Catabolic Ornithine Carbamoyltransferase Activity Facilitates Growth of Staphylococcus aureus in Defined Medium Lacking Glucose and Arginine.

Previous studies have found that arginine biosynthesis in Staphylococcus aureus is repressed via carbon catabolite repression (CcpA), and proline is used as a precursor. Unexpectedly, however, robust growth of S. aureus is not observed in complete defined medium lacking both glucose and arginine (CDM-R). Mutants able to grow on agar-containing defined medium lacking arginine (CDM-R) were selected and found to contain mutations within ahrC, encoding the canonical arginine biosynthesis pathway repressor (AhrC), or single nucleotide polymorphisms (SNPs) upstream of the native arginine deiminase (ADI) operon arcA1B1D1C1. Reverse transcription-PCR (RT-PCR) studies found that mutations within ccpA or ahrC or SNPs identified upstream of arcA1B1D1C1 increased the transcription of both arcB1 and argGH, encoding ornithine carbamoyltransferase and argininosuccinate synthase/lyase, respectively, facilitating arginine biosynthesis. Furthermore, mutations within the AhrC homologue argR2 facilitated robust growth within CDM-R. Complementation with arcB1 or arcA1B1D1C1, but not argGH, rescued growth in CDM-R. Finally, supplementation of CDM-R with ornithine stimulated growth, as did mutations in genes (proC and rocA) that presumably increased the pyrroline-5-carboxylate and ornithine pools. Collectively, these data suggest that the transcriptional regulation of ornithine carbamoyltransferase and, in addition, the availability of intracellular ornithine pools regulate arginine biosynthesis in S. aureus in the absence of glucose. Surprisingly, ~50% of clinical S. aureus isolates were able to grow in CDM-R. These data suggest that S. aureus is selected to repress arginine biosynthesis in environments with or without glucose; however, mutants may be readily selected that facilitate arginine biosynthesis and growth in specific environments lacking arginine. IMPORTANCE Staphylococcus aureus can cause infection in virtually any niche of the human host, suggesting that it has significant metabolic versatility. Indeed, bioinformatic analysis suggests that it has the biosynthetic capability to synthesize all 20 amino acids. Paradoxically, however, it is conditionally auxotrophic for several amino acids, including arginine. Studies in our laboratory are designed to assess the biological function of amino acid auxotrophy in this significant pathogen. This study reveals that the metabolic block repressing arginine biosynthesis in media lacking glucose is the transcriptional repression of ornithine carbamoyltransferase encoded by arcB1 within the native arginine deiminase operon in addition to limited intracellular pools of ornithine. Surprisingly, approximately 50% of S. aureus clinical isolates can grow in media lacking arginine, suggesting that mutations are selected in S. aureus that allow growth in particular niches of the human host.

A Diverse Panel of Clinical Acinetobacter baumannii for Research and Development.

Over the past two decades, Acinetobacter baumannii has emerged as a leading cause of nosocomial infections worldwide. Of particular concern are panresistant strains, leading the World Health Organization (WHO) to designate carbapenem-resistant A. baumannii as a priority 1 (critical) pathogen for research and development of new antibiotics. A key component in supporting this effort is accessibility to diverse and clinically relevant strains for testing. Here, we describe a panel of 100 diverse A. baumannii strains for use in this endeavor. Whole-genome sequencing was performed on 3,505 A. baumannii isolates housed at the Multidrug-Resistant Organism Repository and Surveillance Network. Isolates were cultured from clinical samples at health care facilities around the world between 2001 and 2017. Core-genome multilocus sequence typing and high-resolution single nucleotide polymorphism (SNP)-based phylogenetic analyses were used to select a final panel of 100 strains that captured the genetic diversity of the collection. Comprehensive antibiotic susceptibility testing was also performed on all 100 isolates using 14 clinically relevant antibiotics. The final 100-strain diversity panel contained representative strains from 70 different traditional Pasteur scheme multilocus sequence types, including major epidemic clones. This diversity was also reflected in antibiotic susceptibility and antimicrobial resistance (AMR) gene content, with phenotypes ranging from pansensitive to panresistant, and over 100 distinct AMR gene alleles identified from 32 gene families. This panel provides the most diverse and comprehensive set of A. baumannii strains for use in developing solutions for combating antibiotic resistance. The panel and all available metadata, including genome sequences, will be available to industry and academic institutions and federal and other laboratories free of charge.

Tracking a serial killer: Integrating phylogenetic relationships, epidemiology, and geography for two invasive meningococcal disease outbreaks.

BACKGROUND: While overall rates of meningococcal disease have been declining in the United States for the past several decades, New York City (NYC) has experienced two serogroup C meningococcal disease outbreaks in 2005-2006 and in 2010-2013. The outbreaks were centered within drug use and sexual networks, were difficult to control, and required vaccine campaigns. METHODS: Whole Genome Sequencing (WGS) was used to analyze preserved meningococcal isolates collected before and during the two outbreaks. We integrated and analyzed epidemiologic, geographic, and genomic data to better understand transmission networks among patients. Betweenness centrality was used as a metric to understand the most important geographic nodes in the transmission networks. Comparative genomics was used to identify genes associated with the outbreaks. RESULTS: Neisseria meningitidis serogroup C (ST11/ET-37) was responsible for both outbreaks with each outbreak having distinct phylogenetic clusters. WGS did identify some misclassifications of isolates that were more distant from the outbreak strains, as well as those that should have been included based on high genomic similarity. Genomes for the second outbreak were more similar than the first and no polymorphism was found to either be unique or specific to either outbreak lineage. Betweenness centrality as applied to transmission networks based on phylogenetic analysis demonstrated that the outbreaks were transmitted within focal communities in NYC with few transmission events to other locations. CONCLUSIONS: Neisseria meningitidis is an ever changing pathogen and comparative genomic analyses can help elucidate how it spreads geographically to facilitate targeted interventions to interrupt transmission.

Complete Genome Sequence of Staphylococcus epidermidis 1457.

Staphylococcus epidermidis 1457 is a frequently utilized strain that is amenable to genetic manipulation and has been widely used for biofilm-related research. We report here the whole-genome sequence of this strain, which encodes 2,277 protein-coding genes and 81 RNAs within its 2.4-Mb genome and plasmid.

Comparative genomics of bacteria in the genus Providencia isolated from wild Drosophila melanogaster.

BACKGROUND: Comparative genomics can be an initial step in finding the genetic basis for phenotypic differences among bacterial strains and species. Bacteria belonging to the genus Providencia have been isolated from numerous and varied environments. We sequenced, annotated and compared draft genomes of P. rettgeri, P. sneebia, P. alcalifaciens, and P. burhodogranariea. These bacterial species that were all originally isolated as infections of wild Drosophila melanogaster and have been previously shown to vary in virulence to experimentally infected flies. RESULTS: We found that these Providencia species share a large core genome, but also possess distinct sets of genes that are unique to each isolate. We compared the genomes of these isolates to draft genomes of four Providencia isolated from the human gut and found that the core genome size does not substantially change upon inclusion of the human isolates. We found many adhesion related genes among those genes that were unique to each genome. We also found that each isolate has at least one type 3 secretion system (T3SS), a known virulence factor, though not all identified T3SS belong to the same family nor are they in syntenic genomic locations. CONCLUSIONS: The Providencia species examined here are characterized by high degree of genomic similarity which will likely extend to other species and isolates within this genus. The presence of T3SS islands in all of the genomes reveal that their presence is not sufficient to indicate virulence towards D. melanogaster, since some of the T3SS-bearing isolates are known to cause little mortality. The variation in adhesion genes and the presence of T3SSs indicates that host cell adhesion is likely an important aspect of Providencia virulence.

Comparative pathology of bacteria in the genus Providencia to a natural host, Drosophila melanogaster.

Bacteria in the genus Providencia are pathogens of many organisms, including humans and insects. We and colleagues have isolated five different strains belonging to four distinct Providencia species as natural infections of Drosophila melanogaster captured in the wild. We found that these isolates vary considerably in pathology to infected D. melanogaster, differing in the level of mortality they cause, their ability to replicate within the host and the level that the fly's immune response is elicited. One interesting bacterium was Providencia sneebia, which causes nearly complete mortality and reaches large numbers in the fly but does not elicit a comparably strong immune response. Through coinfection experiments, we determined that P. sneebia avoids recognition by the immune system. We tested for biofilm formation and replication within D. melanogaster cells as possible mechanisms for P. sneebia escape from host immunity, but did not find evidence for either. D. melanogaster and Providencia provide a powerful system for studying general host-pathogen interactions, and for understanding how the well-studied immune model host D. melanogaster interacts with its natural bacterial pathogens.

Disease pathology: wasting energy fighting infection.

Drosophila melanogaster infected with Mycobacterium marinum suffer metabolic wasting similar to that seen in humans suffering from tuberculosis. This wasting is linked to insulin signaling and hastens host death.