Mark T. Anderson

B-Cation arrangements in double perovskites

A survey of A′A″B′B″O6 double perovskites, which encompasses over 300 compounds synthesized from about 1950 to the present, shows that three distinct B-cation arrangements are known: random, rock salt, and layered. Examples are provided to illustrate the most common symmetries of these B-cation sublattices and to show how they can be assigned based on powder diffraction patterns. A comprehensive examination of the factors that influence B-cation arrangement, namely the charge, size, and electronic configuration of the B cations, and the AB size ratio, is presented. Special consideration is given to the layered sublattice because of its rarity and its potential importance in two-dimensional materials, such as cuprate superconductors.

Opportunity and Means: Horizontal Gene Transfer from the Human Host to a Bacterial Pathogen

ABSTRACT The acquisition and incorporation of genetic material between nonmating species, or horizontal gene transfer (HGT), has been frequently described for phylogenetically related organisms, but far less evidence exists for HGT between highly divergent organisms. Here we report the identification and characterization of a horizontally transferred fragment of the human long interspersed nuclear element L1 to the genome of the strictly human pathogen Neisseria gonorrhoeae. A 685-bp sequence exhibiting 98 to 100% identity to copies of the human L1 element was identified adjacent to the irg4 gene in some N. gonorrhoeae genomes. The L1 fragment was observed in ~11% of the N. gonorrhoeae population sampled but was not detected in Neisseria meningitidis or commensal Neisseria isolates. In addition, N. gonorrhoeae transcripts containing the L1 sequence were detected by reverse transcription-PCR, indicating that an L1-derived gene product may be produced. The high degree of identity between human and gonococcal L1 sequences, together with the absence of L1 sequences from related Neisseria species, indicates that this HGT event occurred relatively recently in evolutionary history. The identification of L1 sequences in N. gonorrhoeae demonstrates that HGT can occur between a mammalian host and a resident bacterium, which has important implications for the coevolution of both humans and their associated microorganisms. IMPORTANCE The interactions between microbes and their hosts are relevant to several aspects of biology, including evolution, development, immunity, and disease. Neisseria gonorrhoeae serves as a particularly informative model for this interaction because it has exclusively coevolved with humans and is not known to be found in any other environment. In addition, investigation of the evolutionary relationship between N. gonorrhoeae and humans has practical implications, since gonorrhea is a prevalent sexually transmitted infection worldwide. This study was undertaken to characterize the horizontal transfer of genetic information from humans to N. gonorrhoeae, an event that has been scarcely recognized between any mammalian host and bacterial pathogen. Here we provide evidence that this genetic exchange was the result of a recent evolutionary event that has been propagated within the gonococcal population. The interactions between microbes and their hosts are relevant to several aspects of biology, including evolution, development, immunity, and disease. Neisseria gonorrhoeae serves as a particularly informative model for this interaction because it has exclusively coevolved with humans and is not known to be found in any other environment. In addition, investigation of the evolutionary relationship between N. gonorrhoeae and humans has practical implications, since gonorrhea is a prevalent sexually transmitted infection worldwide. This study was undertaken to characterize the horizontal transfer of genetic information from humans to N. gonorrhoeae, an event that has been scarcely recognized between any mammalian host and bacterial pathogen. Here we provide evidence that this genetic exchange was the result of a recent evolutionary event that has been propagated within the gonococcal population.

The Bordetella Bfe System: Growth and Transcriptional Response to Siderophores, Catechols, and Neuroendocrine Catecholamines

Ferric enterobactin utilization by Bordetella bronchiseptica and Bordetella pertussis requires the BfeA outer membrane receptor. Under iron-depleted growth conditions, transcription of bfeA is activated by the BfeR regulator by a mechanism requiring the siderophore enterobactin. In this study, enterobactin-inducible bfeA transcription was shown to be TonB independent. To determine whether other siderophores or nonsiderophore catechols could be utilized by the Bfe system, various compounds were tested for the abilities to promote the growth of iron-starved B. bronchiseptica and induce bfeA transcription. The BfeA receptor transported ferric salmochelin, corynebactin, and the synthetic siderophores TRENCAM and MECAM. Salmochelin and MECAM induced bfeA transcription in iron-starved Bordetella cells, but induction by corynebactin and TRENCAM was minimal. The neuroendocrine catecholamines epinephrine, norepinephrine, and dopamine exhibited a remarkable capacity to induce transcription of bfeA. Norepinephrine treatment of B. bronchiseptica resulted in BfeR-dependent bfeA transcription, elevated BfeA receptor production, and growth stimulation. Pyrocatechol, carbidopa, and isoproterenol were similarly strong inducers of bfeA transcription, whereas tyramine and 3,4-dihydroxymandelic acid demonstrated low inducing activity. The results indicate that the inducer structure requires a catechol group for function and that the ability to induce bfeA transcription does not necessarily correlate with the ability to stimulate bacterial growth. The expanded range of catechol siderophores transported by the BfeA receptor demonstrates the potential versatility of the Bordetella Bfe iron retrieval system. The finding that catecholamine neurotransmitters activate bfeA transcription and promote growth suggests that Bordetella cells can perceive and may benefit from neuroendocrine catecholamines on the respiratory epithelium.

Norepinephrine mediates acquisition of transferrin-iron in Bordetella bronchiseptica.

Previous research demonstrated that the sympathoadrenal catecholamine norepinephrine could promote the growth of Bordetella bronchiseptica in iron-restricted medium containing serum. In this study, norepinephrine was demonstrated to stimulate growth of this organism in the presence of partially iron-saturated transferrin but not lactoferrin. Although norepinephrine is known to induce transcription of the Bordetella bfeA enterobactin catechol xenosiderophore receptor gene, neither a bfeA mutant nor a bfeR regulator mutant was defective in growth responsiveness to norepinephrine. However, growth of a tonB mutant strain was not enhanced by norepinephrine, indicating that the response to this catecholamine was the result of high-affinity outer membrane transport. The B. bronchiseptica genome encodes a total of 19 known and predicted iron transport receptor genes, none of which, when mutated individually, were found to confer a defect in norepinephrine-mediated growth stimulation in the presence of transferrin. Labeling experiments demonstrated a TonB-dependent increase in cell-associated iron levels when bacteria grown in the presence of (55)Fe-transferrin were exposed to norepinephrine. In addition, TonB was required for maximum levels of cell-associated norepinephrine. Together, these results demonstrate that norepinephrine facilitates B. bronchiseptica iron acquisition from the iron carrier protein transferrin and this process may represent a mechanism by which some bacterial pathogens obtain this essential nutrient in the host environment.

The BfeR Regulator Mediates Enterobactin-Inducible Expression of Bordetella Enterobactin Utilization Genes

Utilization of the enterobactin siderophore by the respiratory pathogens Bordetella pertussis and Bordetella bronchiseptica is dependent on the BfeA outer membrane receptor. This study determined that production of BfeA was increased significantly in iron-starved bacteria upon supplementation of cultures with enterobactin. A 1.01-kb open reading frame, designated bfeR, encoding a predicted positive transcriptional regulator of the AraC family was identified upstream and divergently oriented from bfeA. In iron-depleted cultures containing enterobactin, a Bordetella bfeR mutant exhibited markedly decreased BfeA receptor production compared to that of the wild-type strain. Additionally, B. pertussis and B. bronchiseptica bfeR mutants exhibited impaired growth with ferric enterobactin as the sole source of iron, demonstrating that effective enterobactin utilization is bfeR dependent. Transcriptional analysis using bfeA-lacZ reporter fusions in wild-type strains demonstrated that bfeA transcription was stimulated in iron-depleted conditions in the presence of enterobactin, compared to modest expression levels in cultures lacking enterobactin. In contrast, bfeA transcription in B. pertussis and B. bronchiseptica bfeR mutants was completely unresponsive to the enterobactin inducer. bfeA transcriptional analyses of a bfeA mutant demonstrated that induction by enterobactin did not require BfeA receptor-mediated uptake of the siderophore. These studies establish that bfeR encodes an enterobactin-dependent positive regulator of bfeA transcription in these Bordetella species.

Bordetella iron transport and virulence

Bordetella pertussis, Bordetella parapertussis, and Bordetella bronchiseptica are pathogens with a complex iron starvation stress response important for adaptation to nutrient limitation and flux in the mammalian host environment. The iron starvation stress response is globally regulated by the Fur repressor using ferrous iron as the co-repressor. Expression of iron transport system genes of Bordetella is coordinated by priority regulation mechanisms that involve iron source sensing. Iron source sensing is mediated by distinct transcriptional activators that are responsive to the cognate iron source acting as the inducer.

Differential expression of Bordetella pertussis iron transport system genes during infection

Temporal expression patterns of the Bordetella pertussis alcaligin, enterobactin and haem iron acquisition systems were examined using alcA–, bfeA– and bhuR–tnpR recombinase fusion strains in a mouse respiratory infection model. The iron systems were differentially expressed in vivo, showing early induction of the alcaligin and enterobactin siderophore systems, and delayed induction of the haem system in a manner consistent with predicted changes in host iron source availability during infection. Previous mixed infection competition studies established the importance of alcaligin and haem utilization for B. pertussis in vivo growth and survival. In this study, the contribution of the enterobactin system to the fitness of B. pertussis was confirmed using wild‐type and enterobactin receptor mutant strains in similar competition infection experiments. As a correlate to the in vivo expression studies of B. pertussis iron systems in mice, sera from uninfected and B. pertussis‐infected human donors were screened for antibody reactivity with Bordetella iron‐repressible cell envelope proteins. Pertussis patient sera recognized multiple iron‐repressible proteins including the known outer membrane receptors for alcaligin, enterobactin and haem, supporting the hypothesis that B. pertussis is iron‐starved and responds to the presence of diverse iron sources during natural infection.

Seminal Plasma Initiates a Neisseria gonorrhoeae Transmission State

ABSTRACT Niche-restricted pathogens are evolutionarily linked with the specific biological fluids that are encountered during infection. Neisseria gonorrhoeae causes the genital infection gonorrhea and is exposed to seminal fluid during sexual transmission. Treatment of N. gonorrhoeae with seminal plasma or purified semen proteins lactoferrin, serum albumin, and prostate-specific antigen each facilitated type IV pilus-mediated twitching motility of the bacterium. Motility in the presence of seminal plasma was characterized by high velocity and low directional persistence. In addition, infection of epithelial cells with N. gonorrhoeae in the presence of seminal plasma resulted in enhanced microcolony formation. Close association of multiple pili in the form of bundles was also disrupted after seminal plasma treatment leading to an increase in the number of single pilus filaments on the bacterial surface. Thus, exposure of N. gonorrhoeae to seminal plasma is proposed to alter bacterial motility and aggregation characteristics to influence the processes of transmission and colonization. IMPORTANCE There are greater than 100 million estimated new cases of gonorrhea annually worldwide. Research characterizing the mechanisms of pathogenesis and transmission of Neisseria gonorrhoeae is important for developing new prevention strategies, since antibiotic resistance of the organism is becoming increasingly prevalent. Our work identifies seminal plasma as a mediator of N. gonorrhoeae twitching motility and microcolony formation through functional modification of the type IV pilus. These findings provide insight into motility dynamics and epithelial cell colonization under conditions that are relevant to sexual transmission. Type IV pili are common virulence factors with diverse functions among bacterial pathogens, and this work identifies interactions between type IV pili and the host environment. Finally, this work illustrates the importance of the host environment and niche-specific fluids on microbial pathogenesis. There are greater than 100 million estimated new cases of gonorrhea annually worldwide. Research characterizing the mechanisms of pathogenesis and transmission of Neisseria gonorrhoeae is important for developing new prevention strategies, since antibiotic resistance of the organism is becoming increasingly prevalent. Our work identifies seminal plasma as a mediator of N. gonorrhoeae twitching motility and microcolony formation through functional modification of the type IV pilus. These findings provide insight into motility dynamics and epithelial cell colonization under conditions that are relevant to sexual transmission. Type IV pili are common virulence factors with diverse functions among bacterial pathogens, and this work identifies interactions between type IV pili and the host environment. Finally, this work illustrates the importance of the host environment and niche-specific fluids on microbial pathogenesis.

Phase Variation Leads to the Misidentification of a Neisseria Gonorrhoeae Virulence Gene

Neisseria gonorrhoeae is the causative agent of gonorrhea and an obligate pathogen of humans. The Opa proteins of these bacteria are known to mediate attachment and internalization by host cells, including neutrophils. The Opa protein repertoire of a typical N. gonorrhoeae isolate is encoded on ∼11 genes distributed throughout the chromosome and is subject to stochastic changes in expression through phase variation. Together, these characteristics make Opa proteins a critical yet unpredictable aspect of any experimental investigation into the interaction of N. gonorrhoeae with host cells. The goal of this study was to identify novel virulence factors of N. gonorrhoeae by assessing the contribution of a set of uncharacterized hydrogen peroxide-induced genes to bacterial survival against neutrophil-mediated killing. To this end, a strain harboring an engineered mutation in the NGO0322 gene was identified that exhibited increased sensitivity to neutrophil-mediated killing, enhanced internalization by neutrophils, and the ability to induce high levels of neutrophil-generated reactive oxygen species. Each of these phenotypes reverted to near wild-type levels following genetic complementation of the NGO0322 mutation. However, after immunoblot analysis of Opa proteins expressed by the isogenic parent, mutant, and genetically complemented strains, it was determined that phase variation had resulted in a disparity between the Opa profiles of these strains. To determine whether Opa phase variation, rather than NGO0322 mutation, was the cause of the observed neutrophil-related phenotypes, NGO0322 function was investigated in N. gonorrhoeae strains lacking all Opa proteins or constitutively expressing the OpaD variant. In both cases, mutation of NGO0322 did not alter survival of gonococci in the presence of neutrophils. These results demonstrate the importance of controlling for the frequent and random variation in Opa protein production by N. gonorrhoeae when investigating host cell interactions.

Neisseria gonorrhoeae and humans perform an evolutionary LINE dance

Horizontal gene transfer is an important mechanism for generating genetic diversity. As the number of sequenced genomes continues to increase, so do the examples of horizontal genetic exchange between both related and divergent organisms. Here we discuss the recent finding that certain strains of the human pathogen Neisseria gonorrhoeae have incorporated a small fragment of human DNA sequence into their genomes. The horizontally acquired sequence exhibits 98-100% nucleotide identity to a 685-bp portion of the highly repetitive retrotransposable element L1 and its presence in the gonococcal genome has been confirmed by multiple molecular techniques. The possibility of similar L1 horizontal gene transfer events having occurred in other bacteria based on genomic sequence evidence is explored. Potential mechanisms of how N. gonorrhoeae was able to acquire and maintain this human sequence are also discussed in addition to the evolutionary implications of such an event.