Genome sequencing and functional genomics investigations of LABs varieties provide rapid and important information about their diversity and their evolution, revealing a significant molecular basis.This study investigated the whole genome sequences of the Enterococcus faecium strain (HG937697), isolated from the mucus of freshwater fish in Tunisian dams. Genomic DNA was extracted using the Quick-GDNA kit and sequenced using the Illumina HiSeq2500 system. Sequences quality assessment was performed using FastQC software. The complete genome annotation was carried out with the Rapid Annotation using Subsystem Technology (RAST) web server then NCBI PGAAP. Results: The Enterococcus faecium R.A73 assembled in 28 contigs consisting of 2,935,283 bps. The genome annotation revealed 2,884 genes in total including 2,834 coding sequences and 50 RNAs containing 3 rRNAs (one rRNA 16s, one rRNA 23s and one rRNA 5s) and 47 tRNAs. Twenty-two genes implicated in bacteriocin production are identified within the Enterococcus faecium R.A73 strain. Conclusion: Data obtained provide insights to further investigate the effective strategy for testing this Enterococcus faecium R.A73 strain in the industrial manufacturing process. Studying their metabolism with bioinformatics tools represents the future challenge and contribution to improving the utilization of the multi-purpose bacteria in food.