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.

Factors associated with the number of drugs in darunavir/cobicistat regimens.

BACKGROUND: Darunavir/cobicistat can be used as mono, dual, triple or more than triple therapy. OBJECTIVES: To assess factors associated with the number of drugs in darunavir/cobicistat regimens. METHODS: A nationwide retrospective cohort study of consecutive HIV-infected patients initiating darunavir/cobicistat in Spain from July 2015 to May 2017. Baseline characteristics, efficacy and safety at 48 weeks were compared according to the number of drugs used. RESULTS: There were 761 patients (75% men, 98% were antiretroviral-experienced, 32% had prior AIDS, 84% had HIV RNA <50 copies/mL and 88% had ≥200 CD4 cells/mm3) who initiated darunavir/cobicistat as mono (n=308, 40%), dual (n=173, 23%), triple (n=253, 33%) or four-drug (n=27, 4%) therapy. Relative to monotherapy, triple therapy was more common in men aged <50 years, with prior AIDS and darunavir plus ritonavir use, and with CD4 cells <200/mm3 and with detectable viral load at initiation of darunavir/cobicistat; dual therapy was more common with previous intravenous drug use, detectable viral load at initiation of darunavir/cobicistat and no prior darunavir plus ritonavir; and four-drug therapy was more common with prior AIDS and detectable viral load at initiation of darunavir/cobicistat. Monotherapy and dual therapy showed a trend to better virological responses than triple therapy. CD4 responses and adverse effects did not differ among regimens. DISCUSSION: Darunavir/cobicistat use in Spain has been tailored according to clinical characteristics of HIV-infected patients. Monotherapy and dual therapy have been common and preferentially addressed to older patients with a better HIV status, suggesting that health issues other than HIV infection may have been strong determinants of its prescription.

Emergence as an outbreak of the HIV-1 CRF19_cpx variant in treatment-naïve patients in southern Spain.

BACKGROUND: CRF19_cpx is a complex circulating recombination form (CRF) of HIV-1. We describe the characteristics of an outbreak of the CRF19_cpx variant among treatment-naïve patients in southern Spain. METHODS: The study was undertaken at the Virgen de la Victoria Hospital, a reference centre for the analysis of HIV-1 genotype in Malaga (Spain). Subtyping was performed through REGA v3.0 and the relationship of our CRF19_cpx sequences, among themselves and regarding other reference sequences from the same variant, was defined by phylogenetic analysis. We used PhyML program to perform a reconstruction of the phylogeny by Maximum Likelihood method as well as further confirmation of the transmission clusters by Bayesian inference. Additionally, we collected demographic, clinical and immunovirological data. RESULTS: Between 2011 and 2016, we detected 57 treatment-naïve patients with the CRF19_cpx variant. Of these, 55 conformed a very well-defined transmission cluster, phylogenetically close to CRF19_cpx sequences from the United Kingdom. The origin of this subtype in Malaga was dated between 2007 and 2010. Over 50% of the patients presented the non-nucleoside reverse transcriptase inhibitor G190A resistance mutation. This variant was mostly represented by young adult Spanish men who had sex with men. Almost half of them were recent seroconverters, though a similar percentage was diagnosed at a late state of HIV infection. Five cases of AIDS and one non-AIDS defined death occurred during follow-up. The majority of patients treated with first-line combination antiretroviral therapy (ART) responded. CONCLUSIONS: We report the largest HIV-1 CRF19_cpx cohort of treatment-naïve patients outside Cuba, almost all emerging as an outbreak in the South of Spain. Half the cases had the G190A resistance mutation. Unlike previous studies, the variant from Malaga seems less pathogenic, with few AIDS events and an excellent response to ART.

Identification of Xenologs and Their Characteristic Low Expression Levels in the Cyanobacterium Synechococcus elongatus.

Horizontal gene transfer (HGT) is a central process in prokaryotic evolution. Once a gene is introduced into a genome by HGT, its contribution to the fitness of the recipient cell depends in part on its expression level. Here we show that in Synechococcus elongatus PCC 7942, xenologs derived from non-cyanobacterial sources exhibited lower expression levels than native genes in the genome. In accord with our observation, xenolog codon adaptation indexes also displayed relatively low expression values. These results are in agreement with previous reports that suggested the relative neutrality of most xenologs. However, we also demonstrated that some of the xenologs detected participated in cellular functions, including iron starvation acclimation and nitrate reduction, which corroborate the role of HGT in bacterial adaptation. For example, the expression levels of some of the xenologs detected are known to increase under iron-limiting conditions. We interpreted the overall pattern as an indication that there is a selection pressure against high expression levels of xenologs. However, when a xenolog protein product confers a selective advantage, natural selection can further modulate its expression level to meet the requirements of the recipient cell. In addition, we show that ORFans did not exhibit significantly lower expression levels than native genes in the genome, which suggested an origin other than xenology.

Sífilis e infección por el virus de la inmunodeficiencia humana: una endemia en hombres que tienen sexo con hombres / Syphilis and human immunodeficiency virus infection: an endemic infection in men who have sex with men

OBJETIVO: Describir las características epidemiológicas, clínicas y analíticas de los hombres que tienen sexo con hombres (HSH) con infección por el VIH diagnosticados de sífilis en la Unidad de Gestión Clínica de Enfermedades Infecciosas del Hospital Virgen de la Victoria de Málaga durante el período 2004-2013. Pacientes y método: Estudio descriptivo de 196 episodios de sífilis en 167 HSH infectados por el VIH (2004-2013). Se recogieron datos epidemiológicos, clínicos y analíticos de todos los pacientes. La incidencia anual de sífilis en HSH con infección por el VIH corresponde al cociente entre el número de episodios de sífilis en HSH en un año dividido por el número de HSH en seguimiento en ese año. RESULTADOS: La incidencia anual osciló entre el 1,2% (2007) y el 7,8% (2012). Presentación asintomática en el 42,8% y diagnóstico coincidente de sífilis e infección por el VIH en el 28,5%. CONCLUSIONES: La incidencia anual de sífilis ha aumentado en los HSH con infección por el VIH. Un tercio de los diagnósticos de infección por el VIH coincidió con el de sífilis y casi la mitad eran cuadros asintomáticos

OBJECTIVE: to analyse epidemiological, clinical, and analytical features of HIV-infected men who have sex with men (MSM) diagnosed with syphilis in the Infectious Diseases Unit (Hospital Virgen de la Victoria, Málaga, Spain) during 2004-2013. PATIENTS AND METHODS: An observational study was conducted on 196 syphilis episodes in 167 MSM infected with HIV (2004-2013). Epidemiological, clinical, and analytical data were collected. Annual syphilis incidence among HIV-MSM is calculated as the number of syphilis episodes among MSM in one year divided by the number of MSM followed up in that year. RESULTS: Incidence ranged from 1.2% (2007) to 7.8% (2012). There were asymptomatic episodes in 42.8% cases, and an HIV-syphilis coincident diagnosis in 28.5%. CONCLUSIONS: The annual incidence of syphilis has increased within HIV infected MSM. One third of the syphilis episodes were simultaneous to HIV diagnosis and near half of them were asymptomatic

[Syphilis and human immunodeficiency virus infection: an endemic infection in men who have sex with men]. / Sífilis e infección por el virus de la inmunodeficiencia humana: una endemia en hombres que tienen sexo con hombres.

OBJECTIVE: to analyse epidemiological, clinical, and analytical features of HIV-infected men who have sex with men (MSM) diagnosed with syphilis in the Infectious Diseases Unit (Hospital Virgen de la Victoria, Málaga, Spain) during 2004-2013. PATIENTS AND METHODS: An observational study was conducted on 196 syphilis episodes in 167 MSM infected with HIV (2004-2013). Epidemiological, clinical, and analytical data were collected. Annual syphilis incidence among HIV-MSM is calculated as the number of syphilis episodes among MSM in one year divided by the number of MSM followed up in that year. RESULTS: Incidence ranged from 1.2% (2007) to 7.8% (2012). There were asymptomatic episodes in 42.8% cases, and an HIV-syphilis coincident diagnosis in 28.5%. CONCLUSIONS: The annual incidence of syphilis has increased within HIV infected MSM. One third of the syphilis episodes were simultaneous to HIV diagnosis and near half of them were asymptomatic.

Lipid changes in HIV-patients switching to the coformulated single tablet FTC/RPV/TDF (Eviplera®). Efficacy and safety analysis. GeSida Study 8114.

INTRODUCTION: Rilpivirine (RPV) has a better lipid profile than efavirenz (EFV) in naïve patients (1). Switching to RPV may be convenient for many patients, while maintaining a good immunovirological control (2). The aim of this study was to analyze lipid changes in HIV-patients at 24 weeks after switching to Eviplera® (emtricitabine/RPV/tenofovir disoproxil fumarate [FTC/RPV/TDF]). MATERIALS AND METHODS: Retrospective, multicentre study of a cohort of asymptomatic HIV-patients who switched from a regimen based on 2 nucleoside reverse transcriptase inhibitors (NRTI)+protease inhibitor (PI)/non nucleoside reverse transcriptase inhibitor (NNRTI) or ritonavir boosted PI monotherapy to Eviplera® during February-December, 2013; all had undetectable HIV viral load for ≥3 months prior to switching. Patients with previous failures on antiretroviral therapy (ART) including TDF and/or FTC/3TC, with genotype tests showing resistance to components of Eviplera®, or who had changed the third drug of the ART during the study period were excluded. Changes in lipid profile and cardiovascular risk (CVR), and efficacy and safety at 24 weeks were analyzed. RESULTS: Among 305 patients included in the study, 298 were analyzed (7 cases were excluded due to lack of data). Men 81.2%, mean age 44.5 years, 75.8% of HIV sexually transmitted. 233 (78.2%) patients switched from a regimen based on 2 NRTI+NNRTI (90.5% EFV/FTC/TDF). The most frequent reasons for switching were central nervous system (CNS) adverse events (31.0%), convenience (27.6%) and metabolic disorders (23.2%). At this time, 293 patients have reached 24 weeks: 281 (95.9%) have continued Eviplera®, 6 stopped it (3 adverse events, 2 virologic failures, 1 discontinuation) and 6 have been lost to follow up. Lipid profiles of 283 cases were available at 24 weeks and mean (mg/dL) baseline vs 24 weeks are: total cholesterol (193 vs 169; p=0.0001), HDL-c (49 vs 45; p=0.0001), LDL-c (114 vs 103; p=0.001), tryglycerides (158 vs 115; p=0.0001), total cholesterol to HDL-c ratio (4.2 vs 4.1; p=0.3). CVR decreased (8.7 vs 7.5%; p= 0.0001). CD4 counts were similar to baseline (653 vs 674 cells/µL; p=0.08), and 274 (96.8%) patients maintained viral suppression. CONCLUSIONS: At 24 weeks after switching to Eviplera®, lipid profile and CVR improved while maintaining a good immunovirological control. Most subjects switched to Eviplera® from a regimen based on NNRTI, mainly EFV/FTC/TDF. CNS adverse events, convenience and metabolic disorders were the most frequent reasons for switching.

Diversity and distribution of Halomonas in Rambla Salada, a hypersaline environment in the southeast of Spain.

We have studied the diversity and distribution of Halomonas populations in the hypersaline habitat Rambla Salada (Murcia, southeastern Spain) by using different molecular techniques. Denaturing gradient gel electrophoresis (DGGE) using specific primers for the 16S rRNA gene of Halomonas followed by a multivariate analysis of the results indicated that richness and evenness of the Halomonas populations were mainly influenced by the season. We found no significant differences between the types of samples studied, from either watery sediments or soil samples. The highest value of diversity was reached in June 2006, the season with the highest salinity. Furthermore, canonical correspondence analysis (CCA) demonstrated that both salinity and pH significantly affected the structure of the Halomonas community. Halomonas almeriensis and two denitrifiers, H. ilicicola and H. ventosae were the predominant species. CARD-FISH showed that the percentage of Halomonas cells with respect to the total number of microorganisms ranged from 4.4% to 5.7%. To study the functional role of denitrifying species, we designed new primer sets targeting denitrification nirS and nosZ genes. Using these primers, we analyzed sediments from the upwelling zone collected in June 2006, where we found the highest percentage of denitrifiers (74%). Halomonas ventosae was the predominant denitrifier in this site.

Blueprint for a minimal photoautotrophic cell: conserved and variable genes in Synechococcus elongatus PCC 7942.

BACKGROUND: Simpler biological systems should be easier to understand and to engineer towards pre-defined goals. One way to achieve biological simplicity is through genome minimization. Here we looked for genomic islands in the fresh water cyanobacteria Synechococcus elongatus PCC 7942 (genome size 2.7 Mb) that could be used as targets for deletion. We also looked for conserved genes that might be essential for cell survival. RESULTS: By using a combination of methods we identified 170 xenologs, 136 ORFans and 1401 core genes in the genome of S. elongatus PCC 7942. These represent 6.5%, 5.2% and 53.6% of the annotated genes respectively. We considered that genes in genomic islands could be found if they showed a combination of: a) unusual G+C content; b) unusual phylogenetic similarity; and/or c) a small number of the highly iterated palindrome 1 (HIP1) motif plus an unusual codon usage. The origin of the largest genomic island by horizontal gene transfer (HGT) could be corroborated by lack of coverage among metagenomic sequences from a fresh water microbialite. Evidence is also presented that xenologous genes tend to cluster in operons. Interestingly, most genes coding for proteins with a diguanylate cyclase domain are predicted to be xenologs, suggesting a role for horizontal gene transfer in the evolution of Synechococcus sensory systems. CONCLUSIONS: Our estimates of genomic islands in PCC 7942 are larger than those predicted by other published methods like SIGI-HMM. Our results set a guide to non-essential genes in S. elongatus PCC 7942 indicating a path towards the engineering of a model photoautotrophic bacterial cell.

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.

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.

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).

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.