Effective and alternative treatment of anastomotic leak after gastrojejunal bypass: endoluminal vacuum therapy.

Anastomotic leak (AL) after with Roux-en-Y gastric bypass (RYGB) has a morbidity rate to 53% and it can be potentially lethal (mortality rate from 0.5 to 10%). In these cases, surgery is usually a challenge, so in recent years minimally invasive endoscopic treatment is gaining ground. Endoluminal vacuum therapy (EVAC) is a promising treatment that is being used in esophagogastric and rectal surgery for the management of AL. We present the case of a patient on his 5th postoperative day of bariatric surgery (RYGB) with an acute abdomen. He was diagnosed of dehiscence of gastrojejunal anastomosis and he needed urgent surgery twice. Subsequently, in control CT, a new anastomotic leak is evidenced. However, given the clinical stability of the patient, it was decided to start EVAC type ESO-Sponge® placed by endoscopy. A total of 4 changes are made every 3-4 days with a total duration of treatment of 15 days. EVAC was removed when the defect presented a 1 mm sized.

Systems metabolic engineering of the primary and secondary metabolism of Streptomyces albidoflavus enhances production of the reverse antibiotic nybomycin against multi-resistant Staphylococcus aureus.

Nybomycin is an antibiotic compound with proven activity against multi-resistant Staphylococcus aureus, making it an interesting candidate for combating these globally threatening pathogens. For exploring its potential, sufficient amounts of nybomycin and its derivatives must be synthetized to fully study its effectiveness, safety profile, and clinical applications. As native isolates only accumulate low amounts of the compound, superior producers are needed. The heterologous cell factory S. albidoflavus 4N24, previously derived from the cluster-free chassis S. albidoflavus Del14, produced 860 µg L-1 of nybomycin, mainly in the stationary phase. A first round of strain development modulated expression of genes involved in supply of nybomycin precursors under control of the common Perm* promoter in 4N24, but without any effect. Subsequent studies with mCherry reporter strains revealed that Perm* failed to drive expression during the product synthesis phase but that use of two synthetic promoters (PkasOP* and P41) enabled strong constitutive expression during the entire process. Using PkasOP*, several rounds of metabolic engineering successively streamlined expression of genes involved in the pentose phosphate pathway, the shikimic acid pathway, supply of CoA esters, and nybomycin biosynthesis and export, which more than doubled the nybomycin titer to 1.7 mg L-1 in the sixth-generation strain NYB-6B. In addition, we identified the minimal set of nyb genes needed to synthetize the molecule using single-gene-deletion strains. Subsequently, deletion of the regulator nybW enabled nybomycin production to begin during the growth phase, further boosting the titer and productivity. Based on RNA sequencing along the created strain genealogy, we discovered that the nyb gene cluster was unfavorably downregulated in all advanced producers. This inspired removal of a part and the entire set of the four regulatory genes at the 3'-end nyb of the cluster. The corresponding mutants NYB-8 and NYB-9 exhibited marked further improvement in production, and the deregulated cluster was combined with all beneficial targets from primary metabolism. The best strain, S. albidoflavus NYB-11, accumulated up to 12 mg L-1 nybomycin, fifteenfold more than the basic strain. The absence of native gene clusters in the host and use of a lean minimal medium contributed to a selective production process, providing an important next step toward further development of nybomycin.

Miramides A-D: Identification of Detoxin-like Depsipeptides after Heterologous Expression of a Hybrid NRPS-PKS Gene Cluster from Streptomyces mirabilis Lu17588.

Natural products derived from plants, fungi or bacteria have been used for years in the medicine, agriculture and food industries as they exhibit a variety of beneficial properties, such as antibiotic, antifungal, anticancer, herbicidal and immunosuppressive activities. Compared to synthetic compounds, natural products possess a greater chemical diversity, which is a reason why they are profitable templates for developing pharmaceutical drug candidates and ongoing research on them is inevitable. Performing heterologous expression with unknown gene clusters is the preferred method to activate gene clusters that are not expressed in the wild-type strain under laboratory conditions; thus, this method offers a way to discover new interesting metabolites. Here, we report the gene cluster assembly of a hybrid NRPS-PKS gene cluster from Streptomyces mirabilis Lu17588, which was heterologously expressed in Streptomyces albus Del14. Four new compounds were produced by the obtained strain, which were named miramides A-D. Isolation and structure elucidation revealed similarity of the isolated compounds to the known depsipeptides rimosamides/detoxins.

Antibody Response 3 Months after 2 Doses of BNT162b2 mRNA COVID-19 Vaccine in Residents of Long-Term Care Facilities.

BACKGROUND: Older adults living in long-term care facilities (LTCFs) are at increased risk for severe outcomes from COVID-19 and were identified as a priority group in COVID-19 vaccination strategies. Emerging evidence suggests vaccine effectiveness in LTCF populations, but data about median and long-term durability of immune response after vaccination are still limited. OBJECTIVES: In this study, we assessed the humoral response to BNT162b2 mRNA COVID-19 vaccine 3 months after the second dose, in a cohort of 495 residents aged ≥65 years from 11 LTCF in Granada, Spain. METHOD: Between April 19 and April 30, 2021, we measured anti-SARS-CoV-2 Spike IgG to evaluate the humoral vaccination response. Antibody titers were reported in binding antibody units (BAU/mL). Bivariate and multivariate logistic regression models were performed to investigate the impact of age, sex, underlying health conditions, and prior COVID-19 infection on the antibody levels. RESULTS: Over 96% of the participants developed an adequate humoral response. We detected higher antibody titers in previously infected individuals, compared with those previously uninfected (B: 1,150.059 BAU/mL, p < 0.001). Moreover, we found a significant inverse association between age and antibody levels (B: -7.943 BAU/mL, p < 0.05). This negative age-dependent response was more noticeable among residents over 85 years old. In contrast, baseline health conditions and cognitive status were not associated with different antibody levels. CONCLUSIONS: These findings support monitoring COVID-19 vaccination response trend in older adults, in order to optimize future disease prevention and control strategies in this vulnerable population.

Bonsecamin: A New Cyclic Pentapeptide Discovered through Heterologous Expression of a Cryptic Gene Cluster.

The intriguing structural complexity of molecules produced by natural organisms is uncontested. Natural scaffolds serve as an important basis for the development of molecules with broad applications, e.g., therapeutics or agrochemicals. Research in recent decades has demonstrated that by means of classic metabolite extraction from microbes only a small portion of natural products can be accessed. The use of genome mining and heterologous expression approaches represents a promising way to discover new natural compounds. In this paper we report the discovery of a novel cyclic pentapeptide called bonsecamin through the heterologous expression of a cryptic NRPS gene cluster from Streptomyces albus ssp. chlorinus NRRL B-24108 in Streptomyces albus Del14. The new compound was successfully isolated and structurally characterized using NMR. The minimal set of genes required for bonsecamin production was determined through bioinformatic analysis and gene deletion experiments. A biosynthetic route leading to the production of bonsecamin is proposed in this paper.

Comparison between Aptima® assays (Hologic) and the CoBAS® 6800 system (Roche) for the diagnosis of sexually transmitted infections caused by Chlamydia trachomatis, Neisseria gonorrhoeae, and Mycoplasma genitalium.

Nowadays, it is of utmost importance to use fully validated assays for molecular-based diagnosis. In the field of sexually transmitted disease (STD), Roche and Hologic provide assays for diagnosing Chlamydia trachomatis (CT), Neisseria gonorrhoeae (NG), Mycoplasma genitalium (MG), and Trichomonas vaginalis (TV). A total of 212 clinical samples were tested. Aptima® Combo 2 (detecting CT and NG), Aptima® M. genitalium and the Aptima® T. vaginalis on the Panther® system were compared to CoBAS® CT/NG and CoBAS® TV/MG running on the CoBAS® 6800 system. To solve the discrepancies, Allplex™ STI Essential assay (Seegene®) and/or Sanger DNA sequencing were used. The diagnostic performance was calculated by mean of the sensitivity and specificity parameters. Aptima® (sensitivity: 98.90%, specificity: 100%), CoBAS® (sensitivity 100%, specificity: 96.67%). The CoBAS® combo (CT/NG) failed detecting NG from an anal/rectum specimen, which is not included into the validated specimens of the assay. Aptima® combo 2 produced two false positives (CT and NG), not detected by the third tests. All the assays showed an optimal diagnostic capacity, meeting the requirements for IVD DNA-based assays. All products work optimally on automatic platforms, minimizing time and risk of contamination during handling.

Dudomycins: New Secondary Metabolites Produced After Heterologous Expression of an Nrps Cluster from Streptomyces albus ssp. Chlorinus Nrrl B-24108.

Since the 1950s, natural products of bacterial origin were systematically developed to be used as drugs with a wide range of medical applications. The available treatment options for many diseases are still not satisfying, wherefore, the discovery of new structures has not lost any of its importance. Beyond the great variety of already isolated and characterized metabolites, Streptomycetes still harbor uninvestigated gene clusters whose products can be accessed using heterologous expression in host organisms. This works presents the discovery of a set of structurally novel secondary metabolites, dudomycins A to D, through the expression of a cryptic NRPS cluster from Streptomyces albus ssp. Chlorinus NRRL B-24108 in the heterologous host strain Streptomyces albus Del14. A minimal set of genes, required for the production of dudomycins, was defined through gene inactivation experiments. This paper also proposes a model for dudomycin biosynthesis.

Novel Fredericamycin Variant Overproduced by a Streptomycin-resistant Streptomyces albus subsp. chlorinus Strain.

Streptomycetes are an important source of natural products potentially applicable in the pharmaceutical industry. Many of these drugs are secondary metabolites whose biosynthetic genes are very often poorly expressed under laboratory cultivation conditions. In many cases, antibiotic-resistant mutants exhibit increased production of natural drugs, which facilitates the identification and isolation of new substances. In this study, we report the induction of a type II polyketide synthase gene cluster in the marine strain Streptomyces albus subsp. chlorinus through the selection of streptomycin-resistant mutants, resulting in overproduction of the novel compound fredericamycin C2 (1). Fredericamycin C2 (1) is structurally related to the potent antitumor drug lead fredericamycin A.

Invaginación intestinal como signo guía de enfermedad celíaca. Un caso clínico pediátrico / Intestinal intussusception as a key sign of Celiac Disease. A pediatric case report

La invaginación intestinal es la causa más frecuente de obstrucción intestinal entre los 6 y los 36 meses de edad. La mayoría son idiopáticas. Se ha descrito la asociación entre la enfermedad celíaca y la invaginación intestinal en la población pediátrica. Se presenta el caso de un varón de 23 meses ingresado por estancamiento ponderal en cuyo estudio ecográfico se observaron invaginaciones íleo-ileales asintomáticas repetidas.

Intestinal intussusception is the most frequent cause of intestinal obstruction between 6 and 36 months of age, the majority being idiopathic. The association between celiac disease and intestinal intussusception in the pediatric population has been described. We present the case of a 23-month-old male admitted due to a failure to thrive. In his ultrasound study recurrent asymptomatic ileo-ileal invaginations were found

[Intestinal intussusception as a key sign of Celiac Disease. A pediatric case report]. / Invaginación intestinal como signo guía de enfermedad celíaca. Un caso clínico pediátrico.

Intestinal intussusception is the most frequent cause of intestinal obstruction between 6 and 36 months of age, the majority being idiopathic. The association between celiac disease and intestinal intussusception in the pediatric population has been described. We present the case of a 23-month-old male admitted due to a failure to thrive. In his ultrasound study recurrent asymptomatic ileo-ileal invaginations were found.

La invaginación intestinal es la causa más frecuente de obstrucción intestinal entre los 6 y los 36 meses de edad. La mayoría son idiopáticas. Se ha descrito la asociación entre la enfermedad celíaca y la invaginación intestinal en la población pediátrica. Se presenta el caso de un varón de 23 meses ingresado por estancamiento ponderal en cuyo estudio ecográfico se observaron invaginaciones íleo-ileales asintomáticas repetidas.

Engineering of Streptomyces lividans for heterologous expression of secondary metabolite gene clusters.

BACKGROUND: Heterologous expression of secondary metabolite gene clusters is used to achieve increased production of desired compounds, activate cryptic gene clusters, manipulate clusters from genetically unamenable strains, obtain natural products from uncultivable species, create new unnatural pathways, etc. Several Streptomyces species are genetically engineered for use as hosts for heterologous expression of gene clusters. S. lividans TK24 is one of the most studied and genetically tractable actinobacteria, which remain untapped. It was therefore important to generate S. lividans chassis strains with clean metabolic backgrounds. RESULTS: In this study, we generated a set of S. lividans chassis strains by deleting endogenous gene clusters and introducing additional φC31 attB loci for site-specific integration of foreign DNA. In addition to the simplified metabolic background, the engineered S. lividans strains had better growth characteristics than the parental strain in liquid production medium. The utility of the developed strains was validated by expressing four secondary metabolite gene clusters responsible for the production of different classes of natural products. Engineered strains were found to be superior to the parental strain in production of heterologous natural products. Furthermore, S. lividans-based strains were better producers of amino acid-based natural products than other tested common hosts. Expression of a Streptomyces albus subsp. chlorinus NRRL B-24108 genomic library in the modified S. lividans ΔYA9 and S. albus Del14 strains resulted in the production of 7 potentially new compounds, only one of which was produced in both strains. CONCLUSION: The constructed S. lividans-based strains are a great complement to the panel of heterologous hosts for actinobacterial secondary metabolite gene expression. The expansion of the number of such engineered strains will contribute to an increased success rate in isolation of new natural products originating from the expression of genomic and metagenomic libraries, thus raising the chance to obtain novel biologically active compounds.

Detection of sexually transmitted disease-causing pathogens from direct clinical specimens with the multiplex PCR-based STD Direct Flow Chip Kit.

Pathogens causing sexually transmitted diseases (STDs) include viruses, bacteria, and parasites. The ability to rapidly and efficiently detect these pathogens in a single reaction still remains a health challenge. The aim of this study was to evaluate the clinical reliability and accuracy of the STD Direct Flow Chip Kit (Vitro, IVD-EC approved), which can simultaneously detect up to 9 different species of STD pathogens at once. This kit enables direct analysis-direct-PCR-of clinical specimens (urine, semen, endocervical, urethral, nasopharyngeal, and perianal swabs) without DNA purification for the following pathogens: Chlamydia trachomatis (serovars A-K and L1-L3), Haemophilus ducreyi, Herpes Simplex Virus (Types I and II), Mycoplasma genitalium, Mycoplasma hominis, Neisseria gonorrhoeae, Treponema pallidum, Trichomonas vaginalis, and Ureaplasma. The Anyplex™ II STI-7 Detection Kit (Seegene, IVD-EC) was used as the reference's method. Existing discordances were resolved using either a third molecular assay or DNA sequencing. Clinical performance was evaluated at two different stages: (i) from purified DNA of three hundred and fifty-eight clinical specimens with a diagnostic sensitivity (SE) and specificity (SP) of 99.4% and 100%, respectively, and an agreement of 99% (kappa index, κ = 0.97) with the reference's method and; (ii) by direct-PCR from six hundred and thirty-three specimens rendering SE, SP, and agreement values of 98.4%, 99.9%, and 98.0% (κ = 0.95), respectively. The STD Direct Flow Chip Kit constitutes a promising alternative to routine procedures in diagnostic, allowing direct analysis of specimens and enabling the detection of a broad panel of pathogens.

Benzanthric Acid, a Novel Metabolite From Streptomyces albus Del14 Expressing the Nybomycin Gene Cluster.

Streptomycetes constitute a diverse bacterial group able to produce a wide variety of secondary metabolites with potential applications in the pharmacy industry. However, the genes responsible for the biosynthesis of these compounds are very frequently inactive or expressed at very low levels under standard laboratory cultivation conditions. Therefore, the activation or upregulation of secondary metabolite biosynthesis genes is a crucial step for the discovery of new bioactive natural products. We have recently reported the discovery of the biosynthetic genes for the antibiotic nybomycin (nyb genes) in Streptomyces albus subsp. chlorinus. The nyb genes were expressed in the heterologous host Streptomyces albus Del14, which produces not only nybomycin, but also a novel compound. In this study, we describe the isolation, purification, and structure elucidation of the new substance named benzanthric acid.

Heterologous Expression of the Nybomycin Gene Cluster from the Marine Strain Streptomyces albus subsp. chlorinus NRRL B-24108.

Streptomycetes represent an important reservoir of active secondary metabolites with potential applications in the pharmaceutical industry. The gene clusters responsible for their production are often cryptic under laboratory growth conditions. Characterization of these clusters is therefore essential for the discovery of new microbial pharmaceutical drugs. Here, we report the identification of the previously uncharacterized nybomycin gene cluster from the marine actinomycete Streptomyces albus subsp. chlorinus through its heterologous expression. Nybomycin has previously been reported to act against quinolone-resistant Staphylococcus aureus strains harboring a mutated gyrA gene but not against those with intact gyrA. The nybomycin-resistant mutants generated from quinolone-resistant mutants have been reported to be caused by a back-mutation in the gyrA gene that restores susceptibility to quinolones. On the basis of gene function assignment from bioinformatics analysis, we suggest a model for nybomycin biosynthesis.

Validación de un método seguro y sencillo para la elaboración de secuencias consenso del virus de la inmunodeficiencia humana a partir de los datos de secuenciación masiva 454 / A safe an easy method for building consensus HIV sequences from 454 massively parallel sequencing data

Objetivo: Generar una secuencia consenso a partir de los datos de secuenciación masiva obtenidos en estudios de resistencias a antiretrovirales, que sea representativa de la secuencia Sanger y que sirva para estudios de epidemiología molecular. Material y métodos: En 62 pacientes se obtuvo la secuencia de transcriptasa reversa-proteasa, mediante Sanger (Trugene-Siemens), y NGS (454GSJunior-Roche). Las secuencias consenso NGS se generaron con Mesquite, seleccionando umbrales 10%, 15% y 20%. Para el estudio filogenético se empleó MEGA. Resultados: Utilizando el umbral 10%, 17/62 pacientes presentaron secuencias pareadas NGS-Sanger, con una mediana de bootstrap del 88% (IQR83,5-95,5). La asociación aumenta a 36/62 pacientes y el bootstrap, a 94% (IQR85,5-98), y alcanza el máximo al 20% en 61/62 pacientes, bootstrap 99% (IQR98-100). Conclusión: Mostramos un método seguro para generar secuencias consenso NGS para su uso en estudios de epidemiología molecular procesadas con umbral 20%, de fácil uso y aplicación en los servicios de microbiología clínica (AU)

Objective: To show how to generate a consensus sequence from the information of massive parallel sequences data obtained from routine HIV anti-retroviral resistance studies, and that may be suitable for molecular epidemiology studies. Material and methods: Paired Sanger (Trugene-Siemens) and next-generation sequencing (NGS) (454 GSJunior-Roche) HIV RT and protease sequences from 62 patients were studied. NGS consensus sequences were generated using Mesquite, using 10%, 15%, and 20% thresholds. Molecular evolutionary genetics analysis (MEGA) was used for phylogenetic studies. Results: At a 10% threshold, NGS-Sanger sequences from 17/62 patients were phylogenetically related, with a median bootstrap-value of 88% (IQR83.5-95.5). Association increased to 36/62 sequences, median bootstrap 94% (IQR85.5-98)], using a 15% threshold. Maximum association was at the 20% threshold, with 61/62 sequences associated, and a median bootstrap value of 99% (IQR98-100). Conclusion: A safe method is presented to generate consensus sequences from HIV-NGS data at 20% threshold, which will prove useful for molecular epidemiological studies (AU)

A safe an easy method for building consensus HIV sequences from 454 massively parallel sequencing data. / Validación de un método seguro y sencillo para la elaboración de secuencias consenso del virus de la inmunodeficiencia humana a partir de los datos de secuenciación masiva 454.

OBJECTIVE: To show how to generate a consensus sequence from the information of massive parallel sequences data obtained from routine HIV anti-retroviral resistance studies, and that may be suitable for molecular epidemiology studies. MATERIAL AND METHODS: Paired Sanger (Trugene-Siemens) and next-generation sequencing (NGS) (454 GSJunior-Roche) HIV RT and protease sequences from 62 patients were studied. NGS consensus sequences were generated using Mesquite, using 10%, 15%, and 20% thresholds. Molecular evolutionary genetics analysis (MEGA) was used for phylogenetic studies. RESULTS: At a 10% threshold, NGS-Sanger sequences from 17/62 patients were phylogenetically related, with a median bootstrap-value of 88% (IQR83.5-95.5). Association increased to 36/62 sequences, median bootstrap 94% (IQR85.5-98)], using a 15% threshold. Maximum association was at the 20% threshold, with 61/62 sequences associated, and a median bootstrap value of 99% (IQR98-100). CONCLUSION: A safe method is presented to generate consensus sequences from HIV-NGS data at 20% threshold, which will prove useful for molecular epidemiological studies.

Los estudios de resistencias a antirretrovirales como herramienta para el análisis de los clusters de transmisión del virus de la inmunodeficiencia humana / Study of human immunodeficiency virus transmission chains in Andalusia: Analysis from baseline antiretroviral resistance sequences

INTRODUCCIÓN Y OBJETIVO: Las secuencias de proteasa y transcriptasa reversa del VIH-1 aportan una información muy valiosa para el manejo de la infección por VIH, más allá de la información de resistencias a los antirretrovirales. En nuestro estudio la hemos utilizado para evaluar las cadenas de transmisión, la transmisión de resistencias entre ellos, y para conocer la distribución espacial de los diferentes subtipos utilizando técnicas de georreferenciación. MÉTODOS: Hemos estudiado 693 pacientes diagnosticados de VIH-1 durante el periodo 2005-2012, todos ellos residentes en Andalucía Oriental. La secuencia del gen pol (transcriptasa reversa y proteasa) se generó utilizando Trugene® HIV Genotyping Kit (Siemens, NAD). La historia evolutiva fue inferida a través de MEGA 5.2 mediante el método de Neighbor-Joining. Para la filogeografía y el estudio de resistencias utilizamos ArcGIS y REGA. RESULTADOS: Doscientos noventa y ocho pacientes se asociaron en 77 clusters diferentes. La mayoría de los cluster estaban formados por parejas (n = 49), de hombres que practican sexo con hombres (n = 26), de nacionalidad española (n = 37), con una edad menor a 45 años (73,5%). Las áreas de mayor heterogeneidad de subtipos fueron el área metropolitana de Granada y las zonas de costa de Almería y Granada. Hemos encontrado 5 cluster con más de 10 individuos. En 15 cluster detectamos mutaciones de resistencia. CONCLUSIONES: Presentamos datos que demuestran que el estudio epidemiológico de los diferentes clusters de transmisión de VIH mediante análisis filogenético se presenta como una herramienta potente y de gran utilidad para la vigilancia y control epidemiológico de la propagación del VIH, que puede ayudar a diseñar actuaciones eficaces para prevenir la diseminación del VIH

INTRODUCTION AND OBJECTIVE: Protease and reverse transcriptase HIV-1 sequences provide useful information for patient clinical management, as well as information on resistance to antiretrovirals. The aim of this study is to evaluate transmission events, transmitted drug resistance, and to georeference subtypes among newly diagnosed patients referred to our center. METHODS: A study was conducted on 693 patients diagnosed between 2005 and 2012 in Southern Spain. Protease and reverse transcriptase sequences were obtained for resistance to cART analysis with Trugene® HIV Genotyping Kit (Siemens, NAD). MEGA 5.2, Neighbor-Joining, ArcGIS and REGA were used for subsequent analysis. RESULTS: The results showed 298 patients clustered into 77 different transmission events. Most of the clusters were formed by pairs (n = 49), of men having sex with men (n = 26), Spanish (n = 37), and below 45 years of age (73.5%). Urban areas from Granada, and the coastal areas of Almeria and Granada showed the greatest subtype heterogeneity. Five clusters were formed by more than 10 patients, and 15 clusters had transmitted drug resistance. CONCLUSIONS: The study data demonstrate how the phylogenetic characterization of transmission clusters is a powerful tool to monitor the spread of HIV, and may contribute to design correct preventive measures to minimize it

Diagnóstico microbiológico de la infección por el virus de la inmunodeficiencia humana / Microbiological diagnosis of human immunodeficiency virus infection

El presente documento intenta reflejar y actualizar las principales tareas y cometidos que un laboratorio de microbiología debería tener para realizar el diagnóstico y seguimiento de los pacientes infectados por el VIH. Se distribuye en 3 apartados: en el primero se trata el diagnóstico serológico, que en los últimos años ha sufrido una importante renovación, y en el que hemos procurado adecuarnos a las demandas diagnósticas y epidemiológicas actuales, para que desde los laboratorios podamos contribuir a no perder oportunidades de diagnóstico. En una segunda parte se describe la determinación de la carga viral plasmática, y se hace una exhaustiva revisión de los avances tecnológicos y de las recomendaciones actuales, además de abordar un tema de enorme interés clínico, la significación de la viremia persistente de bajo grado. Finalmente, en el tercer apartado se desarrolla el tema de las resistencias a los fármacos antirretrovirales tanto en pacientesnaive como en fracaso, analizando la transcriptasa reversa, la proteasa y la integrasa, e incorporando como novedades las técnicas de determinación del tropismo viral, y el papel de las variantes minoritarias

This document attempts to update the main tasks and roles of the Clinical Microbiology laboratory in HIV diagnosis and monitoring. The document is divided into three parts. The first deals with HIV diagnosis and how serological testing has changed in the last few years, aiming to improve diagnosis and to minimize missed opportunities for diagnosis. Technological improvements for HIV Viral Load are shown in the second part of the document, which also includes a detailed description of the clinical significance of low-level and very low-level viremia. Finally, the third part of the document deals with resistance to antiretroviral drugs, incorporating clinical indications for integrase and tropism testing, as well as the latest knowledge on minority variants

[Study of human immunodeficiency virus transmission chains in Andalusia: analysis from baseline antiretroviral resistance sequences]. / Los estudios de resistencias a antirretrovirales como herramienta para el análisis de los clusters de transmisión del virus de la inmunodeficiencia humana.

INTRODUCTION AND OBJECTIVE: Protease and reverse transcriptase HIV-1 sequences provide useful information for patient clinical management, as well as information on resistance to antiretrovirals. The aim of this study is to evaluate transmission events, transmitted drug resistance, and to georeference subtypes among newly diagnosed patients referred to our center. METHODS: A study was conducted on 693 patients diagnosed between 2005 and 2012 in Southern Spain. Protease and reverse transcriptase sequences were obtained for resistance to cART analysis with Trugene(®) HIV Genotyping Kit (Siemens, NAD). MEGA 5.2, Neighbor-Joining, ArcGIS and REGA were used for subsequent analysis. RESULTS: The results showed 298 patients clustered into 77 different transmission events. Most of the clusters were formed by pairs (n=49), of men having sex with men (n=26), Spanish (n=37), and below 45 years of age (73.5%). Urban areas from Granada, and the coastal areas of Almeria and Granada showed the greatest subtype heterogeneity. Five clusters were formed by more than 10 patients, and 15 clusters had transmitted drug resistance. CONCLUSIONS: The study data demonstrate how the phylogenetic characterization of transmission clusters is a powerful tool to monitor the spread of HIV, and may contribute to design correct preventive measures to minimize it.