RESUMO
Dual co-infection with both HSV-1 and HSV-2 is rare, with few cases reported in the literature. In this case report, we describe the successful use of unbiased metagenomic next-generation sequencing (mNGS) as a rapid and alternative method for confirming dual genital herpes co-infection. Our case involves a 74-year-old woman who presented with genital lesions and initially tested positive for both HSV-1 and HSV-2 via the Luminex ARIES HSV 1&2 assay. The entire mNGS process, from nucleic acid extraction to result analysis, was completed in less than 48 h. Using mNGS, we identified mapped reads specific to either HSV-1 or HSV-2 and screened the sequences to rule out mis-genotyping by the Luminex ARIES assay. Notably, the generated sequences can reveal sequence variations within multiple gene regions, demonstrating the potential of mNGS for identifying novel HSV-1 and HSV-2 variants. Our findings suggest that mNGS can serve as a rapid and reliable alternative confirmatory method for dual genital herpes infections, providing valuable information to guide appropriate treatment options for patients. By eliminating the need for prior knowledge of causative agents, mNGS offers an unbiased approach for detecting and characterizing viral co-infections.
RESUMO
The HIV genotypic resistance test (GRT) is a standard of care for the clinical management of HIV/AIDS patients. In recent decades, population or Sanger sequencing has been the foundation for drug resistance monitoring in clinical settings. However, the advent of high-throughput or next-generation sequencing has caused a paradigm shift towards the detection and characterization of low-abundance covert mutations that would otherwise be missed by population sequencing. This is clinically significant, as these mutations can potentially compromise the efficacy of antiretroviral therapy, causing poor virologic suppression. Therefore, it is important to develop a more sensitive method so as to reliably detect clinically actionable drug-resistant mutations (DRMs). Here, we evaluated the diagnostic performance of a laboratory-developed, high-throughput, sequencing-based GRT using 103 archived clinical samples that were previously tested for drug resistance using population sequencing. As expected, high-throughput sequencing found all the DRMs that were detectable by population sequencing. Significantly, 78 additional DRMs were identified only by high-throughput sequencing, which is statistically significant based on McNemar's test. Overall, our results complement previous studies, supporting the notion that the two methods are well correlated, and the high-throughput sequencing method appears to be an excellent alternative for drug resistance testing in a clinical setting.
Assuntos
Fármacos Anti-HIV , Infecções por HIV , Soropositividade para HIV , HIV-1 , Humanos , HIV-1/genética , Farmacorresistência Viral/genética , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Genótipo , Mutação , Fármacos Anti-HIV/farmacologia , Fármacos Anti-HIV/uso terapêuticoRESUMO
We compared the performance of five assays for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) detection on nasopharyngeal swab samples: Roche "cobas," Luminex "ARIES," MiRXES "Fortitude," Altona "RealStar," and Thermo Fisher Scientific "TaqPath." A total of 94 nasopharyngeal swab samples were obtained from 80 confirmed coronavirus disease 2019 cases in the first 2 weeks of illness (median, 7 days; range, 2-14 days) and 14 healthy controls. After collection, all samples were transported to the hospital clinical laboratory within 24 h. These samples were tested on all five assays within 3 days of sample receipt. Of the 94 samples, 69 yielded the same result on all platforms, resulting in an agreement of 73.4% (69 of 94). Of these, 14 were the healthy control swabs which all tested negative, demonstrating good specificity across all platforms. The ARIES assay had the lowest detection rate (68.8%), followed by Fortitude (85.0%), RealStar (86.3%), cobas (95.0%), and TaqPath (100%). Statistically significant differences were observed for ARIES, Fortitude, and RealStar when compared against the best performing TaqPath using McNemar's χ2 test. A consensus result was established based on the results obtained by the cobas, Fortitude, RealStar, and TaqPath. Six discrepancies had failed to reach a consensus and were adjudicated using the Cepheid Xpert Xpress SARS-CoV-2. Overall, the TaqPath and cobas assays were the most sensitive at detecting their designated SARS-CoV-2 gene targets. On the other hand, the ARIES assay was the least sensitive, thus warranting the need for assay re-optimization before go-live at the testing laboratory.
Assuntos
Teste de Ácido Nucleico para COVID-19 , COVID-19/diagnóstico , SARS-CoV-2/isolamento & purificação , Genes Virais/genética , Humanos , Nasofaringe/virologia , RNA Viral/genética , SARS-CoV-2/genética , Sensibilidade e EspecificidadeRESUMO
JAK2 (V617F) allelic burden is the main genetic driver behind and a potential differentiator between individual myeloproliferative neoplasm (MPN) subtypes. This study aimed to explore the relationship between JAK2 (V617F) allelic burden, MPN subtypes and their clinico-haematological manifestations in a Singapore-based cohort. Analysis was performed on a retrospectively collected dataset of 128 patients diagnosed with JAK2 (V617F) positive Philadelphia-negative MPNs between 2016 to 2017 in Singapore. Genomic analysis was conducted on blood samples via DNA extraction and Droplet Digital Polymerase Chain Reaction (ddPCR). The mean age was 62.4 (SD=14.1). 85 out of the 128 (66.4%) patients were male. There was a statistically significant difference in allelic burdens between the different MPN disease subtypes χ2(3) = 9.064, p=0.028, with essential thrombocytosis (ET) patients having the lowest mean JAK2 percentage allelic burden (26.5%). Patients with an allelic burden >50% had higher leukocyte counts (MWU 1016.5, p=0.001), haemoglobin levels (MWU 1287.0, p=0.045), lactate dehydrogenase levels (MWU 611.5, p=0.001), and lower platelet levels (MWU 1164.0, p=0.008). Subgroup analysis revealed none of these correlations was significant in the ET subgroup. The results are largely in concordance with previous research in Asian cohorts demonstrating the association between allelic burden and clinico-haematological manifestations of MPN. However, in the ET subgroup, the JAK2 (V617F) allelic burden do not correlate positively for haematological parameters which is only seen in Asian patients.
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Assuntos
Povo Asiático/genética , Janus Quinase 2/genética , Mutação , Transtornos Mieloproliferativos/patologia , Cromossomo Filadélfia , Idoso , Alelos , Feminino , Seguimentos , Humanos , Masculino , Pessoa de Meia-Idade , Transtornos Mieloproliferativos/epidemiologia , Transtornos Mieloproliferativos/genética , Prognóstico , Estudos Retrospectivos , Singapura/epidemiologiaAssuntos
Teste de Ácido Nucleico para COVID-19/métodos , COVID-19/diagnóstico , Nasofaringe/virologia , Manejo de Espécimes/métodos , Adulto , Proteínas do Nucleocapsídeo de Coronavírus/análise , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Fosfoproteínas/análise , Poliproteínas/análise , Reação em Cadeia da Polimerase em Tempo Real , SARS-CoV-2/isolamento & purificação , Proteínas Virais/análiseAssuntos
Janus Quinase 2/genética , Mutação , Transtornos Mieloproliferativos/genética , Trombocitose/genética , Clopidogrel/uso terapêutico , Feminino , Humanos , Janus Quinase 2/metabolismo , Pessoa de Meia-Idade , Transtornos Mieloproliferativos/diagnóstico , Transtornos Mieloproliferativos/tratamento farmacológico , Inibidores da Agregação Plaquetária/uso terapêuticoRESUMO
BACKGROUND: Human herpes simplex virus type 1 (HSV-1) and type 2 (HSV-2) are responsible for a plethora of human diseases, of which cutaneous and mucocutaneous infections are the most prevalent. In its most severe form, HSV infection can cause meningitis/encephalitis. We compared the Luminex ARIES HSV 1&2 assay (Luminex Corp., Austin, TX, USA), an automated sample-to-result molecular solution, to two non-automated HSV DNA assays. METHODS: A total of 116 artificial controls were used to determine the analytical performance of the ARIES assay. Controls were prepared by spiking universal transport medium (UTM) and cerebrospinal fluid (CSF) samples from patients who tested negative for HSV by an in-house HSV-1 and -2 DNA assay with reference materials (SeraCare Life Sciences, MA, USA; ZeptoMetrix Corp., MA, USA). Another 117 clinical samples were then used to compare the clinical performance of the ARIES assay with those of an in-house assay and the FTD Neuro 9 assay (Fast Track Diagnostics, Junglinster, Luxembourg). RESULTS: The analytical sensitivity (95% limit of detection) of the ARIES assay was 318 copies/mL (UTM samples) and 935 copies/mL (CSF samples) for HSV-1 strain 96 and 253 copies/mL (UTM samples) and 821 copies/mL (CSF samples) for HSV-2 strain 09. No cross-reactivity was observed in samples spiked with 14 non-HSV microorganisms. Compared with the reference result (agreement between the in-house and FTD Neuro 9 results), the ARIES assay had overall concordance rates of 98.2% (111/113) and 100% (113/113) for HSV-1 and HSV-2, respectively. CONCLUSIONS: The ARIES assay appears to be an excellent alternative for rapid detection and differentiation of HSV in skin and genital infections, meningitis, and encephalitis.