Serum and Urethral Antibody Response in Mycoplasma genitalium -Infected Men.

ABSTRACT: The antibody response to Mycoplasma genitalium in serum and urethral secretions of men with nongonococcal urethritis was examined longitudinally. Serum and urethral antibodies reacted primarily with the MgpB and MgpC adhesins. Serum antibodies persisted throughout follow-up, whereas urethral antibodies waned despite organism persistence. Declining antibodies may facilitate chronic infection.

Appraisal of Fungus-Derived Xanthoquinodins as Broad-Spectrum Anti-Infectives Targeting Phylogenetically Diverse Human Pathogens.

Xanthoquinodins make up a distinctive class of xanthone-anthraquinone heterodimers reported as secondary metabolites from several fungal species. Through a collaborative multi-institutional screening program, a fungal extract prepared from a Trichocladium sp. was identified that exhibited strong inhibitory effects against several human pathogens (Mycoplasma genitalium, Plasmodium falciparum, Cryptosporidium parvum, and Trichomonas vaginalis). This report focuses on one of the unique samples that exhibited a desirable combination of biological effects: namely, it inhibited all four test pathogens and demonstrated low levels of toxicity toward HepG2 (human liver) cells. Fractionation and purification of the bioactive components and their congeners led to the identification of six new compounds [xanthoquinodins NPDG A1-A5 (1-5) and B1 (6)] as well as several previously reported natural products (7-14). The chemical structures of 1-14 were determined based on interpretation of their 1D and 2D NMR, HRESIMS, and electronic circular dichroism (ECD) data. Biological testing of the purified metabolites revealed that they possessed widely varying levels of inhibitory activity against a panel of human pathogens. Xanthoquinodins A1 (7) and A2 (8) exhibited the most promising broad-spectrum inhibitory effects against M. genitalium (EC50 values: 0.13 and 0.12 µM, respectively), C. parvum (EC50 values: 5.2 and 3.5 µM, respectively), T. vaginalis (EC50 values: 3.9 and 6.8 µM, respectively), and P. falciparum (EC50 values: 0.29 and 0.50 µM, respectively) with no cytotoxicity detected at the highest concentration tested (HepG2 EC50 > 25 µM).

In Vitro Susceptibility and Resistance of Mycoplasma genitalium to Nitroimidazoles.

Mycoplasma genitalium is a sexually transmitted reproductive tract pathogen of men and women. M. genitalium infections are increasingly difficult to treat due to poor efficacy of doxycycline and acquired resistance to azithromycin and moxifloxacin. A recent clinical trial suggested that metronidazole may improve cure rates for women with pelvic inflammatory disease and reduced the detection of M. genitalium when included with standard doxycycline plus ceftriaxone treatment. As data regarding susceptibility of mycoplasmas to nitroimidazoles are lacking in the scientific literature, we determined the in vitro susceptibility of 10 M. genitalium strains to metronidazole, secnidazole, and tinidazole. MICs ranged from 1.6 to 12.5 µg/mL for metronidazole, 3.1 to 12.5 µg/mL for secnidazole, and 0.8 to 6.3 µg/mL for tinidazole. None of these agents was synergistic with doxycycline in checkerboard broth microdilution assays. Tinidazole was superior to metronidazole and secnidazole in terms of MIC and time-kill kinetics and was bactericidal (>99.9% killing) at concentrations below reported serum concentrations. Mutations associated with nitroimidazole resistance were identified by whole-genome sequencing of spontaneous resistant mutants, suggesting a mechanism for reductive activation of the nitroimidazole prodrug by a predicted NAD(P)H-dependent flavin mononucleotide (FMN) oxidoreductase. The presence of oxygen did not affect MICs of wild-type M. genitalium, but a nitroimidazole-resistant mutant was defective for growth under anaerobic conditions, suggesting that resistant mutants may have a fitness disadvantage in anaerobic genital sites. Clinical studies are needed to determine if nitroimidazoles, especially tinidazole, are effective for eradicating M. genitalium infections in men and women.

Update in Epidemiology and Management of Mycoplasma genitalium Infections.

Mycoplasma genitalium is a frequent cause of urogenital syndromes in men and women and is associated with adverse sequelae in women. M genitalium also infects the rectum, and may cause proctitis, but rarely infects the pharynx. Diagnosis requires nucleic acid amplification testing. Antibiotic resistance is widespread: more than half of infections are resistant to macrolides and fluoroquinolone resistance is increasing. Resistance-guided therapy is recommended for symptomatic patients, involving initial treatment with doxycycline to reduce organism load followed by azithromycin for macrolide-sensitive infections or moxifloxacin for macrolide-resistant infections. Neither screening nor tests of cure are recommended in asymptomatic persons.

Ascending Reproductive Tract Infection in Pig-Tailed Macaques Inoculated with Mycoplasma genitalium.

Mycoplasma genitalium is a sexually transmitted bacterial pathogen that causes urogenital disease in men and women. M. genitalium infections can persist for months to years and can ascend to the upper reproductive tract in women where it is associated with serious sequelae including pelvic inflammatory disease, tubal factor infertility, and preterm birth. An animal model is needed to understand immune evasion strategies that allow persistence, mechanisms of ascending infection, and factors associated with clearance. In earlier studies, we determined that pig-tailed macaques are susceptible to cervical infection; however, not all primates were successfully infected, persistence varied between animals, and ascension to the upper reproductive tract was not observed after 4 or 8 weeks of follow-up. Building on our previous findings, we refined our inoculation methods to increase infection rates, extended observation to 18 weeks, and comprehensively sampled the upper reproductive tract to detect ascending infection. With these improvements, we established infection in all (3/3) primates inoculated with M. genitalium and demonstrated lower tract persistence for 16 to 18 weeks. Ascension to the upper reproductive tract at endpoint was observed in two out of three primates. All three primates developed serum and local antibodies reacting primarily to the MgpB and MgpC adherence proteins. Elevated genital polymorphonuclear leukocytes (PMNs) and inflammatory cytokines and chemokines, erythema of the ectocervix in one primate, and histologic evidence of vaginitis and endocervicitis in two primates suggest a mild to moderate inflammatory response to infection. This model will be valuable to understand the natural history of M. genitalium infection including mechanisms of persistence, immune evasion, and ascension to the upper reproductive tract.

Azithromycin and Doxycycline Resistance Profiles of U.S. Mycoplasma genitalium Strains and Their Association with Treatment Outcomes.

Mycoplasma genitalium is a sexually transmitted bacterium associated with nongonococcal urethritis (NGU) in men and cervicitis, endometritis, and pelvic inflammatory disease in women. Effective treatment is challenging due to the inherent, and increasingly acquired, antibiotic resistance in this pathogen. In our treatment trial conducted from 2007 to 2011 in Seattle, WA, we demonstrated poor efficacy of azithromycin (AZM) and doxycycline (DOX) against M. genitalium among men with NGU. In the present study, we cultured M. genitalium from 74 of 80 (92.5%) PCR-positive men at enrollment (V-1) and defined the MICs of AZM (N = 56 isolates) of DOX (N = 62 isolates). Susceptibility to AZM was bimodal; MICs were >8 µg/ml (44.6%) and <0.004 µg/ml (55.4%) for these isolates. The association of MIC with treatment efficacy was determined for men initially treated with either AZM (N = 30) or DOX (N = 24). Men treated with AZM were more likely to experience microbiologic treatment failure (P < 0.001) if infected with isolates that had AZM MICs of >8 µg/ml (18/18 men) than those with isolates that had AZM MICs of <0.004 µg/ml (1/12 men). Clinical treatment failure also was more likely to occur (P = 0.002) with AZM MICs of >8 µg/ml (12/18 men) than with AZM MICs of <0.004 µg/ml (1/12 men). In contrast, DOX MICs ranged from <0.125 to 2 µg/ml and were not correlated with microbiologic (P = 0.71) or clinical treatment (P = 0.41) failure, demonstrating no relationship between DOX MICs and treatment efficacy. Given the rapid spread of AZM resistance and the emergence of quinolone resistance, the current second-line therapy, monitoring MICs and evaluating other potential treatments for M. genitalium will be critical.

Sequence variation and immunogenicity of the Mycoplasma genitalium MgpB and MgpC adherence proteins during persistent infection of men with non-gonococcal urethritis.

Mycoplasma genitalium is a sexually transmitted bacterial pathogen that infects men and women. Antigenic variation of MgpB and MgpC, the immunodominant adherence proteins of M. genitalium, is thought to contribute to immune evasion and chronic infection. We investigated the evolution of mgpB and mgpC sequences in men with non-gonococcal urethritis persistently infected with M. genitalium, including two men with anti-M. genitalium antibodies at enrollment and two that developed antibodies during follow-up. Each of the four patients was persistently infected with a different strain type and each patient produced antibodies targeting MgpB and MgpC. Amino acid sequence evolution in the variable regions of MgpB and MgpC occurred in all four patients with changes observed in single and multiple variable regions over time. Using the available crystal structure of MgpC of the G37 type strain we found that predicted conformational B cell epitopes localize predominantly to the variable region of MgpC, amino acids that changed during patient infection lie in these epitopes, and variant amino acids are in close proximity to the conserved sialic acid binding pocket. These findings support the hypothesis that sequence variation functions to avoid specific antibodies thereby contributing to persistence in the genital tract.

Design and Validation of Transcription-Mediated-Amplification Nucleic Acid Amplification Tests for Mycoplasma genitalium.

Mycoplasma genitalium is a sexually transmitted bacterium linked to adverse sexual and reproductive health outcomes in women and men. M. genitalium is difficult to culture, and in the absence of validated amplified molecular methods for diagnosis of infection, there is no reference standard available for use as a comparator for the validation of new M. genitalium diagnostic tests. We evaluated the analytical and clinical performance of three transcription-mediated amplification (TMA) tests for M. genitalium, each targeting unique rRNA sequences, for use as a composite comparator for clinical validation of the Aptima Mycoplasma genitalium (AMG) assay, an in vitro diagnostic (IVD) TMA test that targets 16 s rRNA of M. genitalium Analytical sensitivity, specificity, and strain inclusivity of all four TMA tests were determined using nine laboratory strains of M. genitalium and 56 nontarget bacteria, protozoa, and viruses. Analytical sensitivity of the tests for M. genitalium ranged from 0.017 to 0.040 genome equivalents/ml. None of the nontarget organisms evaluated cross-reacted with any test. A composite comparator reference standard consisting of the 3 alternate (Alt) TMA tests was used to evaluate the clinical performance of the AMG assay by testing residual vaginal swab, female urine, and male urine specimens obtained from 1,400 adult subjects from three U.S. clinical sites. Using this reference standard to establish infected specimen status, the sensitivity, specificity, and overall agreement of the AMG IVD assay were 100%, 99.9%, and 99.9%, respectively. These results demonstrate the utility of molecular composite reference standard methodology for the clinical validation of future IVD tests for this organism.

Mycoplasma genitalium Nonadherent Phase Variants Arise by Multiple Mechanisms and Escape Antibody-Dependent Growth Inhibition.

Antigenic variation of the immunodominant MgpB and MgpC proteins has been suggested to be a mechanism of immune evasion of the human pathogen Mycoplasma genitalium, a cause of several reproductive tract disease syndromes. Phase variation resulting in the loss of adherence has also been documented, but the molecular mechanisms underlying this process and its role in pathogenesis are still poorly understood. In this study, we isolated and characterized 40 spontaneous, nonadherent phase variants from in vitro-passaged M. genitalium cultures. In all cases, nonadherence was associated with the loss of MgpBC protein expression, attributable to sequence changes in the mgpBC expression site. Phase variants were grouped into seven classes on the basis of the nature of the mutation. Consistent with the established role of RecA in phase variation, 31 (79.5%) variants arose via recombination with MgPa repeat regions that contain mgpBC variable sequences. The remaining mutants arose via nonsense or frameshift mutations. As expected, revertants were obtained for phase variants that were predicted to be reversible but not for those that arose via an irreversible mechanism. Furthermore, phase variants were enriched in M. genitalium cultures exposed to antibodies reacting to the extracellular, conserved C terminus of MgpB but not in cultures exposed to antibodies reacting to an intracellular domain of MgpB or the cytoplasmic HU protein. Genetic characterization of the antibody-selected phase variants confirmed that they arose via reversible and irreversible recombination and point mutations within mgpBC These phase variants resisted antibody-mediated growth inhibition, suggesting that phase variation promotes immune evasion.

Experimental Infection of Pig-Tailed Macaques (Macaca nemestrina) with Mycoplasma genitalium.

Mycoplasma genitalium is an underappreciated cause of human reproductive tract disease, characterized by persistent, often asymptomatic, infection. Building on our previous experiments using a single female pig-tailed macaque as a model for M. genitalium infection (G. E. Wood, S. L. Iverson-Cabral, D. L. Patton, P. K. Cummings, Y. T. Cosgrove Sweeney, and P. A. Totten, Infect Immun 81:2938-2951, 2013, https://doi.org/10.1128/IAI.01322-12), we cervically inoculated eight additional animals, two of which were simultaneously inoculated in salpingeal tissue autotransplanted into abdominal pockets. Viable M. genitalium persisted in the lower genital tract for 8 weeks in three animals, 4 weeks in two, and 1 week in one; two primates resisted infection. In both animals inoculated in salpingeal pockets, viable M. genitalium was recovered for 2 weeks. Recovery of viable M. genitalium from lower genital tract specimens was improved by diluting the specimen in broth and by Vero cell coculture. Ascension to upper reproductive tract tissues was not detected, even among three persistently infected animals. M. genitalium-specific serum antibodies targeting the immunodominant MgpB and MgpC proteins appeared within 1 week in three animals inoculated both cervically and in salpingeal pockets and in one of three persistently infected animals inoculated only in the cervix. M. genitalium-specific IgG, but not IgA, was detected in cervical secretions of serum antibody-positive animals, predominantly against MgpB and MgpC, but was insufficient to clear M. genitalium lower tract infection. Our findings further support female pig-tailed macaques as a model of M. genitalium infection, persistence, and immune evasion.

Analysis of the Mycoplasma genitalium MgpB Adhesin to Predict Membrane Topology, Investigate Antibody Accessibility, Characterize Amino Acid Diversity, and Identify Functional and Immunogenic Epitopes.

Mycoplasma genitalium is a sexually transmitted pathogen and is associated with reproductive tract disease that can be chronic in nature despite the induction of a strong antibody response. Persistent infection exacerbates the likelihood of transmission, increases the risk of ascension to the upper tract, and suggests that M. genitalium may possess immune evasion mechanism(s). Antibodies from infected patients predominantly target the MgpB adhesin, which is encoded by a gene that recombines with homologous donor sequences, thereby generating sequence variation within and among strains. We have previously characterized mgpB heterogeneity over the course of persistent infection and have correlated the induction of variant-specific antibodies with the loss of that particular variant from the infected host. In the current study, we examined the membrane topology, antibody accessibility, distribution of amino acid diversity, and the location of functional and antigenic epitopes within the MgpB adhesin. Our results indicate that MgpB contains a single transmembrane domain, that the majority of the protein is surface exposed and antibody accessible, and that the attachment domain is located within the extracellular C-terminus. Not unexpectedly, amino acid diversity was concentrated within and around the three previously defined variable regions (B, EF, and G) of MgpB; while nonsynonymous mutations were twice as frequent as synonymous mutations in regions B and G, region EF had equal numbers of nonsynonymous and synonymous mutations. Interestingly, antibodies produced during persistent infection reacted predominantly with the conserved C-terminus and variable region B. In contrast, infection-induced antibodies reacted poorly with the N-terminus, variable regions EF and G, and intervening conserved regions despite the presence of predicted B cell epitopes. Overall, this study provides an important foundation to define how different segments of the MgpB adhesin contribute to functionality, variability, and immunogenicity during persistent M. genitalium infection.

Persistence, immune response, and antigenic variation of Mycoplasma genitalium in an experimentally infected pig-tailed macaque (Macaca nemestrina).

Mycoplasma genitalium is a sexually transmitted pathogen associated with several acute and chronic reproductive tract disease syndromes in men and women. To evaluate the suitability of a pig-tailed macaque model of M. genitalium infection, we inoculated a pilot animal with M. genitalium strain G37 in the uterine cervix and in salpingeal pockets generated by transplanting autologous Fallopian tube tissue subcutaneously. Viable organisms were recovered throughout the 8-week experiment in cervicovaginal specimens and up to 2 weeks postinfection in salpingeal pockets. Humoral and cervicovaginal antibodies reacting to MgpB were induced postinoculation and persisted throughout the infection. The immunodominance of the MgpB adhesin and the accumulation of mgpB sequence diversity previously observed in persistent human infections prompted us to evaluate sequence variation in this animal model. We found that after 8 weeks of infection, sequences within mgpB variable region B were replaced by novel sequences generated by reciprocal recombination with an archived variant sequence located elsewhere on the chromosome. In contrast, mgpB region B of the same inoculum propagated for 8 weeks in vitro remained unchanged. Notably, serum IgG reacted strongly with a recombinant protein spanning MgpB region B of the inoculum, while reactivity to a recombinant protein representing the week 8 variant was reduced, suggesting that antibodies were involved in the clearance of bacteria expressing the original infecting sequence. Together these results suggest that the pig-tailed macaque is a suitable model to study M. genitalium pathogenesis, antibody-mediated selection of antigenic variants in vivo, and immune escape.

RecA mediates MgpB and MgpC phase and antigenic variation in Mycoplasma genitalium, but plays a minor role in DNA repair.

Mycoplasma genitalium, a sexually transmitted human pathogen, encodes MgpB and MgpC adhesins that undergo phase and antigenic variation through recombination with archived 'MgPar' donor sequences. The mechanism and molecular factors required for this genetic variation are poorly understood. In this study, we estimate that sequence variation at the mgpB/C locus occurs in vitro at a frequency of > 1.25 × 10(-4) events per genome per generation using a quantitative anchored PCR assay. This rate was dramatically reduced in a recA deletion mutant and increased in a complemented strain overexpressing RecA. Similarly, the frequency of haemadsorption-deficient phase variants was reduced in the recA mutant, but restored by complementation. Unlike Escherichia coli, inactivation of recA in M. genitalium had a minimal effect on survival after exposure to mitomycin C or UV irradiation. In contrast, a deletion mutant for the predicted nucleotide excision repair uvrC gene showed growth defects and was exquisitely sensitive to DNA damage. We conclude that M. genitalium RecA has a primary role in mgpB/C-MgPar recombination leading to antigenic and phase variation, yet plays a minor role in DNA repair. Our results also suggest that M. genitalium possesses an active nucleotide excision repair system, possibly representing the main DNA repair pathway in this minimal bacterium.

Chronic immobilization stress alters aspects of emotionality and associative learning in the rat.

Chronic stress significantly alters limbic neuroarchitecture and function, and potentiates emotionality in rats. Chronic restraint stress (CRS) increases aggression among familiar rats, potentiates anxiety, and enhances fear conditioning. Chronic immobilization stress (CIS) induces anxiety behavior and dendritic hypertrophy in the basolateral amygdala, which persist beyond a recovery period. However, little else is known about the emotional impact of CIS as a model of chronic stress or depression. Therefore, the authors present two experiments examining emotional and learned responses to CIS. In Experiment I, the authors examine individual differences in behaviors during and after CIS, specifically: struggling, aggression, learned helplessness, inhibitory avoidance, and escape behavior. In Experiment II, the authors confirm the effects of CIS on aggression and struggling during immobilization, and correlate individual responses with aspects of conditioned fear. Here the authors report significant effects of CIS on aggression, inhibitory avoidance, escape, as well as learned aspects of fear (i.e., fear conditioning) and inescapable stress (i.e., struggling and helplessness). These results emphasize the emotional and learned responses to CIS evident during and after the stress treatment, as well as the importance of individual differences.

Uncovering the mechanisms of estrogen effects on hippocampal function.

Estrogens have direct effects on the brain areas controlling cognition. One of the most studied of these regions is the dorsal hippocampal formation, which governs the formation of spatial and episodic memories. In laboratory animals, most investigators report that estrogen enhances synaptic plasticity and improves performance on hippocampal-dependent cognitive behaviors. This review summarizes work conducted in our laboratory and others toward identifying estrogen's actions in the hippocampal formation, and the mechanisms for these actions. Physiologic and pharmacologic estrogen affects cognitive behavior in mammals, which may be applicable to human health and disease. The effects of estrogen in the hippocampal formation that lead to modulation of hippocampal function include effects on cell morphology, synapse formation, signaling, and excitability that have been studied in laboratory mice, rats, and primates. Finally, estrogen may signal through both nuclear and extranuclear hippocampal estrogen receptors to achieve its downstream effects.

Tianeptine increases brain-derived neurotrophic factor expression in the rat amygdala.

Chronic restraint stress affects hippocampal and amygdalar synaptic plasticity as determined by electrophysiological, morphological and behavioral measures, changes that are inhibited by some but not all antidepressants. The efficacy of some classes of antidepressants is proposed to involve increased phosphorylation of cAMP response element binding protein (CREB), leading to increased expression of neurotrophic factors, such as brain-derived neurotrophic factor (BDNF). Conversely, some studies suggest that acute and chronic stress downregulate BDNF expression and activity. Accordingly, the aim of the current study was to examine total and phosphorylated CREB (pCREB), as well as BDNF mRNA and protein levels in the hippocampus and amygdala of rats subjected to chronic restraint stress in the presence and absence of the antidepressant tianeptine. In the hippocampus, chronic restraint stress increased pCREB levels without affecting BDNF mRNA or protein expression. Tianeptine administration had no effect upon these measures in the hippocampus. In the amygdala, BDNF mRNA expression was not modulated in chronic restraint stress rats given saline in spite of increased pCREB levels. Conversely, BDNF mRNA levels were increased in the amygdala of chronic restraint stress/tianeptine rats in the absence of changes in pCREB levels when compared to non-stressed controls. Amygdalar BDNF protein increased while pCREB levels decreased in tianeptine-treated rats irrespective of stress conditions. Collectively, these results demonstrate that tianeptine concomitantly decreases pCREB while increasing BDNF expression in the rat amygdala, increases in neurotrophic factor expression that may participate in the enhancement of amygdalar synaptic plasticity mediated by tianeptine.

Deficient hippocampal c-fos expression results in reduced anxiety and altered response to chronic stress in female mice.

Stress response is an important neuroendocrine function. Overt or prolonged stress hormone secretion can lead to various disease states. The hippocampus plays an important role in the negative feedback onto the major player in the stress response, the hypothalamo-pituitary-adrenal axis. The transcription factor c-Fos is activated in the hippocampus following a number of stressors, including restraint stress. To determine whether c-fos modulates stress response, we previously generated mutant mice carrying a hippocampal mutation of the c-fos gene. In the current study, we found that female mutant mice display lower anxiety-like behavior than female wild-type mice in the elevated plus maze, whereas male mice are apparently normal. While both male and female mutant mice exhibit normal diurnal glucocorticoid (CORT) production and normal responses to acute restraint stress, female mutant mice habituated faster than female wild-type mice in response to chronic restraint stress. These findings suggest that hippocampal c-fos plays a role in gender-dependent response to stress.

Chronic restraint stress decreases the expression of glutathione S-transferase pi2 in the mouse hippocampus.

Chronic restraint stress in mice affects hippocampal structure and function. Mice were subjected to daily restraint for 3 weeks, and gene expression in hippocampus was compared to controls using large-scale cDNA microarrays. We found that 444 genes were differentially expressed, and further analysis of 6 genes by real-time reverse transcription PCR confirmed that 3 of them were downregulated by stress. These 3 genes, growth factor receptor-bound protein 2 (Grb2), phosphatidylinositol-4-phosphate 5-kinase, type 1 beta (Pip5k1b), and glutathione S-transferase, pi2 (Gstp2), were also analyzed by in situ hybridization. The downregulation of Gstp2 may induce an increase of oxidative damage in the pyramidal cells of the CA1 and CA3 regions and granular layer of the dentate gyrus, leading to structural and functional damage. Those regions are affected by stress, and our results could help understand further the mechanisms involved in the occurrence of stress-related disorders.

Regulation of nif expression in Methanococcus maripaludis: roles of the euryarchaeal repressor NrpR, 2-oxoglutarate, and two operators.

The methanogenic archaean Methanococcus maripaludis can use ammonia, alanine, or dinitrogen as a nitrogen source for growth. The euryarchaeal nitrogen repressor NrpR controls the expression of the nif (nitrogen fixation) operon, resulting in full repression with ammonia, intermediate repression with alanine, and derepression with dinitrogen. NrpR binds to two tandem operators in the nif promoter region, nifOR(1) and nifOR(2). Here we have undertaken both in vivo and in vitro approaches to study the way in which NrpR, nifOR(1), nifOR(2), and the effector 2-oxoglutarate (2OG) combine to regulate nif expression, leading to a comprehensive understanding of this archaeal regulatory system. We show that NrpR binds as a dimer to nifOR(1) and cooperatively as two dimers to both operators. Cooperative binding occurs only with both operators present. nifOR(1) has stronger binding and by itself can mediate the repression of nif transcription during growth on ammonia, unlike the weakly binding nifOR(2). However, nifOR(2) in combination with nifOR(1) is critical for intermediate repression during growth on alanine. Accordingly, NrpR binds to both operators together with higher affinity than to nifOR(1) alone. NrpR responds directly to 2OG, which weakens its binding to the operators. Hence, 2OG is an intracellular indicator of nitrogen deficiency and acts as an inducer of nif transcription via NrpR. This model is upheld by the recent finding (J. A. Dodsworth and J. A. Leigh, submitted for publication) in our laboratory that 2OG levels in M. maripaludis vary with growth on different nitrogen sources.

Stress-induced structural remodeling in hippocampus: prevention by lithium treatment.

Chronic restraint stress, psychosocial stress, as well as systemic or oral administration of the stress-hormone corticosterone induces a morphological reorganization in the rat hippocampus, in which adrenal steroids and excitatory amino acids mediate a reversible remodeling of apical dendrites on CA3 pyramidal cell neurons of the hippocampus. This stress-induced neuronal remodeling is accompanied also by behavioral changes, some of which can be prevented with selective antidepressant and anticonvulsive drug treatments. Lithium is an effective treatment for mood disorders and has neuroprotective effects, which may contribute to its therapeutic properties. Thus, we wanted to determine whether lithium treatment could prevent the effects of chronic stress on CA3 pyramidal cell neuroarchitecture and the associated molecular and behavioral measures. Chronic lithium treatment prevented the stress-induced decrease in dendritic length, as well as the stress-induced increase in glial glutamate transporter 1 (GLT-1) mRNA expression and the phosphorylation of cAMP-response element binding in the hippocampus. Lithium treatment, however, did not prevent stress effects on behavior in the open field or the plus-maze. These data demonstrate that chronic treatment with lithium can protect the hippocampus from potentially deleterious effects of chronic stress on glutamatergic activation, which may be relevant to its therapeutic efficacy in the treatment of major depressive disorder and bipolar disorder.