those from iron-starved cells at 26°C (stationary and exponential

those from iron-starved cells at 26°C (stationary and exponential phase, respectively; Table 4). Table 4 Reaction rates for four Y. pestis enzyme classes comparing -Fe vs. +Fe conditions   Reaction ratea) (nmol min-1 mL-1); (U mL-1)b) Reaction ratea) (nmol min-1 mL-1); (U mL-1)b) Enzyme +Fe, exp, n = 4 e) -Fe, early, n = 5 e) p-value f) +Fe, stat, n = 4 e) -Fe, late, n = 5 e) p-value f) Aconitase c) 2.31 1.14 0.019 4.98 1.82 0.008 Pyruvate oxidase

c) 167.5 1307 0.0001 463.0 2405 click here 0.0001 Catalase d) 82.5 31.8 0.0001 93.4 29.0 0.0001 Superoxide dismutase d) 887.8 426.9 0.002 448.5 312.5 0.234 a) Spectrophotometric assays in 96-well microtiter plates were used for the determination of enzyme reaction rates. Total protein concentrations

in crude cell lysates were the same for all samples used in a given enzyme assay: aconitase, 0.5 mg/mL; pyruvate oxidase, 0.4 mg/mL; catalase, 0.15 mg/mL; superoxide dismutase, 1.1 μg/mL. b) Units ml-1 was the definition for the superoxide dismutase reaction rate. All assays were performed in duplicate. c) Reaction rates from the linear part of the slope of the absorbance change over time. d) Reaction rates from endpoint assays. e) Number of biological replicates of cell lysates (n); exp: abbreviation for exponential, early: early growth phase equivalent to exp. phase (-Fe); average OD600 = 0.66 (+Fe) and OD600 = 0.47 (-Fe); stat: abbreviation for stationary growth phase, late: late growth phase equivalent Nutlin-3a datasheet to stat. phase (-Fe); average OD600 = 2.0 (+Fe) and OD600 = 0.81 (-Fe). True exponential and stationary growth phases were not observed for cell cultures in iron-free media. f) p-values were calculated from to comparison of reaction

rates (+ Fe vs. -Fe) using a two-tailed t-test method. The question STK38 arose whether iron-starved Y. pestis cells activated a different metabolic route of pyruvate degradation able to produce reducing equivalents (NADH and UQH2) for the electron transport chain. Pyruvate oxidase (PoxB) degrades pyruvate to acetate and is a flavin-dependent, iron-independent enzyme that generates UQH2 [52]. The pyruvate oxidase pathway indeed appeared to be important, as judged by the strong abundance increase of PoxB#39 (Figure 4) under -Fe conditions. The flavin cofactor may be recruited from redox activities of two flavodoxins. FldA3#44 was quite abundant and moderately increased in iron-depleted cells (Figure 4). FldA was identified in faint 2D spots and not reproducibly quantitated. PoxB activity measurements revealed excellent correlation between enhanced abundances and increased reaction rates in iron-starved cells. PoxB activities were 5.3-fold and 7.8-fold higher in lysates of iron-starved cells than in lysates of iron-replete cells at 26°C (stationary and exponential phase, respectively; Table 4). Electron transport chains are localized in the IM, a fact that compromised the analysis of subunits of these IM protein complexes in 2D gels.

For region upstream from the arp2 gene (B), horizontal lines belo

For region upstream from the arp2 gene (B), horizontal lines below learn more the sequences delimitate the putative stems regions and dashed lines indicate the loop part. To determine which genes were co-transcribed, RT-PCR amplification of core region was performed by grouping ORFs two by two or three by three. For ICESt1, amplifications of orfR/arp1/orfQ and orfP/arp2, respectively, were positive while that of the orfQ/orfP junction was negative (see additional file 1: S1B). These data comfort the hypothesis of a two-operon organization for ICESt1 (see additional file 1: S1A) with a functional rho-independent transcription

terminator located between the two operons. By contrast, for ICESt3, all the RT-PCR amplifications of the regulation module were positive (see additional file 1: S1D) indicating a co-transcription of all the regulation genes (see additional file 1: S1C). The free energy of the transcriptional terminator detected between orf385B and orfQ genes in ICESt3 (Figure 1) was calculated with the mFold software [19]. It is different from the one for ICESt1 (ΔG = -4.3 kcal.mol-1 for ICESt3 and ΔG = -8.2 kcal.mol-1 for ICESt1). This difference could explain why all genes of the regulation module of ICESt3 can be co-transcribed while two independent transcriptional units were found in ICESt1. We then examined the

activity of the promoter located upstream from the orfQ gene by Rapid Amplification of cDNA ends (5′ RACE). For both elements, the start point (A nucleotide) was located seven nucleotides downstream from a -10 box separated by 17 nt PX-478 mouse from a -35 box, which overlapped the rho-independent transcription terminator (Figure 1A). This result is consistent with the S. thermophilus promoter consensus sequence (TTGACA – 17 nt – TATAAT) [20]. Therefore, both ICEs possess a functional PorfQ promoter. However, it was previously showed that ICESt3 differs from ICESt1 by a -1 frameshift in the 5′ end of its orfQ gene (orfQ1) [11]. A second RBS, that could enable the translation from an initiation codon located downstream, was identified in silico (Figure 1A). All together, buy Staurosporine these data suggest that

the orfQ2 gene of ICESt3 is truncated of 54 nucleotides at its 5′ end compared to the orfQ gene of ICESt1. All RT-PCR amplifications targeting co-transcription of the sixteen conjugation-recombination genes of ICESt1 and ICESt3 gave amplicons (see additional file 1: S1B and S1D). Therefore, these genes are transcribed as a single polycistronic mRNA of about 14.6 kb (see additional file 1: S1A and S1C). To map more precisely the 5′ end of these transcripts, other sets of primers were designed in the arp2/orfN intergenic region. For ICESt1, these results (data not shown) combined with 5′ RACE experiments confirmed the predicted conjugation-recombination promoter, Pcr, with a -10 box (TATAAT) located seven nucleotides upstream from the transcription start point (A) nucleotide (Figure 1B).

To increase the efficiency of combined treatments, particularly t

To increase the efficiency of combined treatments, particularly the combination of DTIC and protons, the order of administration of drugs and radiation was inversed. The new experimental set up was conceived knowing the position on the time scale where the best effect of each single treatment with FM, DTIC or protons was reached [10]. The HTB140 cells were irradiated with protons, incubated for 4 days, when FM or DTIC was added to the cells, and then incubated for another 3 days. In this way it was enabled that the incubation periods providing the best single effects of protons and

drugs coincide at the same time. The described combination of protons and FM reduced cell proliferation to ~40% and clonogenic survival to ~50%, while there was ~80% of viable cells estimated by the SRB assay (Figure 1). With respect to the single treatments the obtained effects were weaker. The Fludarabine concentration time interval between irradiation and drug treatment might be considered as long because the multiplicity of microcolonies 4 days after irradiation could underestimate the effects of drug treatment, particularly for the clonogenic assay. An overestimation of cell viability by the SRB assay could be ascribed to the excess of proteins coming from the dead cells that were indistinguishable from those of surviving cells [23]. DNA damaging agents also produce morphological changes of cells, such as an increased cell size and therefore

protein content [29]. This might also explain the overestimated viability obtained by the SRB assay. Comparing the inactivation levels obtained in this experiment to those of the two experiments that PRIMA-1MET were previously described [11, 12], the best effect was obtained when the HTB140 cells were treated with FM before proton irradiation and incubated for 7 days [12]. The combination of protons and DTIC reduced cell proliferation to ~32% while after single treatments this level was higher (Figure 2). Again, an overestimation of viability was obtained by SRB assay [23, 29]. According to cell proliferation

and survival, the poor efficiency of the single DTIC treatment was overcome when it was introduced following proton irradiation. The cells that were damaged by protons and would most likely survive were additionally damaged in a similar way by the DTIC treatment [30]. Rutecarpine As a result, the obtained cell inactivation levels were better than those of the two previously reported experiments [11, 12]. Analysing the effects of the two administration procedures of radiation and drugs, in general there was not an appreciable improvement with respect to the single treatments. In each of them there was a moderate improvement with the combination of just one drug and radiation. All studied agents affect cellular DNA, but they differ in the type of damage they induce. Protons, as well as conventional radiation, induce oxidative changes in DNA bases together with the single- and double-strand breaks [31].

The ratio

between chlorophyll fluorescence at 735 nm and

The ratio

between chlorophyll fluorescence at 735 nm and that at 700 nm (F735/F700) is linearly proportional to chlorophyll content (Gitelson et al. 1999). Conversely, as discussed in Question 24, the F M and F O values are not related to the chlorophyll content in leaves (Dinç et al. 2012). It may also be noted that there are simple chlorophyll meters on the market (CL-01, Hansatech Instruments, UK; SPAD meter, Minolta, Japan; CCM-200, Opti-Sciences, USA) that can be learn more used to follow changes in the leaf chlorophyll content (see e.g., Cassol et al. 2008; Dinç et al. 2012). These measurements can then be calibrated against measurements of the chlorophyll extracted from leaf areas measured before with the chlorophyll

meter (see e.g., Dinç et al. 2012). Chl measurements on dark-adapted leaves seem to give more reproducible results than measurements made on light-adapted leaves (Ceppi and Schansker, unpublished data, 2008). If the chlorophyll meter is used over the day on the same leaf, the readings change (Mishra, unpublished data, 2010), e.g., due to chloroplast movements, which change the absorbance properties of the leaf (see Wada 2013 for a review on chloroplast movements). Chloroplasts are known ABT-263 research buy to re-arrange themselves inside the cell in response to the ambient blue light intensity, adapting the absorbance properties of the leaf to the circumstances (Sakai et al. 2001; Kasahara et al. 2002). This does not only affect chlorophyll meter measurements, but also normal fluorescence measurements (Brugnoli and Björkman 1992). In practice, values measured using Quisqualic acid a Chl meter are often used as indicators for relative Chl changes. In that case, we assume that the measured values are a linear function of the leaf chlorophyll content between zero and the value measured on control leaves. However, in that case, it is important to test the validity of this assumption for each plant species and for each stress studied (Mishra, unpublished data, 2013). Question 26. Is it possible

to compare different leaves? It is easy to take randomly two leaves from two plants of the same species and to make a fluorescence measurement. But is it truly possible to compare these two measurements? It is likely that a difference in maximum fluorescence amplitude will be observed. Especially, when studying OJIP transients, the kinetics are often more interesting than the absolute amplitude, and in that case, the difference in the fluorescence amplitude is eliminated by double normalization between F O and F M. Arithmetically, this is done in the following way: (F t − FO)/(F M − F O). The effect of this calculation is to rescale each fluorescence value in a range going from 0 (corresponding to F O) to 1 (corresponding to F M). For a comparison of the kinetics of the individual rise phases of the OJIP transient, the same approach can be used.

1, 3, 6, 19 and 21, who were infected by 2 genotypes, still have

1, 3, 6, 19 and 21, who were infected by 2 genotypes, still have a major one across both gastric niches, and that was also true in 2 (no. 14, 27) patients having 3, and 1 (no. 17) patient having 4 genotypes represented in their infections. – indicating that the patients have non-dominant babA and babB genotype in the isolates of antrum or corpus. The patients’ number was according to our previous study [22]. Among those 12 patients infected with more than one genotype (Table 2), Trichostatin A in vivo the frequency of the major dominant genotype, A B combined with AB AB, in the antrum

was higher compared with that in the corpus (75% [9/12] vs. 16.2% [2/12], p = 0.012, odds ratio: 15). However, 6 of 12 patients lacked a dominant genotype in their corpus isolates. Sequence analysis and comparison At locus A, each patient’s antrum and corpus isolates had specific substitutions of amino acids in the region of BabA (Figure 2 and Table 3). However, there was no obvious difference between the antrum and corpus isolates in the sequencing region, except from patient no. 27 (amino acid 134 and 198). We also found 5 different nonsynonymous substitutions at amino acid 161 in 6 patients’ isolates, as compared with strain J99. The same scenario (sequence specificity in individual patients’ strains but not between the antrum and corpus isolates) was in the babB sequences. Figure 1 PCR banding patterns of babAB genotypes. (A) Primer pairs used for gene detection at locus A and B. The forward primers, HypDF1 and S18F1, located in the upstream region of babA or babB, are paired with BabAR1 or

BabBR1 Amrubicin primers to determine whether the gene at locus A and B is babA or babB. (B) PCR banding patterns of genotype A B, AB B, A AB and AB AB. The AB B genotype showed two bacterial populations in the single-colony isolate, the dominant as babA and the minor as babB, at locus A. The strain with A AB genotype represented a dominant population of babB and a minor population of babA at locus B. The combination of AB B and A AB was defined as an AB AB genotype. Lane M1, a 100 bp molecular marker; lane M2, l HindIII marker. The size of PCR products at locus A and B was 2.1-2.6 kb and 1.0-1.5 kb, respectively. Table 3 The amino acid substitutions in BabA encoded by babA at locus A   The location of amino acid substitutions Case No.

1, 0 1) eV The yellow-curved isosurfaces stand for the charge de

1, 0.1) eV. The yellow-curved isosurfaces stand for the charge density of 0.6 a.u.-3. Since VOs are more likely to form at the subsurface (LaO layer) than the surface in the Pd-containing FeO2-terminated surface, we placed another VO in the same LaO layer (Figure  2 group III (a) to (c)). If two VOs are both located at the subsurface, the second Pd atom tends to substitute the Fe atom either at the second FeO2 layer forming a pair of Pd atoms (Figure  2 group III (b)) or on the surface

forming the PdO2 layer (Figure  2 group III (c)). The difference in energy between these two configurations is less than 0.05 eV. Thus, the additional VO stabilizes the PdO2-layer exposed to the vacuum. Thus Selleckchem PR 171 far, we have assumed the existence of VO. However, the concentration of VOs depends on their formation energy. Therefore, we have to verify the stability of surfaces containing VO(s) with different concentrations of Pd by taking the formation energy of VOs into account to further strengthen our conclusion. We calculated the relative energies of surfaces containing Pd m VOn (m =1 and 2 and n =0, 1, and 2) relative to the perfect slab (without VOs) with Pd inside the bulk of LFO (see Figure  2 group

I (a)). Note that the systems we have discussed here are surfaces with Pd atom(s) and VO(s) located on/near the surface. The relative energies (ΔE rel) as a function of the chemical potential of oxygen (Δμ O ) are shown in Figure  4. The corresponding SB431542 purchase geometries for the Pd m VOn -containing surfaces are all included in Figure  2. Since two Pd atoms fail to segregate near/at the surface adjacently without VOs, the results for the Pd2-containing Cediranib (AZD2171) perfect surface excluded from Figure  4. The ΔE rel for each colored line is calculated as: (1) (2) (3) (4) (5) where the first items in Equations 1 to 5 on the right-hand side are the total energies of the Pd m VOn -containing

(m =1 and 2 and n =0, 1, and 2) surfaces, with their subscripts describing their compositions. represents the total energy of the reference surface that contains one solid-solution state of Pd inside the bulk. The in Equations 4 and 5 is the total energy of the pristine FeO2-terminated surface. The μ O is the chemical potential of oxygen. The chemical potentials of oxygen in the LFO bulk and gas phase are equal under equilibrium conditions. The μ O based on an ideal gas approximation is directly connected with the partial pressure (p (O2)) and temperature (T) by (6) (7) in which k is the Boltzmann constant and p 0 is taken to be the standard pressure. is the total energy of an isolated O2 molecule. The item is determined by using thermodynamic data from the thermochemical tables [26]. Hence, we can obtain the formation energy of VO(s) based on Equations 2 to 5 by subtracting the item in Equation 1.

Three separate experiments

showed consistent results and

Three separate experiments

showed consistent results and representative examples are shown. Standard deviation represents variation between biological replicates. Selleck PP2 Asterisks indicate significant differences (P ≤ 0.05) in accumulation compared with the parental isolate or with addition of an EI. Panel A, Fold-change in level of ethidium bromide accumulated by R2 and mutants. Panel B, Fold-change in level of ethidium bromide accumulated by R2 and mutants with addition of EIs. Panel C, Fold-change in level of ethidium bromide accumulated by DB and mutants. Panel D, Fold-change in level of ethidium bromide accumulated by DB and mutants with addition of EIs. Dark grey, selleck chemicals llc no EI; light grey, CCCP; white, PAβN. Discussion The two-step deletion strategy we have described was used for creating unmarked deletions in the adeFGH and adeIJK efflux pump operons, separately and together, in two clinical MDR A. baumannii isolates. It is an improvement from the simple method for gene replacement in A. baumannii described by Aranda et al (2010) that uses an antibiotic resistance cassette [12]. To adapt the method first described for use in MDR A. baumannii, we introduced a tellurite resistance cassette into the pMo130 suicide vector created by Hamad et al (2009) to facilitate the selection of MDR A. baumannii transconjugants with the suicide plasmid inserted

into the genome, i.e. first crossover products [8]. It was helpful to first ascertain the growth inhibitory concentration of tellurite for the parental A. baumannii strain so the number of transconjugants (first crossover) that are false positives can be minimized by using a suitable tellurite concentration. Passaging the first crossover recombinants in media containing sucrose provided the selection pressure for loss of the plasmid by a second crossover, leading to the formation of white colonies when sprayed with pyrocathechol. The main advantage of this method, which does not use antibiotic selection for the gene deletion mutants, Vasopressin Receptor is its application for generating multiple gene deletions in a single strain as we have

demonstrated by creating DBΔadeFGHΔadeIJK and R2ΔadeFGHΔadeIJK mutants. This is particularly important because the majority of A. baumannii strains are MDR or extensively drug-resistant (XDR). Other than the MDR strains described in this study, we have also tested this method in a carbapenem-susceptible A. baumannii strain (data not shown). Un-marked deletion mutants are especially useful for ascertaining the contribution of each efflux pump to MDR as the presence of antibiotic resistance cassettes in the mutants may complicate the interpretation of antimicrobial susceptibility. We believe that the marker-less method would allow the impact of each efflux system on antimicrobial resistance to be clearly defined.

Ekberg and Wildhagen (1996) found that long-term sickness absence

Ekberg and Wildhagen (1996) found that long-term sickness absence is associated with worse ratings in quality of life after 1-year and that pain did not diminish during the follow-up year. Information on the severity and impact of the diseases is important for decision-making GDC-0973 manufacturer in preventive policy. Moreover, the incidence rate, the severity

and the impact of a disease can provide arguments when deciding for which diseases preventive activities should be financed. In general, registries of occupational diseases do not provide information on the severity or impact of the diseases (Karjalainen and Niederlaender 2004). Despite variations in registration guidelines in different countries, general occupational disease registries probably contain the relatively more severe cases of occupational disease, which result in relatively higher costs. Therefore, it might be relevant Idasanutlin purchase for policy purposes to perform follow-up studies of the cases from registries. In addition, periodically executed small-scale follow-up studies linked to registries will probably

be less expensive and more efficient than a series of cohort studies. The aim of this study was to investigate the perceived severity and the consequences of the upper extremity disorders that are registered as occupational diseases. Severity, functional impairment, quality of life and sickness absence were assessed over the course of 1 year and compared with reference data on the general working population. Methods Population In the Netherlands, occupational physicians are obliged to notify cases of occupational diseases to the registry of the NCvB. Besides classic occupational diseases like occupational asthma or mesothelioma, this registry also covers work-related diseases like work-related depression or musculoskeletal diseases. The registry distinguishes eleven categories of work-related specific disorders of the upper extremity: radiating neck complaints; rotator cuff syndrome; epicondylitis (lateral and medial); ulnar nerve compression at the elbow (cubital tunnel syndrome); radial nerve compression

(radial tunnel syndrome); flexor–extensor peritendinitis or tenosynovitis of the forearm–wrist Cell press region; de Quervain’s disease; carpal tunnel syndrome; ulnar nerve compression at the wrist (Guyon canal syndrome); Raynaud’s phenomenon and peripheral neuropathy associated with hand-arm vibration; and osteoarthrosis of distal upper extremity joints. In addition, a twelfth category of non-specific upper extremity musculoskeletal disorders has been described (Sluiter et al. 2001). We asked occupational physicians, who had participated in an NCvB sentinel surveillance project, to recruit patients, who had been diagnosed with a work-related upper extremity disorder, to participate in this study and to ask them to fill out an informed consent form. After signing the form, the patients received a questionnaire.

J Vasc Interv Radiol 2008, 19:1693–1698 PubMedCrossRef 24 Fava M

J Vasc Interv Radiol 2008, 19:1693–1698.PubMedCrossRef 24. Fava M, Meneses L, Loyola S, Tevah J, Bertoni H, Huete I, Mellado P: Carotid artery dissection: endovascular treatment. Report of 12 patients. Catheter Cardiovasc Interv 2008, 71:694–700.PubMedCrossRef 25. Schulte S, Donas KP, Pitoulias GA, Horsch S: Endovascular treatment of iatrogenic and traumatic carotid artery dissection. Cardiovasc Intervent Radiol 2008, 31:870–874.PubMedCrossRef 26. DuBose J, Recinos G, Teixeira PG, Inaba K, Demetriades D: Endovascular stenting for the treatment of traumatic internal carotid injuries: expanding

experience. J Trauma 2008, 65:1561–1566.PubMedCrossRef 27. Siomin V, Angelov L, Li L, Vogelbaum MA: Results of a survey of neurosurgical practice patterns regarding the prophylactic

use of anti-epilepsy drugs in patients with brain tumors. J Neurooncol 2005, 74:211–215.PubMedCrossRef 28. Kim YJ, Xiao Y, Mackenzie CF, Gardner SD: Availability of trauma specialists in level I and II trauma centers: a national survey. J Trauma 2007, 63:676–683.PubMedCrossRef 29. Berry C, Sandberg DI, Hoh DJ, Krieger MD, McComb JG: Use of cranial fixation pins in pediatric neurosurgery. Neurosurgery 2008, 62:913–918. discussion 918–919PubMedCrossRef 30. Lebude B, Yadla S, Albert T, Anderson DG, Harrop JS, Hilibrand A, Maltenfort M, Sharan A, Vaccaro AR, Ratliff JK: Defining “”Complications”" in Spine Surgery: Neurosurgery and Orthopedic Spine Surgeons’ Survey. Geneticin chemical structure J Spinal Disord Tech 2010,23(8):493–500.PubMedCrossRef

31. Glotzbecker MP, Bono CM, Harris MB, Brick G, Heary RF, Wood KB: Surgeon practices regarding postoperative thromboembolic prophylaxis after high-risk spinal surgery. Spine (Phila Pa 1976) 2008, 33:2915–2921.CrossRef 32. American College of Surgeons Committee on Trauma: National Trauma Data Bank. Chicago, IL; 2010. 33. Hollingworth W, Nathens AB, Kanne JP, Crandall ML, Crummy TA, Hallam DK, Wang MC, Jarvik JG: The diagnostic accuracy PDK4 of computed tomography angiography for traumatic or atherosclerotic lesions of the carotid and vertebral arteries: a systematic review. Eur J Radiol 2003, 48:88–102.PubMedCrossRef 34. Hoit DA, Schirmer CM, Weller SJ, Lisbon A, Edlow JA, Malek AM: Angiographic detection of carotid and vertebral arterial injury in the high-energy blunt trauma patient. J Spinal Disord Tech 2008, 21:259–266.PubMedCrossRef 35. Biffl WL, Egglin T, Benedetto B, Gibbs F, Cioffi WG: Sixteen-slice computed tomographic angiography is a reliable noninvasive screening test for clinically significant blunt cerebrovascular injuries. J Trauma 2006, 60:745–751. discussion 751–742PubMedCrossRef 36. Bub LD, Hollingworth W, Jarvik JG, Hallam DK: Screening for blunt cerebrovascular injury: evaluating the accuracy of multidetector computed tomographic angiography. J Trauma 2005, 59:691–697.PubMed 37.

Within the group of closely related strains RtTA1, R leguminosar

Within the group of closely related strains RtTA1, R. leguminosarum bv. viciae 3841 (Rlv), R. etli CFN42 (Rhe),

RltWSM2304 and RltWSM1325 clusters of replicons carrying the most similar replication systems can be distinguished. They comprise pRleTA1d-pRL12-p42f-pRLG201-pR132501 and pRleTA1b-pRL11-p42e-pRLG202-pR132502, respectively. Therefore, detection of positive hybridization signals with probes derived from rep genes of RtTA1 chromid-like replicons (i.e. pRleTA1b or pRleTA1d) to any of the replicons of the sampled strains allowed regarding those as a chromid-like. Based on the similarity of replication-partition genes detected in our assays, we divided the replicons of the studied strains into three genome compartments: chromosome, 3-Methyladenine in vivo chromid-like and ‘other plasmids’ (i.e. those replicons which gave a hybridization signal with molecular probes originating from repA and repC genes of pRleTA1a or pRleTA1c, as well as those that gave no signal with any rep probes of RtTA1 replication genes). The compartment designated ‘other plasmids’ also comprised pSym. Such replicon division was taken into consideration in the subsequent analyses of distribution of other markers in the studied strains. SB-715992 order Variability of chromosomal and plasmid marker location In further studies, the extent of gene content diversity in the sampled nodule isolates was examined. We aimed to estimate whether, besides repA and repC displacement

events, we could demonstrate changes in the location

of the chromosomal and plasmid genes. The same experimental approach was used, i.e. a series of Southern hybridizations with different genes with a well-defined chromosomal or plasmid location in RtTA1 (Table 1) [36]. For assays of chromosomal marker variability, essential bacterial genes were chosen: rpoH2, dnaK, dnaC, rrn, lpxQ as well as genes that are not essential or with unspecified essentiality but chromosomal in RtTA1, i.e. bioA, stbB, exoR, pssL (Pss-I) and rfbADBC (Pss-V) (Table 1). In addition, location of fixGH genes was assayed, even though they click here are known to be plasmid located on the sequenced RltWSM2304, RltWSM1325 [33, 34], Rlv [6] and Rhe [5] genomes, but chromosomal in RtTA1 [36]. A majority of the studied genes (rpoH2, dnaK, dnaC, rrn, lpxQ, bioA, stbB, exoR and pssL) were located on the chromosome in all the sampled strains, showing considerable conservation of chromosomal markers (Figure 3). Exceptionally, the Pss-V region was identified on the chromosome of the K3.6, K5.4 and RtTA1 but it was missing in the other strains (Figure 3) Moreover, fixGH symbiosis-related genes, which were chromosomal in the RtTA1, K3.6, K4.15 and K5.4 strains, were located mainly in the genome compartment designated as ‘other plasmids’ (pSym to be exact) in the remaining strains. The variable location of fixGH genes which were found on the chromosome, pSyms and chromid-like replicons (K12.