Doctor of Philosophy

dissertationA diverse array of insect species harbor maternally transmitted mutualistic bacterial endosymbionts that perform a variety of functions within their hosts. Many of these associations are obligate in nature with the insect relying on the bacterial symbiont to provide nutrients that are lacking in the insect's natural diet. These obligate endosymbionts often show a highly reduced genome size and maintain only a small fraction of the gene inventory of free-living bacteria. Some of the smallest known bacterial genomes are from obligate endosymbionts that have been associated with their insect hosts for long periods of time. In addition to their small size, the genomes of ancient obligate symbionts also show an increased rate of DNA and polypeptide sequence evolution as well as a nucleotide composition bias that results in an increased ratio of adenine and thymine residues. Despite extensive study of these ancient endosymbionts, little is known about their origins. To address this issue and to better understand the forces shaping genomes in the early stages of an endosymbiotic association, this work focuses on two bacteria: strain HS, a recently characterized free-living bacterium that likely served as a progenitor to the Sodalis-allied clade of bacterial endosymbionts, and the Sitophilus oryzae primary endosymbiont (SOPE), a very recent established maternally transmitted obligate endosymbiont of the rice weevil. The complete genome sequencing of these two bacteria along with comparative genomic analyses revealed that SOPE has undergone a very rapid degeneration of its genome, losing nearly half of its coding capacity, a massive expansion of insertion sequence (IS) elements and numerous intragenomic rearrangements facilitated by the IS elements. Surprisingly, these changes have happened very recently since strain HS and SOPE shared a common ancestor approximately 28,000 years ago

Quorum Sensing Primes the Oxidative Stress Response in the Insect Endosymbiont, Sodalis glossinidius

quorum sensing system relies on the function of two regulatory proteins; SogI (a LuxI homolog) synthesizes a signaling molecule, characterized as N-(3-oxohexanoyl) homoserine lactone (OHHL), and SogR1 (a LuxR homolog) interacts with OHHL to modulate transcription of specific target genes. and SOPE. and SOPE indicates the potential for neofunctionalization to occur during the process of genome degeneration

The draft genome sequence and annotation of the desert woodrat Neotoma lepida

We present the de novo draft genome sequence for a vertebrate mammalian herbivore, the desert woodrat (Neotoma lepida). This species is of ecological and evolutionary interest with respect to ingestion, microbial detoxification and hepatic metabolism of toxic plant secondary compounds from the highly toxic creosote bush (Larrea tridentata) and the juniper shrub (Juniperus monosperma). The draft genome sequence and annotation have been deposited at GenBank under the accession LZPO01000000

Patterns of host gene expression associated with harboring a foregut microbial community

Abstract Background Harboring foregut microbial communities is considered a key innovation that allows herbivorous mammals to colonize new ecological niches. However, the functions of these chambers have only been well studied at the molecular level in ruminants. Here, we investigate gene expression in the foregut chamber of herbivorous rodents and ask whether these gene expression patterns are consistent with results in ruminants. We compared gene expression in foregut tissues of two rodent species: Stephen’s woodrat (Neotoma stephensi), which harbors a dense foregut microbial community, and the lab rat (Rattus norvegicus), which lacks such a community. Results We found that woodrats have higher abundances of transcripts associated with smooth muscle processes, specifically a higher expression of the smoothelin-like 1 gene, which may assist in contractile properties of this tissue to retain food material in the foregut chamber. The expression of genes associated with keratinization and cornification exhibited a complex pattern of differences between the two species, suggesting distinct molecular mechanisms. Lab rats exhibited higher abundances of transcripts associated with immune function, likely to inhibit microbial growth in the foregut of this species. Conclusions Some of our results were consistent with previous findings in ruminants (high expression of facilitative glucose transporters, lower expression of B4galnt2), suggestive of possible convergent evolution, while other results were unclear, and perhaps represent novel host-microbe interactions in rodents. Overall, our results suggest that harboring a foregut microbiota is associated with changes to the functions and host-microbe interactions of the foregut tissues

Retention of strain HS orthologs in <i>S. glossinidius</i> and SOPE according to COG functional category.

<p>The dark shaded component of each bar refers to intact genes retained in both <i>S. glossinidius</i> and SOPE. The intermediate shaded component refers to intact genes retained in only <i>S. glossinidius</i> (upper bar) or SOPE (lower bar) and the lighter shaded component refers to genes that are either absent or disrupted in both <i>S. glossinidius</i> and SOPE. The COG categories are organized in five larger groups with red representing genes involved in information storage and processing, blue representing genes involved in cellular processes and signaling, black representing genes involved in metabolism, green representing genes with poorly characterized functions, and yellow representing components of phages and IS-elements.</p

Posttransfusion Sepsis Attributable to Bacterial Contamination in Platelet Collection Set Manufacturing Facility, United States

During May 2018‒December 2022, we reviewed transfusion-transmitted sepsis cases in the United States attributable to polymicrobial contaminated apheresis platelet components, including Acinetobacter calcoaceticus‒baumannii complex or Staphylococcus saprophyticus isolated from patients and components. Transfused platelet components underwent bacterial risk control strategies (primary culture, pathogen reduction or primary culture, and secondary rapid test) before transfusion. Environmental samples were collected from a platelet collection set manufacturing facility. Seven sepsis cases from 6 platelet donations from 6 different donors were identified in patients from 6 states; 3 patients died. Cultures identified Acinetobacter calcoaceticus‒baumannii complex in 6 patients and 6 transfused platelets, S. saprophyticus in 4 patients and 4 transfused platelets. Whole-genome sequencing showed environmental isolates from the manufacturer were closely related genetically to patient and platelet isolates, indicating the manufacturer was the most probable source of recurrent polymicrobial contamination. Clinicians should maintain awareness of possible transfusion-transmitted sepsis even when using bacterial risk control strategies

Alignments of three regions of the <i>S. glossinidius</i>, strain HS, and SOPE chromosomes.

<p>Alignments of three regions of the <i>S. glossinidius</i>, strain HS, and SOPE chromosomes, corresponding to SG0948–SG0977 (A), ps_SGL0466–SG0918 (B) and ps_SGL0318–ps_SGL0330 (C) in the most recent <i>S. glossinidius</i> annotation <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002990#pgen.1002990-Belda1" target="_blank">[25]</a>. Putative ORFs and intergenic regions are drawn according to scale, oriented according to their inferred direction of transcription and color-coded according to COG functional categories. While all of the depicted strain HS genes have intact reading frames, the status of their orthologs in <i>S. glossinidius</i> and SOPE are shown in the outer bars (green = intact, purple = inactivated). Nonsense mutations (premature stop codons) are depicted by purple diamonds, and frameshifting indels are depicted by purple triangles. Light grey connecting bars are syntenic nucleotide alignments, while brown bars illustrate IS-element acquisitions that occur more frequently in SOPE.</p

Phylogeny of strain HS and related <i>Sodalis</i>-allied endosymbionts and free-living bacteria based on maximum likelihood analyses of a 1.46 kb fragment of 16S rRNA and a 1.68 kb fragment of <i>groEL</i>.

<p>Insect endosymbionts that do not have proper nomenclature are designed by the prefix “E”, followed by the name of their insect host. The numbers adjacent to nodes indicate maximum likelihood bootstrap values shown for nodes with bootstrap support >70%.</p

Allelic spectrum of pseudogene mutations in strain HS orthologs found in SOPE and <i>S. glossinidius</i>.

<p>Numbers in parentheses indicate the number of pseudogenes of strain HS orthologs found in the SOPE or <i>S. glossinidius</i> genome sequences. Nonsense mutations are classified as point mutations that catalyze the incorporation of a premature stop codon in the reading frame of a strain HS ortholog, independent of the presence of any frameshift resulting from an indel.</p>a<p>Nonsense mutations are classified as point mutations that catalyze the incorporation of a premature stop codon in the reading frame of an HS ortholog, independent of the presence of any frameshifting indel.</p