Detection of High Level of Co-Infection and the Emergence of Novel SARS CoV-2 Delta-Omicron and Omicron-Omicron Recombinants in the Epidemiological Surveillance of Andalusia.

Recombination is an evolutionary strategy to quickly acquire new viral properties inherited from the parental lineages. The systematic survey of the SARS-CoV-2 genome sequences of the Andalusian genomic surveillance strategy has allowed the detection of an unexpectedly high number of co-infections, which constitute the ideal scenario for the emergence of new recombinants. Whole genome sequence of SARS-CoV-2 has been carried out as part of the genomic surveillance programme. Sample sources included the main hospitals in the Andalusia region. In addition to the increase of co-infections and known recombinants, three novel SARS-CoV-2 delta-omicron and omicron-omicron recombinant variants with two break points have been detected. Our observations document an epidemiological scenario in which co-infection and recombination are detected more frequently. Finally, we describe a family case in which co-infection is followed by the detection of a recombinant made from the two co-infecting variants. This increased number of recombinants raises the risk of emergence of recombinant variants with increased transmissibility and pathogenicity.

Macrolide and fluoroquinolone resistance of Mycoplasma genitalium in southern Spain, 2018-2019.

OBJECTIVES: In recent years, resistance in Mycoplasma genitalium (MG) to first-line (azithromycin) and second-line (moxifloxacin) treatment has been increasingly reported worldwide, however, no data regarding the south of Spain are available. METHODS: To determine resistance rates, MG-positive samples collected from June 2018 to June 2019 were analysed by sequencing the 23S rRNA and parC genes. RESULTS: A total of 77 patients (24 men having sex with men (MSM), 30 heterosexual men and 23 women) were included. Resistance-associated mutations against macrolide and fluoroquinolones were found in 36.4% (95% CI 25.7% to 48.1%) and 9.1% (95% CI 3.7% to 17.8%) of the patients, respectively. Being MSM and having had another STI in the last year were significantly associated with macrolide-resistant MG infection, while no associations were found with resistance to fluoroquinolones. CONCLUSIONS: Testing for resistance to first-line and second-line drugs against MG should be recommended for the general population and mandatory for the MSM population. We suggest that empiric azithromycin use for STI management should be avoided.

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.

Genetic diversity of HIV in seminal plasma remains higher than in blood after short-term antiretroviral therapy.

OBJECTIVE: To provide insight on viral kinetics and genetic diversity of HIV in seminal plasma at baseline and 1 month after initiating antiretroviral therapy (ART). PATIENTS AND METHODS: Blood and seminal samples from patients with newly diagnosed HIV were obtained before ART initiation (T0) and 1 month after ART initiation (T1). HIV env genetic diversity was studied using deep sequencing Nextera and V3 chemistry in a MiSeq Illumina platform. The number of viral quasispecies (5% cut-off) and Shannon Index were used to analyse diversity. RESULTS: Forty-seven ART-naive patients were recruited between September 2016 and November 2018. At enrolment, the number of quasispecies in blood (median 4 (IQR 2-5)) was lower than in the seminal compartment (median 6, (IQR 4-8)) (p<0.01); the Shannon Index was also higher (p<0.001) in the seminal compartment than in blood (1.77 vs 0.64). At T1, for the 13 patients with detectable HIV in both blood/seminal plasma, viral diversity remained higher (p=0.139) in seminal plasma (median 2 (IQR 1-4.5)) than in blood (median 1 (IQR 1-1.5)) Integrase inhibitors (INI)-based regimens achieved higher levels of undetectability and led more frequently to lower variability (p<0.001) than protease inhibitors (PI) or non-nucleoside reverse transcriptase inhibitors (NNRTI). CONCLUSION: We provide here further evidence of a larger genetic diversity in seminal plasma, both at diagnosis and short term after ART initiation. Our results strengthen previous findings on HIV diversity in seminal plasma. In addition, INIs decrease variability more rapidly than PI and NNRTI in both blood and seminal plasma.

High efficacy of resistance-guided retreatment of HCV patients failing NS5A inhibitors in the real world.

BACKGROUND & AIMS: Most hepatitis C virus (HCV)-infected patients failing NS5A inhibitors develop resistance-associated substitutions (RASs). Here we report the use of resistance-guided retreatment of patients who failed prior NS5A inhibitor-containing regimens in the GEHEP-004 cohort. This is the largest direct-acting antiviral (DAA)-resistance cohort study conducted in Spain. We aim to provide indications on how to use resistance information in settings where sofosbuvir/velpatasvir/voxilaprevir may not be available. METHODS: GEHEP-004 is a prospective multicenter cohort enrolling HCV-infected patients treated with interferon (IFN)-free DAA regimens. Prior to retreatment, population-based sequencing of HCV NS3, NS5A and NS5B genes was performed. After receiving a comprehensive resistance interpretation report, the retreatment regimen was chosen and the sustained virological response (SVR) at 12 weeks after treatment completion (SVR12) was recorded. RESULTS: A total of 342 patients experiencing virological failure after treatment with sofosbuvir/ledipasvir±ribavirin (54%), sofosbuvir/daclatasvir±ribavirin (23%), or paritaprevir-ritonavir/ombitasvir±dasabuvir±ribavirin (20%) were studied. After a resistance report, 186 patients were retreated. An SVR12 was achieved for 88.1% of the patients who failed after sofosbuvir/ledipasvir±ribavirin, 83.3% of the patients who failed after sofosbuvir/daclatasvir±ribavirin, 93.7% of the patients who failed after paritaprevir-ritonavir+ombitasvir±dasabuvir±ribavirin. CONCLUSIONS: In our study, we show how resistance-guided retreatment in conjunction with an interpreted report allows patients to achieve SVR rates close to 90%. We hypothesize that SVR rates may even be improved if resistance data are discussed between experienced virologists and treating clinicians. We believe that our data may be relevant for countries where the access to new DAA combination regimens is limited. LAY SUMMARY: Hepatitis C infection can be cured with currently available antiviral agents. Only a small proportion of patients experience treatment failure, however, in absolute numbers, a high number of patients may require retreatment. Highly effective combinations of antivirals are also available for retreatment. However, these antivirals might not be available in resource-limited settings. Herein, we show how, by analyzing the cause of resistance, retreatment efficacy with old drugs can get very close to the efficacy of new drug combinations.

Surveillance of transmitted drug resistance to integrase inhibitors in Spain: implications for clinical practice.

BACKGROUND: Integrase strand-transfer inhibitors (INSTIs) constitute at present one of the pillars of first-line ART. OBJECTIVES: To study the prevalence of and the trend in transmitted drug resistance (TDR) to INSTIs in ART-naive patients in Spain. METHODS: During the period 2012-17, 1109 patients from CoRIS were analysed. The Stanford algorithm v8.7 was used to evaluate TDR and transmission of clinically relevant resistance. To describe individual mutations/polymorphisms, the most recent IAS list (for INSTIs) and the 2009 WHO list update (for the backbone NRTIs used in combination with INSTIs in first-line treatment) were used. RESULTS: Clinically relevant resistance to the INSTI class was 0.2%: T66I, 0.1%, resistance to elvitegravir and intermediate resistance to raltegravir; and G163K, 0.1%, intermediate resistance to raltegravir and elvitegravir. No clinical resistance to dolutegravir or bictegravir was observed. The prevalence of INSTI TDR following the IAS-USA INSTI mutation list was 2.6%, with no trend towards changes in the prevalence throughout the study period. The overall prevalence of NRTI WHO mutations was 4.3%, whereas clinically relevant resistance to tenofovir, abacavir and emtricitabine/lamivudine was 1.7%, 1.9% and 0.7%, respectively. CONCLUSIONS: Given the low prevalence of clinically relevant resistance to INSTIs and first-line NRTIs in Spain, it is very unlikely that a newly diagnosed patient will present with clinical resistance to a first-line INSTI-based regimen. These patients may not benefit from INSTI and NRTI baseline resistance testing.

Association of breast and gut microbiota dysbiosis and the risk of breast cancer: a case-control clinical study.

BACKGROUND: Breast cancer ranks first in women, and is the second cause of death in this gender. In addition to genetics, the environment contributes to the development of the disease, although the factors involved are not well known. Among the latter is the influence of microorganisms and, therefore, attention is recently being paid to the mammary microbiota. We hypothesize that the risk of breast cancer could be associated with the composition and functionality of the mammary/gut microbiota, and that exposure to environmental contaminants (endocrine disruptors, EDCs) might contribute to alter these microbiota. METHODS: We describe a case-control clinical study that will be performed in women between 25 and 70 years of age. Cases will be women diagnosed and surgically intervened of breast cancer (stages I and II). Women with antecedents of cancer or advanced tumor stage (metastasis), or who have received antibiotic treatment within a period of 3 months prior to recruitment, or any neoadjuvant therapy, will be excluded. Controls will be women surgically intervened of breast augmentation or reduction. Women with oncological, gynecological or endocrine history, and those who have received antibiotic treatment within a period of 3 months prior to recruitment will also be excluded. Blood, urine, breast tissue and stool samples will be collected. Data regarding anthropometric, sociodemographic, reproductive history, tumor features and dietary habits will be gathered. Metabolomic studies will be carried out in stool and breast tissue samples. Metagenomic studies will also be performed in stool and breast tissue samples to ascertain the viral, fungal, bacterial and archaea populations of the microbiota. Quantitation of estrogens, estrogen metabolites and EDCs in samples of serum, urine and breast tissue will also be performed. DISCUSSION: This is the first time that the contribution of bacteria, archaea, viruses and fungi together with their alteration by environmental contaminants to the risk of breast cancer will be evaluated in the same study. Results obtained could contribute to elucidate risk factors, improve the prognosis, as well as to propose novel intervention studies in this disease. TRIAL REGISTRATION: ClinicalTrials.gov NCT03885648 , 03/25/2019. Retrospectively registered.

Increasing importance of European lineages in seeding the hepatitis C virus subtype 1a epidemic in Spain.

BackgroundReducing the burden of the hepatitis C virus (HCV) requires large-scale deployment of intervention programmes, which can be informed by the dynamic pattern of HCV spread. In Spain, ongoing transmission of HCV is mostly fuelled by people who inject drugs (PWID) infected with subtype 1a (HCV1a).AimOur aim was to map how infections spread within and between populations, which could help formulate more effective intervention programmes to halt the HCV1a epidemic in Spain.MethodsEpidemiological links between HCV1a viruses from a convenience sample of 283 patients in Spain, mostly PWID, collected between 2014 and 2016, and 1,317, 1,291 and 1,009 samples collected abroad between 1989 and 2016 were reconstructed using sequences covering the NS3, NS5A and NS5B genes. To efficiently do so, fast maximum likelihood-based tree estimation was coupled to a flexible Bayesian discrete phylogeographic inference method.ResultsThe transmission network structure of the Spanish HCV1a epidemic was shaped by continuous seeding of HCV1a into Spain, almost exclusively from North America and European countries. The latter became increasingly relevant and have dominated in recent times. Export from Spain to other countries in Europe was also strongly supported, although Spain was a net sink for European HCV1a lineages. Spatial reconstructions showed that the epidemic in Spain is diffuse, without large, dominant within-country networks.ConclusionTo boost the effectiveness of local intervention efforts, concerted supra-national strategies to control HCV1a transmission are needed, with a strong focus on the most important drivers of ongoing transmission, i.e. PWID and other high-risk populations.

Microenvironment Eradication of Hepatitis C: A Novel Treatment Paradigm.

OBJECTIVES: Prisons are major reservoirs of hepatitis C virus (HCV) in which a therapeutic approach has been particularly difficult so far. Our aim was to create a permanent program of HCV elimination in a prison based on a "test and treat" strategy. METHODS: This open-label clinical trial was conducted in the Spanish prison "El Dueso" between May 2016 and July 2017. Viremic patients were treated with a ledipasvir-sofosbuvir regimen (8-12 weeks) according to the 2015 Spanish Guidelines. A teleconsultation program was established to follow-up patients from the hospital. Non-responders were submitted for a phylogenetic analysis and offered retreatment. An evaluation of new cases of HCV infection was performed every 6 months and upon release in all inmates. RESULTS: 847 (99.5%) inmates accepted to participate. HCV antibodies were present in 110 (13.0%) and 86 (10.2%) had detectable viremia. Most of them were genotype 1 or 3 (82.6%) and had

Obesity and Asthma: A Missing Link.

Obesity and asthma are two chronic conditions that affect millions of people. Genetic and lifestyle factors such as diet, physical activity, and early exposure to micro-organisms are important factors that may contribute to the escalating prevalence of both conditions. The prevalence of asthma is higher in obese individuals. Recently, two major phenotypes of asthma with obesity have been described: one phenotype of early-onset asthma that is aggravated by obesity, and a second phenotype of later-onset asthma that predominantly affects women. Systemic inflammation and mechanical effect, both due to the expansion of the adipose tissue, have been proposed as the main reasons for the association between obesity and asthma. However, the mechanisms involved are not yet fully understood. Moreover, it has also been suggested that insulin resistance syndrome can have a role in the association between these conditions. The intestinal microbiota is an important factor in the development of the immune system, and can be considered a link between obesity and asthma. In the obese state, higher lipopolysaccharide (LPS) serum levels as a consequence of a microbiota dysbiosis have been found. In addition, changes in microbiota composition result in a modification of carbohydrate fermentation capacity, therefore modifying short chain fatty acid (SCFA) levels. The main objective of this review is to summarize the principal findings that link obesity and asthma.

Usefulness of Integrase resistance testing in proviral HIV-1 DNA in patients with Raltegravir prior failure.

BACKGROUND: In our study, we have hypothesized that proviral DNA may show the history of mutations that emerged at previous failures to a Raltegravir containing regimen, in patients who are currently undetectable and candidates to simplification to a Dolutegravir containing regimen, in order to decide on once a day or twice a day dosing. METHODS: We have performed a pilot, observational, retrospective, non interventional study, including 7 patients infected by HIV-1, all with a history of previous failure to a RAL containing regimen, that were successfully salvaged and had reached viral suppression. A genotypic viral Integrase region study was available for each patient at the moment of RAL failure. After an average (IQR) time of 48 months (29-53) Integrase resistance mutations in proviral DNA were studied. RESULTS: All the patients were infected by HIV-1 B subtypes, with a mean age of 55 (range 43 to 56), originating from Spain, and 4 were women. Median viral load (log) and CD4 count at the moment of the study on proviral DNA was of 1.3 log cp/ml (range 0-1.47) and 765.5 cells/µL (range; 436.75-1023.75). The median time (IQR) between previous failure to RAL and the study on proviral DNA was 48 (29-53) months. At Raltegravir failure, N155H was detected in four patients, and other secondary mutations were detected in five patients (71.4 %). In proviral DNA, N155H was detected by population sequencing in three patients (42.8 %), and UDS demonstrated a 9.77 % relative abundance of N155H in the remaining patient. Sanger sequencing correctly identified all the secondary mutations. CONCLUSION: This is a pilot study that demonstrates the possibility of properly identifying N155H and some secondary mutations 29-53 months after failure.

Improved prediction of salvage antiretroviral therapy outcomes using ultrasensitive HIV-1 drug resistance testing.

BACKGROUND: The clinical relevance of ultrasensitive human immunodeficiency virus type 1 (HIV-1) genotypic resistance testing in antiretroviral treatment (ART)-experienced individuals remains unknown. METHODS: This was a retrospective, multicentre, cohort study in ART-experienced, HIV-1-infected adults who initiated salvage ART including, at least 1 ritonavir-boosted protease inhibitor, raltegravir or etravirine. Presalvage ART Sanger and 454 sequencing of plasma HIV-1 were used to generate separate genotypic sensitivity scores (GSS) using the HIVdb, ANRS, and REGA algorithms. Virological failure (VF) was defined as 2 consecutive HIV-1 RNA levels ≥200 copies/mL at least 12 weeks after salvage ART initiation, whereas subjects remained on the same ART. The ability of Sanger and 454-GSS to predict VF was assessed by receiver operating characteristic (ROC) curves and survival analyses. RESULTS: The study included 132 evaluable subjects; 28 (21%) developed VF. Using HIVdb, 454 predicted VF better than Sanger sequencing in the ROC curve analysis (area under the curve: 0.69 vs 0.60, Delong test P = .029). Time to VF was shorter for subjects with 454-GSS < 3 vs 454-GSS ≥ 3 (Log-rank P = .003) but not significantly different between Sanger-GSS < 3 and ≥3. Factors independently associated with increased risk of VF in multivariate Cox regression were a 454-GSS < 3 (HR = 4.6, 95 CI, [1.5, 14.0], P = .007), and the number of previous antiretrovirals received (HR = 1.2 per additional drug, 95 CI, [1.1, 1.3], P = .001). Equivalent findings were obtained with the ANRS and REGA algorithms. CONCLUSIONS: Ultrasensitive HIV-1 genotyping improves GSS-based predictions of virological outcomes of salvage ART relative to Sanger sequencing. This may improve the clinical management of ART-experienced subjects living with HIV-1. CLINICAL TRIALS REGISTRATION: NCT01346878.

Molecular and phylogenetic characterization of the monkeypox outbreak in the South of Spain.

Background and Aim: Until the May 2022 Monkeypox (MPXV) outbreak, which spread rapidly to many non-endemic countries, the virus was considered a viral zoonosis limited to some African countries. The Andalusian circuit of genomic surveillance was rapidly applied to characterize the MPXV outbreak in the South of Spain. Methods: Whole genome sequencing was used to obtain the genomic profiles of samples collected across the south of Spain, representative of all the provinces of Andalusia. Phylogenetic analysis was used to study the relationship of the isolates and the available sequences of the 2022 outbreak. Results: Whole genome sequencing of a total of 160 MPXV viruses from the different provinces that reported cases were obtained. Interestingly, we report the sequences of MPXV viruses obtained from two patients who died. While one of the isolates bore no noteworthy mutations that explain a potential heightened virulence, in another patient the second consecutive genome sequence, performed after the administration of tecovirimat, uncovered a mutation within the A0A7H0DN30 gene, known to be a prime target for tecovirimat in its Vaccinia counterpart. In general, a low number of mutations were observed in the sequences reported, which were very similar to the reference of the 2022 outbreak (OX044336), as expected from a DNA virus. The samples likely correspond to several introductions of the circulating MPXV viruses from the last outbreak. The virus sequenced from one of the two patients that died presented a mutation in a gene that bears potential connections to drug resistance. This mutation was absent in the initial sequencing before treatment.

Gut Microbiota Composition Can Predict Colonization by Multidrug-Resistant Bacteria in SARS-CoV-2 Patients in Intensive Care Unit: A Pilot Study.

The SARS-CoV-2 infection has increased the number of patients entering Intensive Care Unit (ICU) facilities and antibiotic treatments. Concurrently, the multi-drug resistant bacteria (MDRB) colonization index has risen. Considering that most of these bacteria are derived from gut microbiota, the study of its composition is essential. Additionally, SARS-CoV-2 infection may promote gut dysbiosis, suggesting an effect on microbiota composition. This pilot study aims to determine bacteria biomarkers to predict MDRB colonization risk in SARS-CoV-2 patients in ICUs. Seventeen adult patients with an ICU stay >48 h and who tested positive for SARS-CoV-2 infection were enrolled in this study. Patients were assigned to two groups according to routine MDRB colonization surveillance: non-colonized and colonized. Stool samples were collected when entering ICUs, and microbiota composition was determined through Next Generation Sequencing techniques. Gut microbiota from colonized patients presented significantly lower bacterial diversity compared with non-colonized patients (p < 0.05). Microbiota in colonized subjects showed higher abundance of Anaerococcus, Dialister and Peptoniphilus, while higher levels of Enterococcus, Ochrobactrum and Staphylococcus were found in non-colonized ones. Moreover, LEfSe analysis suggests an initial detection of Dialister propionicifaciens as a biomarker of MDRB colonization risk. This pilot study shows that gut microbiota profile can become a predictor biomarker for MDRB colonization in SARS-CoV-2 patients.

Molecular characterization of patients with chronic hepatitis C virus infection in Jordan: implications on response to direct-acting antiviral therapy.

OBJECTIVES: To investigate the molecular characteristics of Hepatitis C Virus (HCV) detected in patients with chronic HCV infection in Jordan. METHODS: The study included 48 Jordanian treatment-naïve patients with active chronic HCV recruited from seven governorates. HCV genotype and the resistance-associated substitutions (RAS) profile were investigated by next-generation sequencing of the NS5B, NS5A, and NS3 regions of HCV. RESULTS: "Unusual genotype 4 subtypes" were detected in four (8.3%) patients (4n-n = 1, 4o-n = 2, 4v-n = 1); one patient (2.1%) was co-infected by genotypes 1b+4a. Overall prevalence of NS5A RASs was 38.3% (3% cutoff); genotype 4a showed the highest NS5A RAS prevalence (n = 11, 55.0%). Overall prevalence of NS3 RASs was 21.8% (7/32), all genotype 1a-infected patients. CONCLUSIONS: We report, for the first time in Jordanian patients with chronic HCV infection, the detection of unusual genotype 4 subtypes 4n, 4o, and 4v. Baseline RASs in NS5A are frequent, with complex RASs patterns in some of the unusual subtypes. Our data support the need for sequencing surveillance programs in sub-Saharan Africa, Asia, and the Middle East and North African region to monitor response to treatment in these subtypes and to facilitate the World Health Organization's 2030 elimination strategy.

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.

[Laboratory diagnosis of HIV infection, viral tropism and resistance to antiretrovirals]. / Diagnóstico de laboratorio de la infección por el VIH, del tropismo viral y de las resistencias a los antirretrovirales.

The accurate diagnosis of HIV infection demands that to consider a positive result, at least three assays with different antigenic base should be used, one of them, Western-Blot being mandatory for confirmation. Fourth generation ELISAs shorten the window phase to 13-15 days, as they now include p24 antigen detection. Proviral DNA or Viral RNA detection by molecular methods have proved useful for addressing complex situations in which serology was inconclusive. Viral load (HIV-RNA) is routinely used to follow-up HIV infected patients and is used for treatment initiation decisions. It is also used to monitor viral failure. When this happens, resistance tests are needed to guide treatment changes. Resistance is also used to assess the transmission of drug resistance to newly diagnosed patients. Finally, before using an anti-CCR5 drug, viral tropism needs to be determined. This can be done using genotypic tests, widely available in many HIV labs, or phenotypic tests, only available at certain sites.

[Virological response to adefovir dipivoxil predicts the long-term development of resistance in previously untreated patients with HBeAg-negative chronic hepatitis B]. / La respuesta virológica al tratamiento con adefovir dipivoxil predice el desarrollo de resistencias a largo plazo en pacientes con hepatitis crónica B HBeAg negativo previamente no tratados.

BACKGROUND AND OBJECTIVE: Adefovir dipivoxil monotherapy in lamivudine-resistant patients is associated with more frequent development of resistance than in naïve patients. The virological response during treatment predicts the risk of developing resistance. The aims of this study were to assess the efficacy of adefovir dipivoxil treatment in naïve and lamivudine-resistant patients and to determine whether virological response predicts the development of adefovir resistance. PATIENTS AND METHOD: This study included 82 patients with HBeAg-negative chronic hepatitis B (CHB) who received adefovir dipivoxil therapy. During active treatment, HBV-DNA values were determined by polymerase chain reaction; in addition, the presence of adefovir resistance-associated mutations was studied in cases of virological breakthrough. RESULTS: Virological response at 12 and 24 months was 59% and 73% in naive patients compared with 40% and 67% in lamivudine-resistant patients, whereas virological breakthrough at 24 months was 9.5% in naïve patients compared with 20% in lamivudine-resistant patients. A small percentage (4%) of patients with virological response at 12 months showed virological breakthrough between 12 and 40 months versus 29.4% of patients without virological response (P=.03). In lamivudine-resistant patients, virological response at 12 months was not a predictive factor for the development of virological breakthrough. CONCLUSIONS: Adefovir dipivoxil monotherapy in lamivudine-resistant patients is associated with an increased tendency to develop virological breakthrough, which cannot be predicted by virological response at 12 months of treatment. In naive patients, an undetectable viral load at 12 months of treatment ensures the absence of virological breakthrough at 40 months of treatment.

Highly accurate whole-genome imputation of SARS-CoV-2 from partial or low-quality sequences.

BACKGROUND: The current SARS-CoV-2 pandemic has emphasized the utility of viral whole-genome sequencing in the surveillance and control of the pathogen. An unprecedented ongoing global initiative is producing hundreds of thousands of sequences worldwide. However, the complex circumstances in which viruses are sequenced, along with the demand of urgent results, causes a high rate of incomplete and, therefore, useless sequences. Viral sequences evolve in the context of a complex phylogeny and different positions along the genome are in linkage disequilibrium. Therefore, an imputation method would be able to predict missing positions from the available sequencing data. RESULTS: We have developed the impuSARS application, which takes advantage of the enormous number of SARS-CoV-2 genomes available, using a reference panel containing 239,301 sequences, to produce missing data imputation in viral genomes. ImpuSARS was tested in a wide range of conditions (continuous fragments, amplicons or sparse individual positions missing), showing great fidelity when reconstructing the original sequences, recovering the lineage with a 100% precision for almost all the lineages, even in very poorly covered genomes (<20%). CONCLUSIONS: Imputation can improve the pace of SARS-CoV-2 sequencing production by recovering many incomplete or low-quality sequences that would be otherwise discarded. ImpuSARS can be incorporated in any primary data processing pipeline for SARS-CoV-2 whole-genome sequencing.