In no case was any evidence of contamination found. Furthermore, real-time PCR assays showed increases of the mmoX gene (encoding for the large hydroxylase subunit of the sMMO) that very closely corresponded with direct microscopic cell counts. Thus, for the first time, clear conclusive proof for the reality of facultative methanotrophy was provided. Remarkably, M. silvestris displayed higher yields, carbon conversion efficiency, Selleckchem Linsitinib and growth rates on acetate than on methane. Specifically,

the growth rate of M. silvestris was 0.053 and 0.033 h−1 on acetate and on methane, respectively, suggesting that acetate may be the preferred growth substrate for this microorganism. Shortly thereafter, another acidophilic methanotroph, Methylocapsa aurea, was also identified that could utilize acetate as the sole growth substrate (maximum OD600 nm=0.3, μ=0.006 h−1). As shown in Table 1, neither larger organic acids (citrate, oxalate, malate) nor any tested sugar (glucose, fructose, maltose) could be used as a sole growth substrate (Dunfield et al.,

2010). In contrast to M. silvestris, however, M. aurea only expresses pMMO. Strain purity was determined via: (1) phase-contrast and electron microscopy of acetate-grown cultures; (2) sequencing of more than 21 16S rRNA gene clones from both acetate- and methane-grown cultures; and (3) streaking onto medium with yeast extract and growing cultures with CH5424802 acetate in the absence of methane. In contrast to M. silvestris, however, M. aurea grew best on methane, with a maximum OD600 nm of 1.2 and μ=0.018 h−1. It is interesting to note that all these facultative methanotrophic species are not only acidophilic, but also members of the Beijerinckiaceae family known to include species with broad substrate

ranges. It could thus be hypothesized that facultative methanotrophy will only thrive in a small subset of acidophilic methanotrophs of this family, in environments where organic acids such as acetate are found primarily in the protonated form due to the prevailing low pH, and are thereby more readily taken up (Axe & Bailey, 1995). Facultative methanotrophy, however, does not extend to all acidophilic Phospholipase D1 methanotrophs of the Beijerinckiaceae family. For example, Methylocapsa acidophila cannot grow on multicarbon compounds such as malate, acetate, ethanol, succinate, or pyruvate (Dedysh et al., 2002, 2005; Dunfield et al., 2010). As a result of these findings, more effort has been spent to find other facultative methanotrophs, and in the past year, other acidophilic methanotrophs of the genus Methylocystis (family Methylocystaceae) were found that could grow on either methane or acetate (Belova et al., 2011). Specifically, Methylocystis strain H2s, a mild acidophile (optimal growth pH of 6.0–6.

Proportion of women who have commenced ART by beginning of week 24 of pregnancy. Proportion of women with a baseline HIV VL >30 000 RNA copies/mL mTOR inhibitor plasma and who do not require treatment

for themselves commencing temporary HAART at the beginning of the second trimester (by beginning of 16 weeks’ gestation). Proportion of women presenting in labour/with ROM/requiring delivery without a documented HIV result having an urgent HIV test result documented and this reactive/positive result acted upon immediately with initiation of the interventions to PMTCT without waiting for further/formal serological confirmation. Proportion of women with HBV coinfection who have LFTs performed 2 weeks after commencing HAART to detect evidence of ARV hepatotoxicity or IRIS. Proportion of women with HCV coinfection who have LFTs performed 2 weeks after commencing HAART to detect evidence of ARV hepatotoxicity or IRIS. Proportion of women

who have invasive prenatal diagnostic testing performed before their HIV status is known. Proportion of emergency CS performed and their indication. Proportion of infants <72 h old, born to untreated HIV-positive mothers, initiating three-drug therapy within 2 h of delivery. Proportion of routine neonatal PEP commenced within 4 h of delivery. Proportion of infants born to HIV-positive mothers who have HIV antibody testing for seroreversion performed at age 15–24 months. One of NVP-BGJ398 clinical trial the major successes in the management of HIV-positive patients has been the PMTCT of HIV-1. With the widespread implementation of routine antenatal screening GBA3 for HIV-1, transmission of HIV-1 from mother to child is now a rare occurrence in the UK. Despite few recent RCTs regarding the use of ART in pregnancy or obstetric intervention, practice continues to evolve. This is largely informed by observational data, theoretical considerations and expert

opinion. At the outset, the aim of the Writing Group was to make these guidelines as clinically relevant and as practical as possible. The Writing Group drew up a list of questions reflecting day-to-day practice and queries. It was acknowledged that the level of evidence for many of these topics was poor but recognized that there was a need to provide guidance. These guidelines have expanded on all areas relevant to the clinical care of HIV-positive pregnant women. The guidelines are intended to inform and aid healthcare workers in the management of pregnant women with HIV. They are not intended to be prescriptive or restrictive and it is recognized that situations will arise where the optimum management may deviate from these recommendations and new data will emerge to better inform practice. A particular focus has been obstetric management. An increasing number of women are aiming for and achieving a vaginal delivery but the rate of emergency CSs has increased.

, 2009). pLM100 with the mutY gene and pLM102 with the mutM gene were electroporated into PAOMY-Mgm separately as previously described (Mandsberg et al., 2009). The method was modified after Oliver (Oliver et al., 2000). To determine

the mutant frequency (MF) to rifampicin and streptomycin, an overnight culture of 20 mL LB media was centrifuged for 10 min at 6000 g, and resuspended in 1 mL of 0.9% NaCl. Serial dilutions of the bacterial culture were made, and 100 μL of appropriate dilution was spread on LB, 300 mg L−1 rifampicin and 500 mg L−1 streptomycin. The plates were incubated at 37 °C for 36 h before the CFU was determined. The CFU on selleck chemicals llc rifampicin and streptomycin plates were compared with the CFU from LB plates. At least five replicates were run for each experiment. The MRs are estimated using a fluctuation experiment, where a culture of each strain was diluted to 2 × 104 cells in 280 μL of LB and grown in 27 microtitre wells to stationary phase, then plated on 100 mg L−1 rifampicin LB agar plates to count the number of mutants. Three wells for each strain were used to estimate the CFU per

well. The expected number of mutations per well was then estimated using the Ma—Sandri—Sarkar (MSS) maximum likelihood method described by Sarkar et al. (1992). The MR was found by dividing the number of mutations by Selleck PFT�� the final CFU per well. The MIC was determined on 105 CFU mL−1 using the E-test system (AB Biodisk, Solna, Sweden), according to instructions of the manufacturer. Experiments were run at least in triplicates. To isolate ciprofloxacin resistant mutants, overnight cultures of PAO1 and PAOMY-Mgm were diluted and plated on 5% blood agar plates containing twofold dilutions of ciprofloxacin (Bagge et al., 2000; Mandsberg et al., 2009). Isolated resistant colonies were grown in LB without antibiotic, twice before E-test was performed with 100 μL of 10−4 dilution of an overnight culture. The strains were cultured in LB to an OD600nm = 1.0. Four millilitres of each culture was harvested, and RNA isolation and purification were performed using RNA Protect Urocanase Bacteria

Reagent and RNeasy Mini Kit (Qiagen, Hilden, Germany). RQ1 RNAse free DNAse (Promega, Madison, WI) was added to remove contaminating DNA. The experiment was run in triplicates. Processing of the P. aeruginosa GeneChip (Affymetrix) was performed at the Department of Clinical Biochemistry, Microarray Core Unit, Rigshospitalet, University of Copenhagen, Denmark. The gene expression analysis was done using arraystar v.3 Software (DNASTAR), http://isim.ku.dk/units/ub/research/paoym_vs_pao1microarray.pdf/. The level of expression of mexB, mexD, mexF and mexX was determined using real-time PCR in adapted isolates from the growth competition assays, and the level of expression of pfpI and PA5148 was determined to verify the microarray data. RNA from the logarithmic phase growth OD600 nm = 0.

In the

LH, single-labeled orexin-ir cells and cells double-labeled for both orexin and Fos, here called orexin/Fos-ir, were counted within an area defined by the presence of the orexin-ir cells. Therefore number, not density, of orexin-ir and orexin/Fos-ir cells per section is reported here. Double-labeled cells in the VTA and LH were not included in measures of Fos-ir cells to provide non-dopaminergic or non-orexinergic cell phenotype-specific insights. Measurements Crizotinib supplier from each tissue section were averaged across sections to create one measurement per subregion per hamster. With data from so many subregions within each hamster, one goal of our statistical approach was to simplify the data and present it at a circuit level by identifying clusters of regions that showed similar patterns of Fos expression across animals. To do so, we used a combination of factor analysis and descriptive correlational analyses to complement RG7420 ic50 previous functional and anatomical findings. Factor analysis, with principal axis factoring and a promax rotation, identified two clusters of subregions. Cluster 1 included Cg1, PrL, IL, AcbC, AcbSh, MePD, MePV, IF, PN, PBP and Tail, and we refer

to regions in this cluster as mesocorticolimbic. Cluster 2 included DM/PeF, LH, VMHM and VMHL, and we refer to regions in this cluster as hypothalamic. We then computed the correlations among the regions within each cluster as well as between the two clusters. The average within-cluster correlation was

0.34 in the mesocorticolimbic cluster and 0.42 in the hypothalamic cluster, based on 55 and six correlations, respectively. These indicate that Fos expression levels in subregions within the same cluster were consistently correlated with one another. We also examined correlations between regions falling into the two different clusters, and here the average of the 44 between-cluster correlations was 0.05, supporting the idea that Fos responses in these two clusters are relatively independent. Fos-ir cell density was next analysed with multilevel modeling treating animal as the upper-level sampling unit and brain region as the lower-level sampling unit. In this analysis, the cluster the region belonged to (mesocorticolimbic vs. hypothalamic) was treated as a within-subject variable, and age (juvenile vs. adult) and swab (blank vs. VS) were treated as between-subject PD184352 (CI-1040) independent variables. Multilevel modeling provides a more powerful analysis than a traditional repeated measures anova because it allows for analysis even if data from all subregions were unavailable for each hamster (as was the case in two juvenile and one adult hamsters due to poor quality tissue sections). The error structure was modeled to impose the traditional homoscedasticity assumption used in anova. Our hypotheses predicted that swab (blank vs. VS) will differentially affect Fos expression in adults and juvenile animals in some subregions.

However, both aerobic cellulose and cellobiose degradation were impaired by higher herbicide concentrations. The analysed bacterial taxa were also metabolically impaired under oxic conditions, which could suggest that they consumed less cellulose; however, the visibly present fungi compensated for

this loss of activity in the presence of pesticides. Agricultural soil is normally well aerated and has only small anoxic microzones. Impairment of anaerobic processes in such soils is probably of minor importance for the overall degradation of cellulose, as this process is mainly aerobic. However, when such soils become water-saturated due to rain, the observed GW-572016 solubility dmso toxic effect of Bentazon and MCPA on anaerobes may be of importance for cellulose degradation. The authors thank Christina Hirsch for technical assistance, M. Schloter, and S. Schulz (Technical University Munich) for providing selleck screening library soil, and the Deutsche Forschungsgemeinschaft (Priority Program 1315), and the University of Bayreuth for funding the study. “
“The DevSR two-component system in Mycobacterium smegmatis consists of the DevS histidine kinase and the DevR response regulator. It is a regulatory system that is involved in the adaptation of mycobacteria to hypoxic and NO stresses. Using the yeast two-hybrid assay and pull-down assay,

it was demonstrated that the phosphoaccepting Asp (Asp54) of DevR is important for protein–protein interactions between DevR and DevS. The negative charge of Asp54 of DevR was shown to play an important role in protein–protein interactions between DevR and DevS. When the Lys104 residue, which is involved in transmission of conformational changes induced by phosphorylation of the response regulator, was replaced with Ala, the mutant form of DevR was not phosphorylated by DevS and functionally inactive in vivo. However, the K104A mutation in

DevR only slightly affected protein–protein interactions between DevR and Atezolizumab manufacturer DevS. “
“Depth-related changes in bacterial community structures and functions were analyzed in a paddy soil profile using denaturing gradient gel electrophoresis (DGGE) and a metabolic profiling technique (BIOLOG ECO plates). Canonical correspondence analysis (CCA) was used to analyze the correlations between the relative abundance of bacterial groups and soil-available elements. DGGE and sequencing analysis revealed 12 classes and one unknown bacterial group. At the family level, Comamonadaceae and Moraxellaceae dominated through the soil profile, while Acidobacteriaceae and Nitrospiraceae dominated in the deepest layer. In addition, Streptococcaceae dominated and was only observed in the deeper layers. Metabolic profiles revealed the greatest carbon source utilization capacity in the surface layer, and no significant differences between upper and deeper soil layers. The carbon sources utilized by microorganisms were different among the different layers.

In this study, we have click here investigated the role of activity directly by measuring changes in medial nucleus of the

trapezoid body (MNTB) neurons in normal hearing mice subjected to 1-h sound stimulation. Broadband (4–12 kHz) chirps were used to activate MNTB neurons tonotopically restricted to the lateral MNTB, as confirmed by c-Fos-immunoreactivity. Following 1-h sound stimulation a substantial increase in Kv3.1b-immunoreactivity was measured in the lateral region of the MNTB, which lasted for 2 h before returning to control levels. Electrophysiological patch-clamp recordings in brainstem slices revealed an increase in high-threshold potassium currents in the lateral MNTB of sound-stimulated mice. Current-clamp and dynamic-clamp experiments

showed that MNTB cells from the sound-stimulated mice were able to maintain briefer action potentials during high-frequency firing than cells from control mice. These results provide evidence that acoustically Fostamatinib clinical trial driven auditory activity can selectively regulate high-threshold potassium currents in the MNTB of normal hearing mice, likely due to an increased membrane expression of Kv3.1b channels. “
“The transition between biofilm and planktonic cells has important consequences during infection. As a model system, we have investigated uropathogenic Escherichia coli (UPEC) strain 536, which forms large biofilm aggregates when grown in iron-restricted tissue culture media. The provision AZD9291 research buy of both inorganic and physiological iron to the media induces dispersal. Aggregates do not disperse upon the addition of exogenous iron when cells are pretreated with either rifampicin or chloramphenicol as inhibitors

of transcription or translation, respectively. Aggregates stain with the cellulose stain Calcofluor White, can be prevented by the addition of cellulase to the growth media, and aggregates are broken down in the absence of exogenous iron when cellulase is added. An extension of this study to 12 UPEC clinical isolates identified seven that form cellulose aggregates under iron restriction, and that disperse upon the provision of iron. Consequently, we hypothesize that iron restriction stimulates the formation of cellulose aggregates, which disperse as a result of new gene expression in response to the provision of iron. An infection is a dynamic process whereby a pathogen will colonize the host, encounter and evade immune killing and acquire nutrients to proliferate. A successful pathogen is able to adapt to its changing environment, and especially to those changes that occur in response to bacterial activities and the damage caused by the pathogen. In the study of bacterial infections, it is important to be aware of the changes that may occur in the environment of the bacterial population during the progression of an infection. Prominent among these changes is the availability of iron, which is an essential nutrient as an enzyme cofactor for most bacteria (Schaible & Kaufmann, 2004).

Cataplexy-like episodes were not observed. The percentage time spent in wakefulness and non-REM (NREM)

DNA Damage inhibitor sleep and the power spectral profile of NREM and REM sleep were unaffected. Control animals, injected with scrambled siRNA, had no sleep changes after injection. Quantification of the knockdown revealed that unilateral microinjection of siRNAs targeting OxR1 into the rat LC on two consecutive days induced a 45.5% reduction of OxR1 mRNA in the LC 2 days following the injections when compared with the contralateral side receiving injections of control (scrambled) siRNAs. This reduction disappeared 4 days after injection. Similarly, unilateral injection of OxR1 siRNA into the LC revealed a marked (33.5%) reduction of OxR1 staining 2 days following injections. In contrast, both the mRNA level and immunohistochemical staining for tyrosine hydroxylase were unaffected. The results indicate that a modest knockdown of OxR1 is sufficient to induce observable http://www.selleckchem.com/products/DAPT-GSI-IX.html sleep changes. Moreover, orexin neurons, by acting on OxR1 in the LC, play a role in the diurnal gating of REM sleep. “
“Stimulation of the vagus nerve produces antiepileptic effects. This is used clinically to treat drug-refractory epilepsies. The mechanisms responsible for these effects depend

on the activation of vagal afferents reaching the nucleus of the solitary tract. This review focuses on the neuroanatomy of the nucleus of the solitary tract and its relation with the nucleus locus coeruleus as a preferential anatomical substrate in producing antiepileptic effects. In fact, following the transient or permanent inactivation of locus coeruleus neurons, some antiepileptic effects of vagus nerve stimulation are lost. The activation of locus coeruleus per se is known to limit the spread of a seizure and the duration of a variety of seizure types. This is due to the fine chemical neuroanatomy of norepinephrine pathways that arise from the locus coeruleus, which produce widespread changes in cortical areas. These

ADP ribosylation factor changes may be sustained by norepinephrine alone, or in combination with its co-transmitters. In addition, vagus nerve stimulation may prevent seizures by activating the serotonin-containing dorsal raphe neurons. “
“Potassium channels comprise the most diverse family of ion channels and play critical roles in a large variety of physiological and pathological processes. In addition to their molecular diversity, variations in their distributions and densities on the axo-somato-dendritic surface of neurons are key parameters in determining their functional impact. Despite extensive electrophysiological and anatomical investigations, the exact location and densities of most K+ channels in small subcellular compartments are still unknown.

Phenylketonuric (PKU) and epileptic mice show altered expression of NIPSNAP1 in the brain. Therefore, the distribution and localization of NIPSNAP1 in rat brain was determined. Results show that NIPSNAP1 is expressed exclusively in neurons including pyramidal neurons in the cerebral cortex, Purkinje neurons in the cerebellum and motor neurons in the spinal cord. Dopaminergic neurons in midbrain and noradrenergic VX-770 supplier neurons in the brainstem, which are affected in PKU, also express NIPSNAP1. NIPSNAP1 is found to be localized in the mitochondrial matrix and can bind dihydrolipoyl-transacylase and -transacetylase components of the BCKA and pyruvate

dehydrogenase complexes in vitro. Our data provide the first experimental evidence for a strictly neuronal expression of this mitochondrial protein in the rat nervous system. “
“Temporal order memory (memory for stimulus order) is crucial for discrimination between familiar objects and depends upon a neural circuit involving the perirhinal cortex (PRH) and medial pre-frontal cortex. This study examined the role of glutamatergic and cholinergic neurotransmission in the encoding or retrieval of temporal order memory, using a task requiring the animals to discriminate between two familiar objects presented

at different intervals. 6-Cyano-7-nitroquinoxaline (CNQX) (AMPA/kainate receptor antagonist), scopolamine (muscarinic receptor antagonist) or 2-amino-5-phosphonopentanoic acid (AP5) (N-methyl-D-aspartate http://www.selleckchem.com/products/icg-001.html receptor antagonist) was administered before sample phase 2 (to be active during encoding) or before test (to be active during retrieval). Unilateral CNQX administration into the PRH and pre-limbic/infra-limbic old cortices (PL/IL) in opposite hemispheres, i.e. to disrupt neurotransmission within the circuit, impaired encoding and retrieval. Administration of scopolamine or AP5 in the PRH–PL/IL circuit impaired encoding. Drug effects in each brain region were then investigated

separately. Intra-PRH CNQX, scopolamine or AP5 disrupted encoding, such that the animals explored the recent object significantly more than the old object. In contrast, intra-PL/IL CNQX, scopolamine or AP5 impaired memory performance such that the animals spent an equal amount of time exploring the objects. CNQX but not AP5 or scopolamine impaired retrieval. Furthermore, CNQX impaired novel object preference when infused into the PRH but not PL/IL following a 3 h delay. Thus, encoding of temporal order memory is mediated by plastic processes involving N-methyl-D-aspartate and muscarinic receptors within the PRH–PL/IL circuit, but these two regions make qualitatively different cognitive contributions to the formation of this memory process.

For some time it was though that

some CB1 antagonists act as inverse agonists (i.e., by blocking a constitutive activity of the CB1 receptors), but the current consensus is that the effects of CB1 antagonists can be attributed solely to blockade of the effects of endocannabinoids (Savinainen et al., 2003; Kano et al., 2009). For example, the basal activity of CB1 receptors was decreased by inhibition of diacylglycerol lipase, the enzyme H 89 that synthesizes the endocannabinoid 2-archidonyl-glycerol (Turu et al., 2007). Accordingly, our results indicate that endocannabinoids are present in the dorsal horn, possibly because their synthesis is triggered by the stimulus used to evoked substance P release. The most likely explanation for the facilitation of substance P release by CB1 receptors is the disinhibition mechanism depicted in Fig. 10. According to this model, the CB1 receptors

producing this effect are located in the presynaptic terminals of GABAergic and opioidergic interneurons in the dorsal horn, where they inhibit neurotransmitter release. As substance P release from primary afferent terminals is inhibited by μ-opioid receptors (Yaksh et al., 1980; Aimone & Yaksh, 1989; Kondo et al., 2005) and GABAB receptors (Malcangio & Bowery, 1993; Marvizon et al., 1999; Riley et al., 2001), reduced agonist binding to these receptors results in a facilitation of substance P find more release. Several lines of evidence support this model. First, it is unlikely that the facilitation of substance P release is mediated by CB1 receptors located in the substance P-containing terminals themselves. While CB1 receptors frequently inhibit neurotransmitter release, no instances of direct facilitation

of neurotransmitter release by this receptor has been found (Kano et al., 2009). Whether CB1 receptors are present in Fossariinae the central terminals of primary afferent terminals has been controversial until recently. Initially, CB1 receptor mRNA and immunoreactivity was detected in some DRG neurons (Hohmann & Herkenham, 1999; Bridges et al., 2003; Binzen et al., 2006; Agarwal et al., 2007). However, other studies found that CB1 receptor immunoreactivity in the dorsal horn was unaffected by rhizotomy (Farquhar-Smith et al., 2000) or by selective CB1 receptor knockout in DRG neurons (Agarwal et al., 2007), suggesting that CB1 receptors may not be transported centrally from the DRG. However, a recent studied (Nyilas et al., 2009) provided solid evidence for the presence of CB1 receptors in C-fiber and Aδ-fiber terminals in the dorsal horn. It remains to be clarified whether CB1 receptors are present in C-fiber terminals that contain substance P (Farquhar-Smith et al., 2000; Khasabova et al., 2004).

, 1997) using S. oneidensis MtrB as the search query. Multiple alignments of MtrB homologs were generated with clustalw (http://www.ebi.ac.uk/Tools/clustalw2/index.html)

(Chenna et al., 2003). β-Barrel architecture of the MtrB homologs was predicted http://www.selleckchem.com/products/crenolanib-cp-868596.html using the program pred-tmbb (Bagos et al., 2004). logo diagrams were generated using the clustalw alignment files (Crooks et al., 2004). mtrB was deleted from the S. oneidensis genome via application of a Shewanella in-frame gene deletion system (Burns & DiChristina, 2009). Regions corresponding to c. 750 bp upstream and downstream of mtrB were independently PCR-amplified and subsequently joined using overlap-extension PCR. Primers for mtrB deletion are listed in Table 2. The resulting fragment was cloned into suicide vector pKO2.0, which does DMXAA ic50 not replicate in S. oneidensis. This construct (designated pKO-mtrB) was

mobilized into wild-type MR-1 via conjugal transfer from E. coli donor strain β2155 λ pir. S. oneidensis strains with the plasmid integrated into the genome were selected on solid LB medium containing gentamycin (15 μg mL−1). Single integrations were verified via PCR with primers flanking the recombination region. Plasmids were resolved from the genomes of single integrants by plating on solid LB medium containing sucrose (10% w/v) with NaCl omitted. In-frame deletions were verified by PCR and direct DNA sequencing (GeneWiz, South Plainfield, NJ). Genetic complementation of ∆mtrB was carried out by cloning wild-type mtrB into broad-host-range cloning vector pBBR1MCS (Kovach et al., 1995) and conjugally transferring the recombinant vector into ∆mtrB via biparental mating procedures (DiChristina et al., 2002). Single amino acid mutations in MtrB (C42A or C45A) were constructed using the Quickchange Lightning site-directed mutagenesis kit (Agilent Technologies, Santa Clara, CA). The mtrB gene and regions c. 750 bp upstream and downstream were PCR-amplified as a single fragment and subsequently cloned into pBBR1MCS. Mutagenesis primers C42A-sense, C42A-antisense, C45A-sense, and C45A-antisense (Table 2) were used Staurosporine datasheet in mutagenesis PCR

according to the manufacturer’s instructions. The resulting PCR products were subsequently transformed into XL10 Gold KanR competent cells (Agilent Technologies). Correct amino acid mutations (C42A or C45A) were verified by direct DNA sequencing using primers MTRB-SeqF and MTRB-SeqR (Table 2). The mutated mtrB constructs were subsequently cloned into suicide vector pKO2.0 and were ‘knocked in’ to the native chromosomal position. Nucleotide sequence changes were verified by PCR and DNA sequencing of S. oneidensis ‘knock-in’ transformants. Genetic complementation of mutant C42A was carried out by cloning wild-type mtrB into broad-host-range cloning vector pBBR1MCS (Kovach et al., 1995) and conjugally transferring the recombinant vector into mutant C42A via biparental mating procedures (DiChristina et al., 2002).