Dispersal history of SARS-CoV-2 in Galicia, Spain.

The dynamics of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) transmission are influenced by a variety of factors, including social restrictions and the emergence of distinct variants. In this study, we delve into the origins and dissemination of the Alpha, Delta, and Omicron-BA.1 variants of concern in Galicia, northwest Spain. For this, we leveraged genomic data collected by the EPICOVIGAL Consortium and from the GISAID database, along with mobility information from other Spanish regions and foreign countries. Our analysis indicates that initial introductions during the Alpha phase were predominantly from other Spanish regions and France. However, as the pandemic progressed, introductions from Portugal and the United States became increasingly significant. The number of detected introductions varied from 96 and 101 for Alpha and Delta to 39 for Omicron-BA.1. Most of these introductions left a low number of descendants (<10), suggesting a limited impact on the evolution of the pandemic in Galicia. Notably, Galicia's major coastal cities emerged as critical hubs for viral transmission, highlighting their role in sustaining and spreading the virus. This research emphasizes the critical role of regional connectivity in the spread of SARS-CoV-2 and offers essential insights for enhancing public health strategies and surveillance measures.

Dispersal history of SARS-CoV-2 in Galicia, Spain.

The dynamics of SARS-CoV-2 transmission are influenced by a variety of factors, including social restrictions and the emergence of distinct variants. In this study, we delve into the origins and dissemination of the Alpha, Delta, and Omicron variants of concern in Galicia, northwest Spain. For this, we leveraged genomic data collected by the EPICOVIGAL Consortium and from the GISAID database, along with mobility information from other Spanish regions and foreign countries. Our analysis indicates that initial introductions during the Alpha phase were predominantly from other Spanish regions and France. However, as the pandemic progressed, introductions from Portugal and the USA became increasingly significant. Notably, Galicia's major coastal cities emerged as critical hubs for viral transmission, highlighting their role in sustaining and spreading the virus. This research emphasizes the critical role of regional connectivity in the spread of SARS-CoV-2 and offers essential insights for enhancing public health strategies and surveillance measures.

Anal Dysplasia Screening in People Living with HIV: Long-Term Follow-Up in a Large Cohort from Northwest Spain.

Anal squamous cell carcinoma (SCC) is not a common disease in the general population, although its incidence is higher in people living with human immunodeficiency virus (PLWH). Anal SCC is caused by human papillomavirus (HPV) infection and arises from premalignant lesions termed squamous intraepithelial lesions (SILs). SIL surveillance programs are based on the early detection and treatment of SILs, especially those with a higher risk of transforming into cancer. An anal surveillance program has been under development in our institution since 2011. In this context, we performed a retrospective cohort study at the anal dysplasia unit of Álvaro-Cunqueiro Hospital (Spain). Epidemiological and clinical data were gathered from our Infectious Diseases Sample Collection (an open sample cohort including PLWH) from January 2011 to January 2022. A total of 493 PLWH were considered, 122 (24.7%) of whom were diagnosed with anal dysplasia at baseline, including 2 cases of anal SCC. Briefly, most of individuals were young men (median age, 38 years old) born in Spain (76%), whose vaccination rate before their inclusion in the program was scarce (<3%). Throughout the study period, 81 (16.4%) cases were diagnosed with high-grade squamous-intraepithelial lesions (HSILs) and 3 with anal SCC. At the baseline, severe immunosuppression (i.e., nadir CD4+ lymphocyte count below 200 cell/µL), and prior diagnosis of condyloma acuminata were more frequent within the group with SILs. Conversely, the baseline CD4+ lymphocyte count was similar among both groups. HPV-16 was related to a higher risk of HSILs (odds ratio: 2.76). At the end of the follow-up, 385 PLWH had been retained in care; one patient had died of anal cancer. Anal dysplasia was common (25% of cases), especially among patients infected by HPV-16, diagnosed with condyloma acuminata, and who were severely immunosuppressed. HPV-16 was the main risk factor for the presentation of HSILs.

The imperative for quality control programs in Monkeypox virus DNA testing by PCR: CIBERINFEC quality control.

To evaluate molecular assays for Mpox diagnosis available in various clinical microbiology services in Spain through a quality control (QC) approach. A total of 14 centers from across Spain participated in the study. The Reference Laboratory dispatched eight serum samples and eight nucleic acid extracts to each participating center. Some samples were spiked with Mpox or Vaccinia virus to mimic positive samples for Mpox or other orthopox viruses. Participating centers provided information on the results obtained, as well as the laboratory methods used. Among the 14 participating centers seven different commercial assays were employed, with the most commonly used kit being LightMix Modular Orthopox/Monkeypox (Mpox) Virus (Roche®). Of the 12 centers conducting Mpox determinations, concordance ranged from 62.5% (n = 1) to 100% (n = 11) for eluates and from 75.0% (n = 1) to 100% (n = 10) for serum. Among the 10 centers performing Orthopoxvirus determinations, a 100% concordance was observed for eluates, while for serum, concordance ranged from 87.5% (n = 6) to 100% (n = 4). Repeatedly, 6 different centers reported a false negative in serum samples for Orthopoxvirus diagnosis, particularly in a sample with borderline Ct = 39. Conversely, one center, using the TaqMan™ Mpox Virus Microbe Detection Assay (Thermo Fisher), reported false positives in Mpox diagnosis for samples spiked with vaccinia virus due to cross-reactions. We observed a positive correlation of various diagnostic assays for Mpox used by the participating centers with the reference values. Our results highlight the significance of standardization, validation, and ongoing QC in the microbiological diagnosis of infectious diseases, which might be particularly relevant for emerging viruses.

The Modification of the Illumina® CovidSeq™ Workflow for RSV Genomic Surveillance: The Genetic Variability of RSV during the 2022-2023 Season in Northwest Spain.

There is growing interest in the molecular surveillance of the Respiratory Syncytial Virus and the monitorization of emerging mutations that could impair the efficacy of antiviral prophylaxis and treatments. A simple, scalable protocol for viral nucleic acid enrichment could improve the surveillance of RSV. We developed a protocol for RSV-A and B amplification based on the Illumina CovidSeq workflow using an RSV primer panel. A total of 135 viral genomes were sequenced from nasopharyngeal samples through the optimization steps of this panel, while an additional 15 samples were used to test the final version. Full coverage of the G gene and over 95% of the coverage of the F gene, the target of the available RSV antivirals or monoclonal antibodies, were obtained. The F:K68N mutation, associated with decreased nirsevimab activity, was detected in our facility. Additionally, phylogenetic analysis showed several sublineages in the 2022-2023 influenza season in Europe. Our protocol allows for a simple and scalable simultaneous amplification of the RSV-A and B whole genome, increasing the yield of RSV sequencing and reducing costs. Its application would allow the world to be ready for the detection of arising mutations in relation to the widespread use of nirsevimab for RSV prevention.

Enhancing SARS-CoV-2 Surveillance through Regular Genomic Sequencing in Spain: The RELECOV Network.

Millions of SARS-CoV-2 whole genome sequences have been generated to date. However, good quality data and adequate surveillance systems are required to contribute to meaningful surveillance in public health. In this context, the network of Spanish laboratories for coronavirus (RELECOV) was created with the main goal of promoting actions to speed up the detection, analyses, and evaluation of SARS-CoV-2 at a national level, partially structured and financed by an ECDC-HERA-Incubator action (ECDC/GRANT/2021/024). A SARS-CoV-2 sequencing quality control assessment (QCA) was developed to evaluate the network's technical capacity. QCA full panel results showed a lower hit rate for lineage assignment compared to that obtained for variants. Genomic data comprising 48,578 viral genomes were studied and evaluated to monitor SARS-CoV-2. The developed network actions showed a 36% increase in sharing viral sequences. In addition, analysis of lineage/sublineage-defining mutations to track the virus showed characteristic mutation profiles for the Delta and Omicron variants. Further, phylogenetic analyses strongly correlated with different variant clusters, obtaining a robust reference tree. The RELECOV network has made it possible to improve and enhance the genomic surveillance of SARS-CoV-2 in Spain. It has provided and evaluated genomic tools for viral genome monitoring and characterization that make it possible to increase knowledge efficiently and quickly, promoting the genomic surveillance of SARS-CoV-2 in Spain.

Proof of concept of the potential of a machine learning algorithm to extract new information from conventional SARS-CoV-2 rRT-PCR results.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has been and remains one of the major challenges modern society has faced thus far. Over the past few months, large amounts of information have been collected that are only now beginning to be assimilated. In the present work, the existence of residual information in the massive numbers of rRT-PCRs that tested positive out of the almost half a million tests that were performed during the pandemic is investigated. This residual information is believed to be highly related to a pattern in the number of cycles that are necessary to detect positive samples as such. Thus, a database of more than 20,000 positive samples was collected, and two supervised classification algorithms (a support vector machine and a neural network) were trained to temporally locate each sample based solely and exclusively on the number of cycles determined in the rRT-PCR of each individual. Overall, this study suggests that there is valuable residual information in the rRT-PCR positive samples that can be used to identify patterns in the development of the SARS-CoV-2 pandemic. The successful application of supervised classification algorithms to detect these patterns demonstrates the potential of machine learning techniques to aid in understanding the spread of the virus and its variants.

Influence of perinatal and childhood exposure to tobacco and mercury in children's gut microbiota.

Background: Early life determinants of the development of gut microbiome composition in infants have been widely investigated; however, if early life pollutant exposures, such as tobacco or mercury, have a persistent influence on the gut microbial community, its stabilization at later childhood remains largely unknown. Objective: In this exposome-wide study, we aimed at identifying the contribution of exposure to tobacco and mercury from the prenatal period to childhood, to individual differences in the fecal microbiome composition of 7-year-old children, considering co-exposure to a width of established lifestyle and clinical determinants. Methods: Gut microbiome was studied by 16S rRNA amplicon sequencing in 151 children at the genus level. Exposure to tobacco was quantified during pregnancy through questionnaire (active tobacco consumption, second-hand smoking -SHS) and biomonitoring (urinary cotinine) at 4 years (urinary cotinine, SHS) and 7 years (SHS). Exposure to mercury was quantified during pregnancy (cord blood) and at 4 years (hair). Forty nine other potential environmental determinants (12 at pregnancy/birth/infancy, 15 at 4 years and 22 at 7 years, such as diet, demographics, quality of living/social environment, and clinical records) were registered. We used multiple models to determine microbiome associations with pollutants including multi-determinant multivariate analysis of variance and linear correlations (wUnifrac, Bray-Curtis and Aitchison ß-diversity distances), single-pollutant permutational multivariate analysis of variance adjusting for co-variates (Aitchison), and multivariable association model with single taxa (MaAsLin2; genus). Sensitivity analysis was performed including genetic data in a subset of 107 children. Results: Active smoking in pregnancy was systematically associated with microbiome composition and ß-diversity (R2 2-4%, p < 0.05, Aitchison), independently of other co-determinants. However, in the adjusted single pollutant models (PERMANOVA), we did not find any significant association. An increased relative abundance of Dorea and decreased relative abundance of Akkermansia were associated with smoking during pregnancy (q < 0.05). Discussion: Our findings suggest a long-term sustainable effect of prenatal tobacco exposure on the children's gut microbiota. This effect was not found for mercury exposure or tobacco exposure during childhood. Assessing the role of these exposures on the children's microbiota, considering multiple environmental factors, should be further investigated.

Silver Nanoparticles Produced by Laser Ablation and Re-Irradiation Are Effective Preventing Peri-Implantitis Multispecies Biofilm Formation.

Implant-associated infection due to biofilm formation is a growing problem. Given that silver nanoparticles (Ag-NPs) have shown antibacterial effects, our goal is to study their effect against multispecies biofilm involved in the development of peri-implantitis. To this purpose, Ag-NPs were synthesized by laser ablation in de-ionized water using two different lasers, leading to the production of colloidal suspensions. Subsequently, part of each suspension was subjected to irradiation one and three times with the same laser source with which it was obtained. Ag-NPs were immobilized on the surface of titanium discs and the resultant materials were compared with unmodified titanium coupons. Nanoparticles were physico-chemically analysed to determine their shape, crystallinity, chemical composition, and mean diameter. The materials were incubated for 90 min or 48 h, to evaluate bacterial adhesion or biofilm formation respectively with Staphylococcus aureus or oral mixed bacterial flora composed of Streptococcus oralis, Actinomyces naeslundii, Veionella dispar, and Porphyromonas gingivalis. Ag-NPs help prevent the formation of biofilms both by S. aureus and by mixed oral bacterial flora. Nanoparticles re-irradiated three times showed the biggest antimicrobial effects. Modifying dental implants in this way could prevent the development of peri-implantitis.

Gastrointestinal colonization by OXA-48-producing Enterobacterales: risk factors for persistent carriage.

Carbapenem-resistant Enterobacterales (CRE) infections are a major health problem. Intestinal colonization is a key factor in developing infection. However, factors associated with persistent colonization by CRE are unknown. The aim of the study was to identify factors associated with persistent CRE gut colonization. This is a retrospective, single-centre, observational study of adult patients with CRE gut colonization between January 2015 and January 2020. Epidemiologic characteristics, comorbidities, infectious events, duration of hospitalization and antimicrobial treatment received in the follow-up period were collected. Colonization was defined as isolation in at least 2 rectal swab culture samples of CRE. Decolonization was defined as 3 negative rectal swab cultures or 2 negative cultures and a negative molecular test. A cohort of 86 patients with CRE gut colonization was selected: 44 patients with spontaneous decolonization (DC) and 42 patients with persistent colonization (PC). The mean follow-up period was 24 months (IQR 14-33) in the DC group vs. 25 months (IQR 16-36) in the PC group (p = 0.478). Patient characteristics were similar between both groups. Colonization by other MDR microorganisms was high (44 patients, 51%) and slightly more common in the PC group (PC 60% vs. DC 43%, p = 0.139). The use of ceftazidime-avibactam was more common among the PC group (PC 33% vs. DC 14%, p = 0.041). We observed a higher percentage of antimicrobial therapy in the previous 30 days (PC 68% vs. DC 57%, p = 0.371) and 90 days (PC 81% vs. DC 82%, p = 0.353) in the PC group. Multivariable analysis showed that patients that have received ceftazidime-avibactam therapy (OR 4.9 95% CI [1.45-16.39], p = 0.010), and those colonized by other MDR microorganisms (OR 2.5, 95% CI [0.96-6.25], p = 0.060) presented a higher risk of PC. Ceftazidime-avibactam use and colonization by other MDR microorganisms might be associated with CRE persistent gut colonization.

Case report: BA.1 subvariant showing a BA.2-like pattern using a variant-specific PCR assay due to a single point mutation downstream the spike 69/70 deletion.

BACKGROUND: SARS-CoV-2 variant tracking is key to the genomic surveillance of the COVID-19 pandemic. While next-generation sequencing (NGS) is commonly used for variant determination, it is expensive and time-consuming. Variant-specific PCR (vsPCR) is a faster, cheaper method that detects specific mutations that are considered variant-defining. These tests usually rely on specific amplification when a mutation is present or a specific melting temperature peak after amplification. CASE PRESENTATION: A discrepant result between vsPCR and NGS was found in seventeen SARS-CoV-2 samples from Galicia, Spain. A cluster of BA.1 Omicron SARS-CoV-2 variant showed a BA.2-like melting temperature pattern due to a point mutation (C21772T) downstream the deletion of the spike amino acids 69/70. As the 69/70 deletion is widely used for differentiation between BA.1 and BA.2 by vsPCR, C21772T can cause BA.1 samples to be misinterpreted as BA.2. Over a thousand BA.1 sequences in the EpiCoV database contain this mutation. CONCLUSIONS: To our knowledge, this is the first case of a point mutation causing a vsPCR algorithm to misclassify BA.1 samples as BA.2. This is an example of how mutations in the probe target area of vsPCR tests based on melting curve analysis can lead to variant misclassification. NGS confirmation of vsPCR results is relevant for the accuracy of the epidemiological surveillance. In order to overcome the possible impact of novel mutations, diagnostic tools must be constantly updated.

In vitro neutralizing activity of BNT162b2 mRNA-induced antibodies against full B.1.351 SARS-CoV-2 variant.

SARS-CoV-2 variation represents a serious challenge to current COVID-19 vaccines. Recent reports suggest that B.1.351 and other variants may escape the neutralization activity of the antibodies generated by current vaccines. Ninety-nine healthcare workers undertaking BNT162b2 mRNA vaccination were sampled at baseline, on the day of the second dose, and 14 days after the latter. Neutralization activity against SARS-CoV-2 B.1, B.1.1.7 and B.1.351 was investigated using a Vero-E6 model. Eleven of the study participants had prior infection with SARS-CoV-2. Neutralization titers against the B.1 and the B.1.1.7 variants were not statistically different and were significantly higher than titers against the B.1.351 variant across pre-exposed and non-pre-exposed vaccinated individuals (p < .01). While all vaccinated individuals presented neutralizing antibodies against B.1 and B 1.1.7 after the second dose, 14% were negative against B.1.351 and 76% had low titers (1/201/80). Pre-exposed vaccinated individuals showed higher titers than non-pre-exposed after the first (median titers of 1/387 versus 1/28, respectively) and the second doses (1/995 versus 1/703, respectively). As high as 72% of the pre-exposed vaccines presented titers >1/80 after a single dose, while only 11% of non-exposed vaccinated individuals had titers >1/80. BNT162b2 mRNA-induced antibodies show a lower in vitro neutralizing activity against B.1.351 variant compared to neutralization against B.1.1.7 or B.1 variants. Interestingly, for individuals pre-exposed to SARS-CoV-2, one dose of BNT162b2 mRNA may be adequate to produce neutralizing antibodies against B.1.1.7 and B.1, while two doses of BNT162b2 mRNA provide optimal neutralizing antibody response against B.1.351 too.

Pooling for SARS-CoV-2 control in care institutions.

BACKGROUND: Workers and residents in Care Homes are considered at special risk for the acquisition of SARS-CoV-2 infection, due to the infectivity and high mortality rate in the case of residents, compared to other containment areas. The role of presymptomatic people in transmission has been shown to be important and the early detection of these people is critical for the control of new outbreaks. Pooling strategies have proven to preserve SARS-CoV-2 testing resources. The aims of the present study, based in our local experience, were (a) to describe SARS-CoV-2 prevalence in institutionalized people in Galicia (Spain) during the Coronavirus pandemic and (b) to evaluate the expected performance of a pooling strategy using RT-PCR for the next rounds of screening of institutionalized people. METHODS: A total of 25,386 Nasopharyngeal swab samples from the total of the residents and workers at Care Homes in Galicia (March to May 2020) were individually tested using RT-PCR. Prevalence and quantification cycle (Cq) value distribution of positives was calculated. Besides, 26 pools of 20 samples and 14 pools of 5 samples were tested using RT-PCR as well (1 positive/pool). Pooling proof of concept was performed in two populations with 1.7 and 2% prevalence. RESULTS: Distribution of SARS-CoV-2 infection at Care Homes was uneven (0-60%). As the virus circulation global rate was low in our area (3.32%), the number of people at risk of acquiring the infection continues to be very high. In this work, we have successfully demonstrated that pooling of different groups of samples at low prevalence clusters, can be done with a small average delay on Cq values (5 and 2.85 cycles for pools of 20 and 5 samples, respectively). CONCLUSIONS: A new screening system with guaranteed protection is required for small clusters, previously covered with individual testing. Our proposal for Care Homes, once prevalence zero is achieved, would include successive rounds of testing using a pooling solution for transmission control preserving testing resources. Scale-up of this method may be of utility to confront larger clusters to avoid the viral circulation and keeping them operative.

Enterovirus D68-associated respiratory and neurological illness in Spain, 2014-2018.

During 2014, enterovirus D68 (EV-D68) outbreaks were described globally, causing severe respiratory diseases in children and, in some cases, subsequent paralysis. In this study, the type characterization of enterovirus (EV) detected in respiratory illnesses and the epidemiology and clinical association of EV-D68 infections in Spain over a five-year period were described. A total of 546 EV-positive samples from hospitalized patients with respiratory infections were included. EV-D68 was the most frequently detected type (46.6%, 191/410 typed EV). Other EV from species A (25.1%), B (27.8%) and C (0.5%) were also identified. EV-D68 infections were more associated with bronchitis while EV-A/B types were more frequent in upper respiratory illness (p < 0.01). EV-D68 was also detected in patients with neurological symptoms (nine meningitis/meningoencephalitis and eight acute flaccid paralysis cases). Phylogenetic analysis of 3'-VP1 region showed most Spanish EV-D68 sequences from 2014 to 2016 belonged to subclades B2/B3, as other American and European strains circulating during the same period. However, those detected in 2017 and 2018 clustered to the emerged subclade D1. In summary, different EV can cause respiratory infections but EV-D68 was the most prevalent, with several strains circulating in Spain at least since 2014. Association between EV-D68 infection and neurological disease was also described.

Prevalencia y distribución de los genotipos del virus de la hepatitis C en Galicia durante el periodo 2000-2015 / Prevalence and distribution of hepatitis C virus genotypes in Galicia during the period 2000-2015

INTRODUCCIÓN: Presentamos el mayor estudio realizado en Galicia sobre prevalencia y distribución de genotipos/subtipos del VHC. MÉTODOS: Estudio retrospectivo recopilando al total de pacientes crónicamente infectados por VHC entre el 1 de enero del 2000 y el 31 de diciembre del 2015 en 3 de las principales áreas sanitarias: Santiago, Pontevedra y Vigo. Recogimos datos sobre prevalencia y distribución de genotipos/subtipos, vías de transmisión, género, edad y coinfección con VIH/VHB. RESULTADOS: Reclutamos a 4.469 pacientes, con una mediana de edad 50 años (RIC 57-45), el 72,3% hombres, el 0,4% coinfectado con otro genotipo, el 20,6% coinfectado con VIH y un 35,2% con VHB. Transmisión parenteral (83,1%), desconocida (15,3%), sexual (1,4%) y vertical (0,2%). Distribución de genotipos: 62,9% VHC-1 (29,2% VHC-1a y 31,9% VHC-1b), 3,4% VHC-2, 21,0% VHC-3, 12,6% VHC-4 y 0,1% VHC-5. CONCLUSIÓN: La distribución de genotipos muestra diferencias significativas respecto a la observada en España. Dicha distribución varía con la edad, el género, la coinfección con VIH o VHB, y dentro de áreas geográficas

INTRODUCTION: We present the largest study conducted in Galicia on the prevalence and distribution of HCV genotypes/subtypes. METHODS: Retrospective study collecting the total number of patients chronically infected by HCV between 2000.01.01 to 2015.12.31 in 3 of the main health areas: Santiago, Pontevedra and Vigo. RESULTS: We collected a total of 4469 patients. The median age was 50 years (IQR 57-45), 72,3% were men, 0,4% were coinfected with another genotype, 20,6% were coinfected with HIV and 35.2% with HBV. The main route of transmission was parenteral (83,1%), followed by unknown (15,3%), sexual (1,4%) and vertical (0,2%). The distribution of genotypes was: 62,9% HCV-1 (29,2% HCV-1a and 31,9% HCV-1b), 3,4% HCV-2, 21,0% HCV-3, 12,6% HCV-4 and 0,1% HCV-5. CONCLUSION: The distribution of genotypes in Galicia shows significant differences with respect to that observed in Spain. This distribution varies with age, gender, coinfection with HIV and/or HBV, and within geographical areas

Prevalence and distribution of hepatitis C virus genotypes in Galicia during the period 2000-2015. / Prevalencia y distribución de los genotipos del virus de la hepatitis C en Galicia durante el periodo 2000-2015.

INTRODUCTION: We present the largest study conducted in Galicia on the prevalence and distribution of HCV genotypes/subtypes. METHODS: Retrospective study collecting the total number of patients chronically infected by HCV between 2000.01.01 to 2015.12.31 in 3of the main health areas: Santiago, Pontevedra and Vigo. RESULTS: We collected a total of 4469 patients. The median age was 50 years (IQR 57-45), 72,3% were men, 0,4% were coinfected with another genotype, 20,6% were coinfected with HIV and 35.2% with HBV. The main route of transmission was parenteral (83,1%), followed by unknown (15,3%), sexual (1,4%) and vertical (0,2%). The distribution of genotypes was: 62,9% HCV-1 (29,2% HCV-1a and 31,9% HCV-1b), 3,4% HCV-2, 21,0% HCV-3, 12,6% HCV-4 and 0,1% HCV-5. CONCLUSION: The distribution of genotypes in Galicia shows significant differences with respect to that observed in Spain. This distribution varies with age, gender, coinfection with HIV and/or HBV, and within geographical areas.

Diagnóstico microbiológico de la infección por virus del papiloma humano / Microbiological diagnosis of human papilloma virus infection

La infección por el virus del papiloma humano (VPH) es la infección de transmisión sexual más frecuente en el mundo. Este virus ocasiona generalmente lesiones benignas, como verrugas genitales, pero también su persistencia ocasiona procesos malignos, como cáncer de cuello de útero (CCU) y, menos frecuentemente, anal, vaginal y de la cavidad orofaríngea. El CCU es una enfermedad muy severa, con alta mortalidad en muchos países. El cribado de CCU con citología ha tenido mucho éxito en estos últimos años, pero hay innumerable evidencia científica para que sea sustituida por la detección del VPH como prueba inicial. Para esto, hay en el mercado gran cantidad de técnicas, siendo aconsejable utilizar sistemas automáticos y pruebas aprobadas por la FDA. Un nuevo algoritmo basado en la detección individualizada de los genotipos 16 y 18 presentes en el 70% de los CCU ha sido propuesto por expertos y su implantación será inmediata en algunos países (AU)

Infection with human papillomavirus (HPV) is the leading cause of sexually transmitted infection worldwide. This virus generally causes benign lesions, such as genital warts, but persistent infection may lead to cervical cancer, anal cancer, vaginal cancer, and oropharyngeal cancer, although less frequently. Cervical cancer is a severe disease with a high mortality in some countries. Screening with cytology has been very successful in the last few years, but nowadays there are numerous studies that confirm that cytology should be replaced with the detection of HPV as a first line test in population based screening. There are several commercially available FDA approved tests for screening of cervical cancer. A new strategy, based on individual detection of the high risk genotypes HPV16 and HPV18, present in 70% of cervical cancer biopsies, has been proposed by some experts, and is going to be implemented in most countries in the future (AU)

Microbiological diagnosis of human papilloma virus infection. / Diagnóstico microbiológico de la infección por virus del papiloma humano.

Infection with human papillomavirus (HPV) is the leading cause of sexually transmitted infection worldwide. This virus generally causes benign lesions, such as genital warts, but persistent infection may lead to cervical cancer, anal cancer, vaginal cancer, and oropharyngeal cancer, although less frequently. Cervical cancer is a severe disease with a high mortality in some countries. Screening with cytology has been very successful in the last few years, but nowadays there are numerous studies that confirm that cytology should be replaced with the detection of HPV as a first line test in population based screening. There are several commercially available FDA approved tests for screening of cervical cancer. A new strategy, based on individual detection of the high risk genotypes HPV16 and HPV18, present in 70% of cervical cancer biopsies, has been proposed by some experts, and is going to be implemented in most countries in the future.

Sequence Analysis of In Vivo-Expressed HIV-1 Spliced RNAs Reveals the Usage of New and Unusual Splice Sites by Viruses of Different Subtypes.

HIV-1 RNAs are generated through a complex splicing mechanism, resulting in a great diversity of transcripts, which are classified in three major categories: unspliced, singly spliced (SS), and doubly spliced (DS). Knowledge on HIV-1 RNA splicing in vivo and by non-subtype B viruses is scarce. Here we analyze HIV-1 RNA splice site usage in CD4+CD25+ lymphocytes from HIV-1-infected individuals through pyrosequencing. HIV-1 DS and SS RNAs were amplified by RT-PCR in 19 and 12 samples, respectively. 13,108 sequences from HIV-1 spliced RNAs, derived from viruses of five subtypes (A, B, C, F, G), were identified. In four samples, three of non-B subtypes, five 3' splice sites (3'ss) mapping to unreported positions in the HIV-1 genome were identified. Two, designated A4i and A4j, were used in 22% and 25% of rev RNAs in two viruses of subtypes B and A, respectively. Given their close proximity (one or two nucleotides) to A4c and A4d, respectively, they could be viewed as variants of these sites. Three 3'ss, designated A7g, A7h, and A7i, located 20, 32, and 18 nucleotides downstream of A7, respectively, were identified in a subtype C (A7g, A7h) and a subtype G (A7i) viruses, each in around 2% of nef RNAs. The new splice sites or variants of splice sites were associated with the usual sequence features of 3'ss. Usage of unusual 3'ss A4d, A4e, A5a, A7a, and A7b was also detected. A4f, previously identified in two subtype C viruses, was preferentially used by rev RNAs of a subtype C virus. These results highlight the great diversity of in vivo splice site usage by HIV-1 RNAs. The fact that four of five newly identified splice sites or variants of splice sites were detected in non-subtype B viruses allows anticipating an even greater diversity of HIV-1 splice site usage than currently known.