Halomonas fontilapidosi sp. nov., a moderately halophilic, denitrifying bacterium

We have made a polyphasic taxonomic study of strain 5CR(T), isolated from Fuente de Piedra, Málaga, southern Spain. The strain is a moderately halophilic, Gram-negative rod, oxidase-positive and motile by a single polar flagellum. It does not produce acids from sugars and shows respiratory metabolism, using oxygen, nitrate and nitrite as terminal electron acceptors. It requires NaCl and grows best with 5-7.5 % w/v at temperatures of between 32 and 45 degrees C within a pH range of 6-8. Its 16S rRNA gene sequence indicates that strain 5CR(T) belongs to the genus Halomonas in the class Gammaproteobacteria. Its closest relatives are Halomonas alimentaria, H. nitroreducens, H. shengliensis and H. ventosae, with the type strains of which our strain showed 16S rRNA gene sequence similarity values of 96.7-97.8 %. DNA-DNA hybridization studies between strain 5CR(T) and H. ventosae CECT 5797(T), the phylogenetically nearest type strain, showed 40 % relatedness. Its G+C content is 65.7 mol%. Its major fatty acids are C(18 : 1)omega7c (31.36 %), C(16 : 0) (25.55 %), C(16 : 1)omega7c/iso-C(15 : 0) 2-OH (23.23 %), C(19 : 0) cyclo omega8c (8.14 %), C(12 : 0) 3-OH (5.76 %) and C(10 : 0) (2.22 %) and the predominant respiratory lipoquinone is ubiquinone with nine isoprene units (Q-9). The proposed name for the novel species is Halomonas fontilapidosi sp. nov., strain 5CR(T) (=CECT 7341(T) =LMG 24455(T)) being the type strain.Dirección General de Investigación Científica y Técnica (BOS2003-0498;CGL2005-05947) and from Plan Andaluz de Investigación, Spain

Halomonas nitroreducens sp. nov., a novel nitrate- and nitrite-reducing species.

We have carried out a polyphasic taxonomic study of strain 11ST, a halophilic, Gram-negative bacterium that is able to respire on nitrate and nitrite in anaerobiosis. Strain 11ST was isolated from a solar saltern in Cahuil, a region next to Pichilemu (Chile). It grows at NaCl concentrations within the range of 3-20 % w/v (optimum 5-7.5 %), temperatures from 4 to 45 degrees C (optimum 20-32 degrees C) and within a pH range of 5-10 (optimum pH 7-9). Its 16S rRNA gene sequence indicates that it belongs to the genus Halomonas in the class Gammaproteobacteria. Its closest relatives are Halomonas alimentaria, H. denitrificans, H. organivorans and H. ventosae, with the type strains of which our strain showed maximum 16S rRNA gene sequence similarity values of 97.1-98.1 %. Its G+C content is 65.3 mol%. DNA-DNA hybridization studies showed 54.2 % relatedness between strain 11ST and H. alimentaria DSM 15356T and 47.2 % relatedness between strain 11ST and H. organivorans CECT 5995T. Lower DNA-DNA hybridization percentages were obtained against the type strains of other related Halomonas species. Its major fatty acids are C12 : 0 3-OH (5.56 %), iso-C15 : 0 2-OH/C16 : 1 omega 7c (22.30 %), C16 : 0 (27.80 %) and C18 : 1 omega 7c (29.92 %). The proposed name for the novel species is Halomonas nitroreducens sp. nov., with strain 11ST (=CECT 7281T =LMG 24185T) being the type strain.Spanish Ministerio de Educación y Cultura (CGL 2005-05947) and Plan Andaluz de Investigació

Denitrification as an important taxonomic marker within the genus Halomonas.

We have made a comprehensive study of the denitrifying species of the genus Halomonas, evaluating both the phylogenetic and phenotypic relationships amongst them and other species of Halomonas. The phylogenetic analysis was based on the 16S rRNA gene sequence as well as those of the three genes essential to a complete denitrification process: narH, nirS and nosZ. The main aim of the phenotypic study was to improve our knowledge of some of the species in question. To this end we investigated the type strain of each species, although in the case of Halomonas cerina we also studied strains R53 and 15CR. In addition to this we investigated some other strains phylogenetically related to Halomonas ventosae, Halomonas denitrificans and Halomonas koreensis that were isolated during this study. We also looked into the conditions under which all these bacteria denitrify. Our results indicate that these denitrifying species of Halomonas are all closely related. A numerical analysis of the phenotypic data demonstrates a high phenotypic similarity (73%) between most of them. In addition, all the denitrifying strains have a high G+C content of between 63 and 74.3 mol%. The results of the phylogenetic study point to two evolutionary lineages for the process. Although phenotypic similarity does not always reflect phylogenetic relatedness, we have found significant congruence between both features in Halomonas, making it clear that denitrifying ability should be considered as an important phenotypic and phylogenetic discriminatory marker within this genus.This research was supported by grants from the Ministry of Technology and Science (CGL2008-02399) and from the Plan Andaluz de Investigación (P06-CVI-01850), Spain

Halomonas cerina sp. nov., a moderately halophilic, denitrifying, exopolysaccharide-producing bacterium

Three bacterial strains were isolated from different saline soils in Spain. The novel strains were moderately halophilic, exopolysaccharide-producing, Gram-negative, non-motile rods. The strains required NaCl and grew best with 7.5-10 % (w/v) NaCl in the medium. They formed wax-coloured colonies, were oxidase-positive and showed respiratory metabolism, using oxygen, nitrate and nitrite as terminal electron acceptors. The novel strains were able to denitrify and did not produce acid from sugars. The DNA G+C contents varied between 62.7 and 66.2 mol%. Phylogenetic analyses based on 16S rRNA gene sequences and sequence signatures of this gene showed that all three novel isolates belonged to the genus Halomonas in the class Gammaproteobacteria and formed an independent phylogenetic line. The most phylogenetically related species were Halomonas alimentaria, Halomonas campaniensis, Halomonas gudaonensis and Halomonas ventosae, with which the novel strains showed 16S rRNA gene sequence similarity values of between 96.3 and 95.2 %. The principal fatty acids of the novel strains were 16 : 0, 18 : 1 omega 7c, 16 : 1 omega 7c and 19 : 0 cyclo omega 8c. The predominant respiratory lipoquinone was ubiquinone with nine isoprene units (Q-9). The name Halomonas cerina sp. nov. is proposed for these isolates. The type strain is SP4T (=CECT 7282T=LMG 24145T).Dirección General de Investigación Científica y Técnica (BOS2003-0498; CGL2005-05947) y Plan Andaluz de Investigación

Diversity of culturable halophilic archaea isolated from Rambla Salada, Murcia (Spain).

We have studied the diversity of culturable halophilic Archaea at Rambla Salada, Murcia (south-eastern Spain). We made 8 samplings at different places in this habitat during the years 2006 and 2007 and isolated a total of 49 strains, which were identified by means of phenotypic tests and the hypervariable V1-V3 region of the 16S rRNA gene sequences (around 500 bp). The ribosomal data showed that the isolates belonged to 12 genera within the Halobacteriaceae family, with Haloferax and Natrinema being the most abundant. Five strains showed less than 97% sequence identity with validly described species and may well represent new taxa. All the strains grew best with around 25% w/v salts, required high concentrations of NaCl and magnesium and produced red to pink colonies. They were facultative anaerobes with both respiratory and fermentative metabolisms. The diversity of the archaeal community was analysed with the MOTHUR package. We identified 14 OTUs at the 3% genetic distance level and found quite high diversity. Rarefaction curves of richness estimators and diversity indices demonstrated that our collection of isolates represented the archaeal community at Rambla Salada that can be isolated under the conditions used in this work. This is the first report to be published on the culturable archaea at Rambla Salada, an area of considerable ecological interest

Synergic effect of metabolic syndrome and lipodystrophy on oxidative stress and inflammation process in treated HIV-patients.

The aim of this study was to assess the effect of lipodystrophy (LD) associated to metabolic syndrome (MS) on oxidative stress and inflammation in a cohort of 243 HIV-infected patients with MS, all of them under three different antiretroviral regimens. We collected immunovirological, biochemical and metabolic data, as well as anthropometric measurements. In addition, cardiovascular risk was also assessed by means of Atherogenic Index of Plasma (API) and Framingham Risk Score. The MS-LD patient set was characterized by a lower initial lymphocyte CD4 count and CD4/CD8 ratio and a higher initial viral load than the group without LD. We also found worse lipidic and glycaemic profiles (with lower HDL-cholesterol and higher triglyceride and glucose levels) in the MS-LD group. BMI, systolic blood pressure and Framingham score were significantly increased compared to MS-Non LD. In addition, patients with MS and LD had significantly higher levels of carbonylated proteins, lipid peroxidation, IL-6 and IL-8, as well as a significant decrease in the levels of leptin, adiponectin and antioxidant activities of catalase, super oxide dismutase and glutathione associated enzymes. In MS-LD HIV-1 patients, a significant negative correlation was found between Framingham Risk Score and the antioxidant biomarkers, however a positive association was found between API and protein-C reactive and carbonylated proteins. Segregating by ART, the above-mentioned conditions were worse within the MS-LD group whose treatment contained protease inhibitors, such as lopinavir. In conclusion, HIV-1 infected patients treated for at least six months, especially with regimens including PIs, showed a worsening of inflammatory process and oxidative stress.This work was supported by the Spanish Ministry of Science and Innovation [SAF 2010-17213]; the Regional Ministry of Health from the Andalusian Government [SAS 111226]; “Miguel Servet Type II” grant [CPI13/00003 to M.I.Q.O. and CPII18/00030 to L.G.S.] from ISCIII, co-funded by the European Regional Development Fund (ERDF); “Nicolas Monardes” research program from the Regional Ministry of Health from the Andalusian Government [C-0030-2018 to M.I.Q.O. and C-0028-2018 to L.G.S.

Pharmacological aspects of darunavir/cobicistat

Rezolsta® (darunavir/cobicistat) es el primer inhibidor de la proteasa potenciado en combinación fija con un nuevo potenciador farmacológico. Darunavir es actualmente el inhibidor de la proteasa de uso preferente con un perfil de eficacia y seguridad bien conocido. Cobicistat es un inhibidor del citocromo P450 3A sin actividad antirretroviral que se usa como potenciador farmacológico y que puede ser una alternativa al ritonavir. Los parámetros farmacológicos de darunavir con cobicistat son similares a los que se producen cuando el potenciador es ritonavir. Los ensayos clínicos de darunavir/cobicistat han demostrado gran eficacia y buena tolerabilidad. El cobicistat inhibe de forma más específica el citocromo P450 3A sin efecto inductor, por lo que presenta un menor perfil de interacciones farmacológicas que el ritonavir. El cobicistat está también coformulado a dosis fijas con otros fármacos: elvitegravir, tenofovir diproxil fumarato y emtricitabina (Stribild®), elvitegravir, tenofovir alafenamina y emtricitabina (Genvoya®) y con atazanavir (Evotaz®). En este capítulo se revisan los aspectos farmacológicos de la combinación darunavir/cobicistat (Rezolsta®

The DORIVIR Study

Objectives: To analyze the efficacy and safety of dolutegravir/rilpivirine (DTG/RPV) in HIV-infected patients who switched from any other antiretroviral therapy (ART). Methods: Open-label, multicenter study including patients who switched to DTG/RPV between February 2015 and February 2016. Efficacy (HIV RNA <50 copies/mL), adverse events, and metabolic changes at 24 weeks were analyzed. Results: A total of 104 participants were included, who switched for the following reasons: toxicity/intolerance (42.3%), convenience (27.8%), and drug interactions (17.3%). Prior regimens are protease inhibitor (56.7%), integrase strand transfer inhibitor (26.9%), and non-nucleoside reverse transcriptase inhibitor (16.3%). Efficacy at 24 weeks was 88.4% (intention to treat) and 96.8% (per protocol). Triglyceride levels were reduced, on average, by 12.7% and a mean decrease of 9.0% in the glomerular filtration rate was observed as well ( P values of .003 and .002, respectively), whereas total cholesterol, HDL cholesterol, LDL cholesterol, creatinine, and glutamic-pyruvic transaminase remained unchanged. No patient discontinued due to adverse events. Conclusions: Dolutegravir/RPV is effective and safe in long-term HIV-infected patients under any prior ART. Toxicity, convenience, and interactions were the main reasons for changing. At 24 weeks, the lipid profile improved with a decrease in triglycerides.The author(s) disclosed receipt of the following financial support for the research, authorship, and/or publication of this article: Funding was received from the Andalusian Society of Infectious Diseases (SAEI) (provided study code SAEI 00/0067)

High Prevalence of Sequences Included in Transmission Clusters Within Newly Diagnosed HIV-1 Patients in Southern Spain (2004-2015)

The presence of transmission clusters (TCs) and their epidemiological characteristics in a treatment-naive cohort of HIV-1 patients in southern Spain over a decade (2004-2015) were evaluated. Protease and reverse transcriptase sequences provided by each genotype test were used in the phylogenetic study, performed first by the neighbor-joining method and then confirmed by Bayesian analysis. We collected clinical, immunovirological, and demographic data for all patients included. Our cohort comprised 757 patients, 428 (56.5%) belonging to a TC. Overall, we found 123 TCs, 21 of them comprising five or more individuals and three with ≥10 sequences. Forty-three TCs (35.0%) remained active. The clustered patients were mainly men (92.8%) who had sex with men (MSM) (81.5%), Spanish (80.6%), and young adults (median age at diagnosis of 32.6 years). They had lower percentages of late diagnosis and AIDS cases (42.1% and 13.6%, respectively), whereas the presence of recent seroconverters (31.1%), HIV-1 B subtypes (79.4%), and transmission drug resistance (20.3%) increased within TCs, with regard to not-clustered individuals. Among the TCs of non-B variants, circulating recombinant forms (CRF) were predominant (87.5%), with the highest frequencies for CRF19_cpx (17.0% of non-B subtype sequences in TCs); CRF02_AG (15.9%); and CRF01_AE (9.1%). In conclusion, over half of our cohort was included within a TC. More than a third of TCs found could be considered active transmission events. Belonging to a TC was related to MSM, Spanish origin, recent seroconversion, high prevalence of resistance mutations, and B HIV subtype. Among the non-B genetic forms in TCs, we found a high prevalence of CRF19_cpx, CRF02_AG, and CRF01_AE variants.This work was mainly supported by the National R+D+I Plan (RD16/0025/0032 project); the Institute of Health Carlos III (ISCIII); and the European Regional Development Fund

Clinical, virological and phylogenetic characterization of a multiresistant HIV-1 strain outbreak in naive patients in southern Spain

Background: We describe the characteristics of an HIV-1 strain with six viral reverse transcriptase mutations (D67N, T69N/D, V118I, V179D, T215S and K219Q), which we have called the Malaga strain. This strain was detected in treatment-naive patients from southern Spain. Methods: The study was undertaken at the Virgen de la Victoria Hospital, Malaga, a reference centre for the study of HIV-1 genotype resistance in Andalusia (the 'Costa del Sol'), Spain. Genotypic resistance testing was done in an automated sequencer. Phylogenetic analysis was performed using a 630 bp region of the reverse transcriptase with the mutations mentioned. Results: Between 2007 and 2014, we detected the Malaga strain in 30 treatment-naive patients. All were MSM, seen at five hospitals on the Costa del Sol. In all cases, the HIV-1 was subtype B with viral tropism R5. Phylogenetic analysis based on the reverse transcriptase sequence showed consistent grouping (with a bootstrap value of the common node of 100%) of the isolates that shared the mutation pattern mentioned. This strain has not been detected elsewhere or in previously treated patients. All of the patients treated with first-line combination ART responded. Conclusions: We report a cluster of an HIV-1 strain with multiple resistance mutations that was transmitted over a period of >8 years, affecting 30 naive patients from the same geographical area. The strain was susceptible to first-line combination ART.Partially funded by the RD12/0017/0017 project (Plan Nacional R+D+I) and cofinanced by Instituto de Salud Carlos III-Subdirección General de Evaluación y el Fondo Europeo de Desarrollo Regional