Efforts to identify EPS-I mutants that produce a short Vsa protei

Efforts to identify EPS-I mutants that produce a short Vsa protein have been unsuccessful. Thus, it cannot be ascertained whether EPS-I is required for efficient adherence when Vsa is short. No mutants that lack Vsa protein have been identified in our robust transposon library, suggesting that these proteins are essential (Dybvig et al., 2010). Past studies have concluded that M. pulmonis cells producing a short Vsa are sensitive to lysis by complement, leading to the hypothesis that the Vsa proteins form a protective

shield (Simmons et al., 2004). Cultures inoculated with mycoplasmas that produce a short Vsa protein have a longer lag phase than cultures inoculated with cells producing a long Vsa but have a rapid growth rate in exponential phase and reach a high titer (Dybvig et al., 1989), suggesting an initial toxicity that is labile and from which the long Vsa can protect. Ferroptosis inhibitor cancer EPS-I may also have a role in protection, rendering EPS-I mutants with a short Vsa protein difficult to isolate. Because it was intriguing that EPS-I promoted cytoadherence, but inhibited biofilm formation, hemadsorption assays were utilized as an additional approach to examine interactions among the mycoplasma and host cells. Hemadsorption has often been used as an indicator for adherence to pulmonary epithelia in multiple species of mycoplasma (Hasselbring et al., 2005). The utter lack of hemadsorption that was observed for mycoplasmas

that produce the VsaG isotype was Methane monooxygenase totally unexpected and perplexing. In all prior studies, variation Caspase inhibitor in Vsa length but not isotype

resulted in phenotypic differences. For example, the Vsa isotype has no known association with any tissue tropism (Gumulak-Smith et al., 2001; Denison et al., 2005). The EPS-I mutants are currently available only in the VsaG background, thus nothing can be said about the role of EPS-I in hemadsorption. However, the remaining Vsa isotypes A, I, and H all exhibit hemadsorption profiles in concurrence with previously published data, with short Vsa-producing strains exhibiting significantly greater hemadsorption than long Vsa-producing strains (Simmons & Dybvig, 2003). Bacterial pathogens generally produce multiple adhesins, and colonization is a complex process. The adhesins and receptors involved in the colonization of the murine host by M. pulmonis are unknown as are the precise roles of the Vsa proteins and the EPS-I polysaccharide. Cells producing a long Vsa protein or lacking EPS-I may retain the ability to colonize animals because although cytoadherence is reduced, it is not eliminated. Mycoplasmal structures resembling the towers of biofilms that develop on glass or plastic surfaces have been observed ex vivo and in vivo on the mouse trachea (Simmons & Dybvig, 2009). Thus, although mutants lacking EPS-I cytoadhere poorly, their enhanced ability to form a biofilm may be a contributing factor to their ability to efficiently colonize animals.

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Secondly, random amplification of polymorphic DNA (RAPD) amplific

Secondly, random amplification of polymorphic DNA (RAPD) amplifications or SmaI digestion Metformin molecular weight allowed us to differentiate (1) A. flavus, A. oryzae and A. minisclerotigenes; (2) A. parasiticus, A. sojae and A. arachidicola; (3) A. tamarii, A. bombycis and A. pseudotamarii. Among the 11 species, only A. parvisclerotigenus cannot be differentiated from A. flavus. Using the results of real-time PCR, RAPD and SmaI digestion, a decision-making tree was drawn up to identify nine of the 11 species of section Flavi. In contrast to

conventional morphological methods, which are often time-consuming, the molecular strategy proposed here is based mainly on real-time PCR, which is rapid and requires minimal handling. Aspergillus section Flavi includes six economically important species that are very closely related morphologically and phylogenetically, and which are often separated into two groups Cetuximab mouse on the basis of their impact on food or human health. The first group includes Aspergillus flavus, Aspergillus parasiticus and Aspergillus nomius, which can cause serious damage to stored food products such as wheat and rye grain, nuts, spices and peanuts (Kurtzman et al.,

1987; Moody & Tyler, 1990a; Samson et al., 2000; Rigo et al., 2002; Hedayati et al., 2007). Furthermore, these species can produce carcinogenic secondary metabolites, the aflatoxins (Klich & Mullaney, 1987; Kurtzman et al., 1987; Yuan et al., 1995; Samson et al., 2000; Hedayati et al., 2007). After Aspergillus fumigatus, A. flavus is known as the second cause of human invasive aspergillosis (Denning, 1998; Latgé, 1999; Hedayati et al., 2007). Often, the name A. flavus is mistakenly used to describe the different species of Aspergillus section Flavi. Other recently described species are included in this group but these species are less important economically or rarely isolated. Indeed,

Aspergillus bombycis was described by Peterson et al. (2001) from nine isolates collected in silkworm-rearing houses. A variety of A. flavus, A. flavus var. parvisclerotigenus, has been raised to species level by Frisvad et al. (2005) as Aspergillus parvisclerotigenus. Aspergillus arachidicola Erastin and Aspergillus minisclerotigenes were described by Pildain et al. (2008). Seven strains of A. arachidicola were isolated in Argentina from Arachis. Some of the 15 strains of A. minisclerotigenes were been described for a long time as A. flavus group II by Geiser et al. (1998a, b, 2000), before being raised to the species rank by Pildain et al. (2008). Hence, many authors have shown evidence that A. flavus sensu lato may consist of several species (Geiser et al., 1998a, b, 2000; Pildain et al, 2008). The second group of the section Flavi comprises the nonproducing aflatoxin species Aspergillus tamarii, Aspergillus oryzae and Aspergillus sojae. The last two have lost the ability to produce aflatoxins (Samson et al., 2000) and are widely used as a koji mold for the production of fermented foods in some Asian countries.

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4% when minority assays for K103N, Y181C and M184V were included

4% when minority assays for K103N, Y181C and M184V were included. This is a 45% (95% CI 15.2–83.7%; P=0.0020) increase in the detection of significant resistance-associated mutations using the more sensitive assays combined with standard genotyping, compared with standard genotyping alone. There was a temporal reduction Selleck AZD5363 of TDR detected by standard methods from 15.4% in 2003 to 9.5% in 2006. This follows similar trends previously observed in UK TDR surveillance [3,5]. Taken together, the minority species methods showed a significant increase in detection over standard genotyping alone, with the M184V assay accounting for almost all of this increase. The 13-fold increase

in detection of M184V was significant (P=0.0005), while the 20% increase in detection of K103N did not reach statistical significance (P=0.5), and the Y181C mutation was not detected in this population by either method. The increased level of M184V detected by the more sensitive assay corresponds

with the observation that this is amongst the most common drug resistance mutations seen in treatment-experienced patients [6]; nevertheless, it is rarely seen in TDR studies using standard genotyping by population sequencing. The high fitness cost of the M184V mutation means that it may rapidly revert to wild-type (M184) levels that are undetectable with standard genotypic methods in the absence of drug pressure. Estimates suggest that M184V will revert to wild type within 6.5 months following seroconversion [14]. By contrast, primary nonnucleoside reverse transcriptase inhibitor (NNRTI) Dactolisib clinical trial mutations such as K103N have a low fitness cost [15]. Estimates of K103N reversion in treatment-naïve patients suggest that its presence is stable

in the plasma RNA for >3 years following seroconversion [16]. The findings we report here support the suggestion that M184V Dichloromethane dehalogenase is as likely to be transmitted as other mutations. Minority M184V/I populations were found in patients achieving successful response to first-line ART combinations containing emtricitabine [8]; consequently, the clinical significance of minority M184V is at present uncertain. Our observation that the M184V mutation occurred in only a minority of recent infections with other drug resistance mutations was surprising. This may indicate that the diagnostic use of minority assays to study only specimens with other resistance, as determined by standard genotypic methods, is inappropriate. The patient specimens were analysed using serological incidence testing to determine whether they came from a recent or long-standing infection. There was no significant difference between these two categories in terms of TDR rates. The issue of examining chronically HIV-infected patients to estimate rates of TDR is controversial because of high fitness cost mutations probably reverting to wild type over an extended period of time [17].

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Interassay variability was 154–903% All the above biomarker as

Interassay variability was 1.54–9.03%. All the above biomarker assays were performed at the Laboratory for Clinical Biochemistry Research under the direction of Dr Russell Tracy, Department of Pathology, University of Vermont. F2-isoprostanes were measured in the Eicosanoid Core Laboratory at Vanderbilt University. Briefly, F2-isoprostanes find more were quantified using gas chromatography–mass spectrometry after

Sep-Pak (Waters Corporation, Milford, MA, USA) and thin layer chromatography purification as pentafluorobenzyl ester and trimethylsilyl ether derivatives utilizing stable isotope dilution techniques with [2H4]-15-F2t-IsoP (Cayman Chemical, Ann Arbor, MI, USA) as an internal standard. The precision of this assay is ±4%, the accuracy is ±95% and the interassay variability is <8%. Important demographic, HIV and cardiovascular factors are described for the group overall, by ATV status (currently Trametinib nmr taking ATV vs. not) and by total bilirubin level (≥75th percentile vs. <75th percentile). The median and interquartile range

(IQR) are reported for continuous variables and the frequency and percentage for categorical variables. All demographic, HIV and cardiovascular factors, as well as endpoints, were compared based on ATV status and total bilirubin level using unpaired t-tests or Wilcoxon rank sum tests as distributionally appropriate for continuous variables, and χ2 tests, Fisher’s exact tests or Pearson exact χ2 tests as appropriate for categorical variables. Spearman correlation coefficients were determined between total bilirubin as a continuous variable and endpoints.

All for above statistical tests were two-sided and considered significant with P < 0.05. No corrections for multiple comparisons were made in this exploratory study. Next, in order to explore the relationship between FMD and total bilirubin in this sample, univariable followed by multivariable linear regressions were performed. In the univariable analysis, all demographic, HIV and cardiovascular factors, and inflammation, coagulation and oxidative stress markers as well as ATV status and total bilirubin as a dichotomized variable by ≥75th percentile compared with <75th percentile and a continuous variable were modelled with FMD as the outcome. In the first multivariable modelling approach, those variables with P < 0.25 were included in three separate multivariable models with ATV status or total bilirubin, as a categorical or continuous variable, as the independent variable of interest. In addition, a second multivariable modelling approach including clinically relevant variables regardless of statistical association was undertaken.

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This issue was raised in focus group discussions of pharmacist pr

This issue was raised in focus group discussions of pharmacist prescribers in Scotland. The need for a workforce of prescribers prompted

research of a large sample of Great Britain pharmacists. Results of research conducted in 2006 highlighted that a minority had taken any prescribing training action, with the majority being at the pre-contemplation stage. However, most strongly agreed/agreed that prescribing would improve patient care, but strongly disagreed/disagreed that they had sufficient pharmacist/technical support. Predictors of prescribing training actions were: colleagues undertaken/undertaking training; awareness of local prescribing networks; postgraduate qualifications; receptivity to change; intrinsic (professional) factors; Cell Cycle inhibitor and extrinsic (infrastructure) factors. We have very recently repeated this research with very similar findings. Research Obeticholic Acid cost in Scotland has demonstrated a lack of strategic direction and policies to support pharmacist prescribing in secondary care hence there is still much to be done to optimise pharmacist

prescribing. In summary, pharmacist prescribing is dynamic and rapidly changing making this a very exciting area of research. Other areas under investigation include pharmacist prescriber pharmacovigilance activities, the transition from supplementary to independent prescribing status, focus on generating solutions to those unable to prescribe and prescribing Sitaxentan within the undergraduate curriculum. There are so many unanswered research questions and we must provide robust evidence on which to base sustainable services, essential in the current political and economic climate. I am fortunate to work with so many talented colleagues

at Robert Gordon University and beyond. My research achievements are the result of collaboration and team working, highly relevant to the conference theme. While time and space do not permit to name them all, I must highlight two key researchers in pharmacist prescribing, Dr Johnson George and Katie MacLure, without them and many others I would not have received this award. “
“Objectives  Diagnosis and management of osteoporosis in hospitals are poor. Effective medications for reducing fracture risk are often underutilised in hospital settings. Studies have shown that improvements in secondary prevention of osteoporosis can occur with the implementation of clinical pathways and are effective in improving the prescription for osteoporosis medications. We aimed to assess the long-term sustainability of the benefit of the osteoporosis pathway implemented at The Queen Elizabeth Hospital, Adelaide, Australia, in 2003. Methods  An audit was performed to review the rate of prescription for osteoporosis therapy 5 years after the implementation of a pharmacist-driven osteoporosis pathway in patients presented with a minimal trauma fracture and admitted to the Department of Orthopaedics at The Queen Elizabeth Hospital.

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In French study of ZDV + lamivudine a small proportion of infants

In French study of ZDV + lamivudine a small proportion of infants required

either blood transfusions or early stop of therapy. Transient lactic acidaemia has been observed in HIV-uninfected infants exposed to HAART in utero and/or ZDV neonatally [79] Combo (all with ZDV) Combo (+ nelfinavir) Mandelbrot 2001 [23] Moodley 2003 [20] Durand-Gasselin 2008 [80] Hirt 2011 [11] Mirochnick 2011 [25] Mothers received two tablets of TDF/FTC at onset of labour and then one tablet daily for 7 days postpartum. This dose resulted in high FTC levels in neonates. Can cause neutropenia, anaemia 13 mg/kg as a single dose within 12 h of life. On the first day of life, neonates received a single dose of NVP syrup (2 mg/kg), within the 12 h after birth a single dose of TDF oral solution (13 mg/kg) and a single dose of FTC oral solution (2 mg/kg), buy BYL719 and for 7 days ZDV syrup (4 mg/kg every 12 h). Single dose administered to neonate after the mothers had received two tablets of TDF/FTC at delivery. Associated with renal dysfunction: monitor

renal function in neonates. Daily dosing regimen: 2 mg/kg once a day for 1st week then 4 mg/kg once a day for 2nd week then stop. Use 4 mg/kg once a day for 2 weeks if mother has received more than 3 days NVP. Single-dose regimen: one 2 mg/kg dose 48–72 h from birth Mono Mono NICHD/HPTN 040/P1043 Mirochnick 2011 [25] 300 mg/m2 Vemurafenib datasheet twice daily 1–2 kg: 40 mg every 12 h 2–6 kg: 80 mg every 12 h Jullien 2006 [83] Verweel 2007 [84] Chadwick 2008 [32] Chadwick 2011 [33] Urien 2011 [35] Some pharmacokinetic studies have suggested that a twice-daily

dose may give low levels in neonates. Frequent dose adjustment for weight gain is advisable. Adrenal dysfunction reported in newborns. Monitor electrolytes. Avoid in premature babies [36]. FDA recommendation (August 2011): the use of Kaletra oral solution should be avoided in premature babies until 14 days after their due date, or in full-term babies <14 days of age unless a healthcare professional believes that the benefit of using Chlormezanone Kaletra oral solution to treat HIV infection immediately after birth outweighs the potential risks. In such cases, FDA strongly recommends monitoring for increases in serum osmolality, serum creatinine and other signs of toxicity. 900 mg/m2 once daily Mon/Wed/Fri <6 months: 120 mg once daily Mon/Wed/Fri 6–12 months: 240 mg once daily Mon/Wed/Fri 8.1.1 Zidovudine monotherapy is recommended if maternal VL is <50 HIV RNA copies/mL at 36 weeks’ gestation or thereafter before delivery (or mother delivered by PLCS while on zidovudine monotherapy). Grading: 1C For women with fully suppressed HIV and a history of zidovudine resistance see discussion below. Zidovudine monotherapy for the infant has been part of the PMTCT strategy since publication of the ACTG 076 results [4].

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Hence, HAART incorporating agents active against HBV (tenofovir a

Hence, HAART incorporating agents active against HBV (tenofovir and emtricitabine) should be continued in this group. In those women with CD4 cell counts of >500 cells/μL with a baseline HBV DNA >2000 IU/mL and/or evidence of fibrosis on biopsy or FK228 Fibroscan, HBV treatment should be continued because of the risk of progressive liver disease if discontinued. In these patients, HAART incorporating tenofovir and emtricitabine

should be continued. Adefovir is an option and has been evaluated against HBV in coinfected patients. It does not select resistance against tenofovir but is less active than tenofovir. Neither entecavir (has antiviral activity to HIV and selects resistance) nor telbivudine (high resistance rates) are suitable Ruxolitinib molecular weight in coinfection. In those with CD4 cell counts over 500 cells/μL who received HAART to prevent MTCT and who are not HBV viraemic (>2000 IU/mL) or have evidence of established liver disease, strong consideration should be given to continuing anti-HBV therapy, in the form of tenofovir-based HAART because of the risk of progression of liver disease in coinfection. Inflammatory flares, which may be severe, particularly in persons with cirrhosis can occur because of viral escape and HBV viraemia, if anti-HBV drugs are stopped. In an RCT comparing lamivudine with placebo for reducing HBV MTCT

in patients with HBV mono-infection, an immediate increase in HBV DNA levels was observed on discontinuation of lamivudine postpartum [15]. Similarly, hepatitis flares among HIV/HBV coinfected patients have been reported upon the discontinuation of lamivudine,

emtricitabine and tenofovir. In the Swiss HIV observational cohort, liver enzyme elevation occurred in 29% of patients who discontinued lamivudine and in 5% this was severe, with three patients presenting with fulminant hepatitis [16] at a median time of 6 weeks after discontinuation. Hepatitis flares that occurred after ART cessation should be treated by resumption of active anti-HBV treatment before significant liver failure occurs. 6.1.17 Mannose-binding protein-associated serine protease In the absence of obstetric complications, normal vaginal delivery can be recommended if the mother has fully suppressed HIV VL on HAART. Grading: 2C No data exist to support any benefit from PLCS in mothers with HBV/HIV coinfection and no robust RCT exists in HBV mono-infected women. In a meta-analysis of mono-infected HBV women (four randomized trials all from China involving 789 people were included) where routine HBV neonatal vaccine and HBIG were used, there was strong evidence that PLCS vs. vaginal delivery could effectively reduce the rate of MTCT of HBV (RR 0.41; 95% CI 0.28–0.60) [17]. However, methodological concerns, including lack of information on randomization procedure, lack of allocation concealment and lack of blinding make the role of PLCS for PMTCT of HBV uncertain.

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These mice were generated using a mixed C57BL/6J and DBA strain a

These mice were generated using a mixed C57BL/6J and DBA strain as background and the coding region of the ghsr locus was precisely deleted and replaced with an in-frame lacZ reporter gene (Abizaid et al., 2006; Diano et al., 2006). click here All animals had free access to tap water at all times and to food unless otherwise specified. Prior to the beginning of the experiments, animals were group-housed under an LD cycle with the onset of light set at 08:00 h [zeitgeber time (ZT) 0], with light intensity ranging between 120 and 180 lux at cage level. Research was conducted according to the guidelines of the Canadian Council on Animal Care and approved by Carleton

University’s Animal Care Committee. GHSR-KO mice (n = 2) living on an LD schedule were taken from their home cage at ≈ ZT 4–6, overdosed with sodium pentobarbital and perfused using a 2% paraformaldehyde solution. The Galunisertib price brains were postfixed overnight in 2% paraformaldehyde, sliced into 50-μm sections on a Vibratome, and stained using the beta-galactocidase staining method described previously (Diano et al., 2006). Briefly, sections were thoroughly rinsed with 10 mm phosphate-buffered saline (PBS; in mm: NaCl, 137; KCl, 2.7; Na2HPO4, 8; KH2PO4, 2.6), rinsed once quickly in cold PBS plus 2 mm MgCl2 (PBS-MgCl2), then incubated in

PBS-MgCl2 for 10 min at 4 °C. Permeability was then increased by incubating in cold PBS with detergent (0.01% sodium desoxycholate and 0.02% NP40) for 10 min at 4 °C, and placed in

Metformin concentration staining solution for 4 h at 37 °C in the staining solution containing (in mm) K3Fe(CN)6, 25; K4Fe(CN)6, 25; MgCl2, 2 in PBS with 1 mg/mL of X-Gal. For the LD condition, WT and GHSR-KO mice (n = 4 per group per time point) were taken from their home cage at ZT 0, ZT 6, ZT 12 or ZT 18. Pairs of animals consisting of one WT and one KO were injected with an overdose of sodium pentobarbital and perfused with 100 mL of saline (0.9%) followed by 100 mL of 4% paraformaldehyde. Brains were postfixed in 4% paraformaldehyde overnight and transferred to a 1% sodium azide solution until being sectioned at a thickness of 60 μm using a Vibratome. Sections were then cryoprotected in Watson’s solution and frozen. One out of four 60-μm sections containing the hypothalamus were processed for cFos immunocytochemistry as described previously (Abizaid et al., 2005). Separate sections through the SCN were processed for PERIOD1 (PER1) or PERIOD2 (PER2) as described previously (Amir et al., 2004). Images from different hypothalamic nuclei were captured with a digital camera connected to an Olympus microscope (Olympus Canada, Markham, ON, Canada), and analysed using Image XSM software (v. 1.91, 2010,http://www.liv.ac.uk/~sdb/ImageSXM/).

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1a) To reduce the number of sequences from the Rhodobacter genus

1a). To reduce the number of sequences from the Rhodobacter genus, we only included one example of each group of sequences with a similarity value of 95% or more (see Table S1). When compared to a 16S-based

phylogeny (Fig. 1b), the RpoN-based tree shows no major changes in the branch distribution, suggesting an ancient origin of rpoN, at least within proteobacteria. Despite the low similarity level between the different copies of rpoN within the Rhodobacter group, all cluster together forming subgroups that correspond with their known or probable function and with their genomic context. This result supports the idea that the rpoN copies present in these strains are the result of several duplication events. Although a low number of sequences are Adriamycin cost available, we tried to deduce the order in which the rpoN copies appeared. To do this, we looked at their distribution in the 16S rRNA gene-based tree (Fig. 1b). The presence of rpoN1 in all the strains see more suggests that this may be the ancestral rpoN gene. If only duplication and deletion events are invoked, rpoN3 would be the first duplicated copy to appear,

because it is present in the early branching Rhodobacter sp. SW2. The R. capsulatus/blasticus group would have lost the rpoN3 gene after its separation from the R. sphaeroides clade and two new duplication events, first within the R. sphaeroides group and finally in the R. sphaeroides 2.4.1/17029 group, led to the appearance of rpoN2 and rpoN4, respectively. Alternatively, all the duplications may have occurred within the R. sphaeroides SPTLC1 clade followed by HGT of rpoN3 to Rhodobacter sp. However, the distribution of the branches within the rpoN3 clade (Fig. 1a) resembles the 16S-based tree, indicating a linear inheritance of this gene. An interesting case is the phylogeny of RpoN1, where the R. capsulatus/blasticus group branches off from the rest of the species. This may be indicative of a different

selective pressure on the rpoN genes of these species, where a single copy of this gene is present. Our results allowed us to establish the genetic context of the rpoN genes sequenced in this work and to compare it with the genetic context of the rpoNs from fully sequenced genomes. As shown in Fig. 2, the rpoN gene from Rv. sulfidophilum is located downstream of the fixCX genes and upstream of a gene similar to hcpH/hpaI (potentially encoding a 2,4-dihydroxyhept-2-ene-1,7-dioic acid aldolase), whereas in R. blasticus, the rpoN gene is flanked by fixCX and nifA, suggesting that the rpoN genes present in these bacteria are involved in nitrogen fixation. The genomic context of the rpoN1 and 2 genes identified in R. azotoformans is identical to that observed in R. sphaeroides 2.4.1., WS8, KD131, ATCC17029, and ATCC17025. In these bacteria, rpoN1 is flanked by nifW and a gene encoding a conserved hypothetical protein (DUF1810).

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1a) To reduce the number of sequences from the Rhodobacter genus

1a). To reduce the number of sequences from the Rhodobacter genus, we only included one example of each group of sequences with a similarity value of 95% or more (see Table S1). When compared to a 16S-based

phylogeny (Fig. 1b), the RpoN-based tree shows no major changes in the branch distribution, suggesting an ancient origin of rpoN, at least within proteobacteria. Despite the low similarity level between the different copies of rpoN within the Rhodobacter group, all cluster together forming subgroups that correspond with their known or probable function and with their genomic context. This result supports the idea that the rpoN copies present in these strains are the result of several duplication events. Although a low number of sequences are FDA-approved Drug Library available, we tried to deduce the order in which the rpoN copies appeared. To do this, we looked at their distribution in the 16S rRNA gene-based tree (Fig. 1b). The presence of rpoN1 in all the strains Buparlisib manufacturer suggests that this may be the ancestral rpoN gene. If only duplication and deletion events are invoked, rpoN3 would be the first duplicated copy to appear,

because it is present in the early branching Rhodobacter sp. SW2. The R. capsulatus/blasticus group would have lost the rpoN3 gene after its separation from the R. sphaeroides clade and two new duplication events, first within the R. sphaeroides group and finally in the R. sphaeroides 2.4.1/17029 group, led to the appearance of rpoN2 and rpoN4, respectively. Alternatively, all the duplications may have occurred within the R. sphaeroides cAMP clade followed by HGT of rpoN3 to Rhodobacter sp. However, the distribution of the branches within the rpoN3 clade (Fig. 1a) resembles the 16S-based tree, indicating a linear inheritance of this gene. An interesting case is the phylogeny of RpoN1, where the R. capsulatus/blasticus group branches off from the rest of the species. This may be indicative of a different

selective pressure on the rpoN genes of these species, where a single copy of this gene is present. Our results allowed us to establish the genetic context of the rpoN genes sequenced in this work and to compare it with the genetic context of the rpoNs from fully sequenced genomes. As shown in Fig. 2, the rpoN gene from Rv. sulfidophilum is located downstream of the fixCX genes and upstream of a gene similar to hcpH/hpaI (potentially encoding a 2,4-dihydroxyhept-2-ene-1,7-dioic acid aldolase), whereas in R. blasticus, the rpoN gene is flanked by fixCX and nifA, suggesting that the rpoN genes present in these bacteria are involved in nitrogen fixation. The genomic context of the rpoN1 and 2 genes identified in R. azotoformans is identical to that observed in R. sphaeroides 2.4.1., WS8, KD131, ATCC17029, and ATCC17025. In these bacteria, rpoN1 is flanked by nifW and a gene encoding a conserved hypothetical protein (DUF1810).

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