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1.
Gigascience ; 132024 Jan 02.
Artigo em Inglês | MEDLINE | ID: mdl-38832467

RESUMO

BACKGROUND: Modern sequencing technologies offer extraordinary opportunities for virus discovery and virome analysis. Annotation of viral sequences from metagenomic data requires a complex series of steps to ensure accurate annotation of individual reads and assembled contigs. In addition, varying study designs will require project-specific statistical analyses. FINDINGS: Here we introduce Hecatomb, a bioinformatic platform coordinating commonly used tasks required for virome analysis. Hecatomb means "a great sacrifice." In this setting, Hecatomb is "sacrificing" false-positive viral annotations using extensive quality control and tiered-database searches. Hecatomb processes metagenomic data obtained from both short- and long-read sequencing technologies, providing annotations to individual sequences and assembled contigs. Results are provided in commonly used data formats useful for downstream analysis. Here we demonstrate the functionality of Hecatomb through the reanalysis of a primate enteric and a novel coral reef virome. CONCLUSION: Hecatomb provides an integrated platform to manage many commonly used steps for virome characterization, including rigorous quality control, host removal, and both read- and contig-based analysis. Each step is managed using the Snakemake workflow manager with dependency management using Conda. Hecatomb outputs several tables properly formatted for immediate use within popular data analysis and visualization tools, enabling effective data interpretation for a variety of study designs. Hecatomb is hosted on GitHub (github.com/shandley/hecatomb) and is available for installation from Bioconda and PyPI.


Assuntos
Metagenômica , Software , Metagenômica/métodos , Viroma/genética , Vírus/genética , Vírus/classificação , Animais , Biologia Computacional/métodos , Genoma Viral , Metagenoma
2.
J Clin Virol ; 173: 105695, 2024 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-38823290

RESUMO

Metagenomics is gradually being implemented for diagnosing infectious diseases. However, in-depth protocol comparisons for viral detection have been limited to individual sets of experimental workflows and laboratories. In this study, we present a benchmark of metagenomics protocols used in clinical diagnostic laboratories initiated by the European Society for Clinical Virology (ESCV) Network on NGS (ENNGS). A mock viral reference panel was designed to mimic low biomass clinical specimens. The panel was used to assess the performance of twelve metagenomic wet lab protocols currently in use in the diagnostic laboratories of participating ENNGS member institutions. Both Illumina and Nanopore, shotgun and targeted capture probe protocols were included. Performance metrics sensitivity, specificity, and quantitative potential were assessed using a central bioinformatics pipeline. Overall, viral pathogens with loads down to 104 copies/ml (corresponding to CT values of 31 in our PCR assays) were detected by all the evaluated metagenomic wet lab protocols. In contrast, lower abundant mixed viruses of CT values of 35 and higher were detected only by a minority of the protocols. Considering the reference panel as the gold standard, optimal thresholds to define a positive result were determined per protocol, based on the horizontal genome coverage. Implementing these thresholds, sensitivity and specificity of the protocols ranged from 67 to 100 % and 87 to 100 %, respectively. A variety of metagenomic protocols are currently in use in clinical diagnostic laboratories. Detection of low abundant viral pathogens and mixed infections remains a challenge, implying the need for standardization of metagenomic analysis for use in clinical settings.


Assuntos
Benchmarking , Metagenômica , Sensibilidade e Especificidade , Vírus , Metagenômica/métodos , Metagenômica/normas , Humanos , Vírus/genética , Vírus/classificação , Vírus/isolamento & purificação , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Sequenciamento de Nucleotídeos em Larga Escala/normas , Viroses/diagnóstico , Viroses/virologia , Biologia Computacional/métodos
3.
Nat Commun ; 15(1): 4888, 2024 Jun 07.
Artigo em Inglês | MEDLINE | ID: mdl-38849332

RESUMO

Chloroxylenol is a worldwide commonly used disinfectant. The massive consumption and relatively high chemical stability of chloroxylenol have caused eco-toxicological threats in receiving waters. We noticed that chloroxylenol has a chemical structure similar to numerous halo-phenolic disinfection byproducts. Solar detoxification of some halo-phenolic disinfection byproducts intrigued us to select a rapidly degradable chloroxylenol alternative from them. In investigating antimicrobial activities of disinfection byproducts, we found that 2,6-dichlorobenzoquinone was 9.0-22 times more efficient than chloroxylenol in inactivating the tested bacteria, fungi and viruses. Also, the developmental toxicity of 2,6-dichlorobenzoquinone to marine polychaete embryos decreased rapidly due to its rapid degradation via hydrolysis in receiving seawater, even without sunlight. Our work shows that 2,6-dichlorobenzoquinone is a promising disinfectant that well addresses human biosecurity and environmental sustainability. More importantly, our work may enlighten scientists to exploit the slightly alkaline nature of seawater and develop other industrial products that can degrade rapidly via hydrolysis in seawater.


Assuntos
Desinfetantes , Desinfecção , Água do Mar , Desinfetantes/química , Desinfetantes/farmacologia , Desinfecção/métodos , Água do Mar/química , Animais , Hidrólise , Poliquetos/efeitos dos fármacos , Fungos/efeitos dos fármacos , Bactérias/efeitos dos fármacos , Clorofenóis/química , Vírus/efeitos dos fármacos , Humanos , Xilenos
4.
BMC Infect Dis ; 24(1): 569, 2024 Jun 07.
Artigo em Inglês | MEDLINE | ID: mdl-38849747

RESUMO

BACKGROUND: Flies are acknowledged as vectors of diseases transmitted through mechanical means and represent a significant risk to human health. The study aimed to determine the prevalence of enteropathogens carried by flies in Pudong New Area to inform strategies for preventing and controlling flies. METHODS: Samples were collected from various locations in the area using cage trapping techniques between April and November 2021, encompassing various habitats such as parks, residential areas, restaurants, and farmers' markets. The main fly species were identified using cryomicrography and taxonomic enumeration, with 20 samples per tube collected from different habitats. Twenty-five enteropathogens were screened using GI_Trial v3 TaqManTM microbial arrays. RESULTS: A total of 3,875 flies were collected from 6,400 placements, resulting in an average fly density of 0.61 flies per cage. M. domestica were the most common species at 39.85%, followed by L. sericata at 16.57% and B. peregrina at 13.14%. Out of 189 samples, 93 tested positive for enteropathogens, with nine different pathogens being found. 12.70% of samples exclusively had parasites, a higher percentage than those with only bacteria or viruses. The study found that M. domestica had fewer enteropathogens than L. sericata and B. peregrina, which primarily harbored B. hominis instead of bacteria and viruses such as E. coli, Astrovirus, and Sapovirus. During spring testing, all three fly species exhibited low rates of detecting enteropathogens. M. domestica were found in residential areas with the highest number of pathogen species, totaling six. In contrast, L. sericata and B. peregrina were identified in farmers' markets with the highest number of pathogen species, totaling six and seven, respectively. CONCLUSIONS: Flies have the potential to serve as vectors for the transmission of enteropathogens, thereby posing a substantial risk to public health.


Assuntos
Insetos Vetores , Animais , Humanos , Insetos Vetores/microbiologia , Bactérias/isolamento & purificação , Bactérias/classificação , Bactérias/genética , China/epidemiologia , Dípteros/microbiologia , Vírus/isolamento & purificação , Vírus/classificação , Vírus/genética , Muscidae/microbiologia
5.
Curr Protoc ; 4(6): e1056, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38856995

RESUMO

Sequence changes in viral genomes generate protein sequence diversity that enables viruses to evade the host immune system, hindering the development of effective preventive and therapeutic interventions. The massive proliferation of sequence data provides unprecedented opportunities to study viral adaptation and evolution. An alignment-free approach removes various restrictions posed by an alignment-dependent approach for studying sequence diversity. The publicly available tool, UNIQmin, offers an alignment-free approach for studying viral sequence diversity at any given rank of taxonomy lineage and is big data ready. The tool performs an exhaustive search to determine the minimal set of sequences required to capture the peptidome diversity within a given dataset. This compression is possible through the removal of identical sequences and unique sequences that do not contribute effectively to the peptidome diversity pool. Herein, we describe a detailed four-part protocol utilizing UNIQmin to generate the minimal set for the purpose of viral diversity analyses, alignment-free at any rank of the taxonomy lineage, using the recent global public health threat Monkeypox virus (MPX) sequence data as a case study. The protocol enables a systematic bioinformatics approach to study sequence diversity across taxonomic lineages, which is crucial for our future preparedness against viral epidemics. This is particularly important when data are abundant, freely available, and alignment is not an option. © 2024 Wiley Periodicals LLC. Basic Protocol 1: Tool installation and input file preparation Basic Protocol 2: Generation of a minimal set of sequences for a given dataset Basic Protocol 3: Comparative minimal set analysis across taxonomic lineage ranks Basic Protocol 4: Factors affecting the minimal set of sequences.


Assuntos
Biologia Computacional , Biologia Computacional/métodos , Proteínas Virais/genética , Genoma Viral/genética , Software , Vírus/genética , Vírus/isolamento & purificação , Vírus/classificação , Peptídeos/química
6.
BMC Ecol Evol ; 24(1): 81, 2024 Jun 13.
Artigo em Inglês | MEDLINE | ID: mdl-38872095

RESUMO

BACKGROUND: New Zealand is home to over 120 native endemic species of skinks and geckos that radiated over the last 20-40 million years, likely driven by the exploitation of diverse habitats formed during the Miocene. The recent radiation of animal hosts may facilitate cross-species virus transmission, likely reflecting their close genetic relationships and therefore relatively low barriers for viruses to emerge in new hosts. Conversely, as animal hosts adapt to new niches, even within specific geographic locations, so too could their viruses. Consequently, animals that have niche-specialised following radiations may be expected to harbour genetically distinct viruses. Through a metatranscriptomic analysis of eight of New Zealand's native skink and gecko species, as well as the only introduced lizard species, the rainbow skink (Lampropholis delicata), we aimed to reveal the diversity of viruses in these hosts and determine whether and how the radiation of skinks and geckos in New Zealand has impacted virus diversity and evolution. RESULTS: We identified a total of 15 novel reptilian viruses spanning 11 different viral families, across seven of the nine species sampled. Notably, we detected no viral host-switching among the native animals analysed, even between those sampled from the same geographic location. This is compatible with the idea that host speciation has likely resulted in isolated, niche-constrained viral populations that have prevented cross-species transmission. Using a protein structural similarity-based approach, we further identified a highly divergent bunya-like virus that potentially formed a new family within the Bunyavirales. CONCLUSIONS: This study has broadened our understanding of reptilian viruses within New Zealand and illustrates how niche adaptation may limit viral-host interactions.


Assuntos
Lagartos , Animais , Lagartos/virologia , Nova Zelândia , Vírus/isolamento & purificação , Vírus/genética , Filogenia
7.
Sci Total Environ ; 943: 173648, 2024 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-38825204

RESUMO

The emergence of SARS-CoV-2 has heightened the need to evaluate the detection of enveloped viruses in the environment, particularly in wastewater, within the context of wastewater-based epidemiology. The studies published over the past 80 years focused primarily on non-enveloped viruses due to their ability to survive longer in environmental matrices such as wastewater or sludge compared to enveloped viruses. However, different enveloped viruses survive in the environment for different lengths of time. Therefore, it is crucial to be prepared to assess the potential infectious risk that may arise from future emerging enveloped viruses. This will require appropriate tools, notably suitable viral concentration methods that do not compromise virus infectivity. This review has a dual purpose: first, to gather all the available literature on the survival of infectious enveloped viruses, specifically at different pH and temperature conditions, and in contact with detergents; second, to select suitable concentration methods for evaluating the infectivity of these viruses in wastewater and sludge. The methodology used in this data collection review followed the systematic approach outlined in the PRISMA (Preferred Reporting Items for Systematic Review and Meta-Analysis) guidelines. Concentration methods cited in the data gathered are more tailored towards detecting the enveloped viruses' genome. There is a lack of suitable methods for detecting infectious enveloped viruses in wastewater and sludge. Ultrafiltration, ultracentrifugation, and polyethylene glycol precipitation methods, under specific/defined conditions, appear to be relevant approaches. Further studies are necessary to validate reliable concentration methods for detecting infectious enveloped viruses. The choice of culture system is also crucial for detection sensitivity. The data also show that the survival of infectious enveloped viruses, though lower than that of non-enveloped ones, may enable environmental transmission. Experimental data on a wide range of enveloped viruses is required due to the variability in virus persistence in the environment.


Assuntos
Esgotos , Águas Residuárias , Esgotos/virologia , Águas Residuárias/virologia , SARS-CoV-2 , Vírus/isolamento & purificação , COVID-19/transmissão
8.
PLoS Biol ; 22(6): e3002643, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38857360

RESUMO

Zebrafish are often used to model host-pathogen interactions, but few models of natural virus infection have been established. A new study in PLOS Biology shows that metatranscriptomics and cohousing experiments can uncover a natural pathogenic virus of zebrafish for laboratory study.


Assuntos
Peixe-Zebra , Peixe-Zebra/virologia , Animais , Interações Hospedeiro-Patógeno , Doenças dos Peixes/virologia , Vírus/genética
9.
J Clin Virol ; 173: 105693, 2024 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-38820916

RESUMO

BACKGROUND: Viral respiratory Infections pose a health risk, especially to vulnerable patient populations. Effective testing programs can detect and differentiate these infections at an early stage, which is particularly important for high-risk clinical departments. The objective of this study was to develop and validate a multiplex PCR-panel for 16 different respiratory viruses on a fully-automated high-throughput platform. METHODS: Three multiplex-PCR assays were designed to run on the cobas5800/6800/8800 systems, consolidating 16 viral targets: RESP1: SARS-CoV-2, influenza-A/B, RSV; RESP2: hMPV, hBoV, hAdV, rhino-/ENV; RESP3: HPIV-1-4, hCoV-229E, hCoV-NL63, hCoV-OC43, hCoV-HKU1. Analytic performance was evaluated using digital-PCR based standards and international reference material. Clinical performance was determined by comparing results from clinical samples with reference assays. RESULTS: Analytical sensitivity (i.e. lower limit of detection (LoD), 95 % probability of detection) was determined as follows: SARS-CoV-2: 29.3 IU/ml, influenza-A: 179.9 cp/ml, influenza-B: 333.9 cp/ml and RSV: 283.1 cp/ml. LoDs of other pathogens ranged between 9.4 cp/ml (hCoV-NL63) and 21,419 cp/ml (HPIV-2). Linearity was verified over 4-7 log-steps with pooled standard differentials (SD) ranging between 0.18-0.70ct. Inter-/intra-run variability (precision) was assessed for all targets over 3 days. SDs ranged between 0.13-0.74ct. Positive agreement in clinical samples was 99.4 % and 95 % for SARS-CoV-2 and influenza-A respectively. Other targets were in the 80-100 % range. Negative agreement varied between 96.3-100 %. DISCUSSION: Lab-developed tests are a key factor for effective clinical diagnostics. The multiplex panel presented in this study demonstrated high performance and provides an easily scalable high-throughput solution for respiratory virus testing, e.g. for testing in high-risk patient populations.


Assuntos
Reação em Cadeia da Polimerase Multiplex , Infecções Respiratórias , Sensibilidade e Especificidade , Humanos , Reação em Cadeia da Polimerase Multiplex/métodos , Infecções Respiratórias/virologia , Infecções Respiratórias/diagnóstico , Ensaios de Triagem em Larga Escala/métodos , Vírus/isolamento & purificação , Vírus/genética , Vírus/classificação , Viroses/diagnóstico , Viroses/virologia , Automação Laboratorial/métodos , SARS-CoV-2/genética , SARS-CoV-2/isolamento & purificação , COVID-19/diagnóstico , COVID-19/virologia , Técnicas de Diagnóstico Molecular/métodos , Técnicas de Diagnóstico Molecular/normas
10.
Int J Biol Macromol ; 269(Pt 2): 132200, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38723834

RESUMO

Viral infections continue to pose a significant global public health threat. Targeting host proteins, such as cluster of differentiation (CD) macromolecules, may offer a promising alternative approach to developing antiviral treatments. CDs are cell-surface biological macromolecules mainly expressed on leukocytes that viruses can use to enter cells, thereby evading immune detection and promoting their replication. The manipulation of CDs by viruses may represent an effective and clever means of survival through the prolonged co-evolution of hosts and viruses. Targeting of CDs is anticipated to hinder the invasion of related viruses, modulate the body's immune system, and diminish the incidence of subsequent inflammation. They have become crucial for biomedical diagnosis, and some have been used as valuable tools for resisting viral infections. However, a summary of the structures and functions of CDs involved in viral infection is currently lacking. The development of drugs targeting these biological macromolecules is restricted both in terms of their availability and the number of compounds currently identified. This review provides a comprehensive analysis of the critical role of CD proteins in virus invasion and a list of relevant targeted antiviral agents, which will serve as a valuable reference for future research in this field.


Assuntos
Antivirais , Viroses , Humanos , Antivirais/farmacologia , Antivirais/uso terapêutico , Antivirais/química , Viroses/tratamento farmacológico , Viroses/virologia , Vírus/efeitos dos fármacos , Animais , Interações Hospedeiro-Patógeno
12.
Pathog Dis ; 822024 Feb 07.
Artigo em Inglês | MEDLINE | ID: mdl-38714349

RESUMO

Respiratory pathogens can cause severe disease and even death, especially in the very young and very old. Studies investigating their prevalence often focus on individuals presenting to healthcare providers with symptoms. However, the design of prevention strategies, e.g. which target groups to vaccinate, will benefit from knowledge on the prevalence of, risk factors for and host response to these pathogens in the general population. In this study, upper respiratory samples (n = 1311) were collected cross-sectionally during winter from 11- and 24-month old children, their parents, and adults ≥60 years of age that were recruited irrespective of seeking medical care. Almost all children, approximately two-thirds of parents and a quarter of older adults tested positive for at least one pathogen, often in the absence of symptoms. Viral interference was evident for the combination of rhinovirus and respiratory syncytial virus. Attending childcare facilities and having siblings associated with increased pathogen counts in children. On average, children showed increased levels of mucosal cytokines compared to parents and especially proinflammatory molecules associated with the presence of symptoms. These findings may guide further research into transmission patterns of respiratory pathogens and assist in determining the most appropriate strategies for the prediction and prevention of disease.


Assuntos
Citocinas , Infecções Respiratórias , Estações do Ano , Humanos , Estudos Transversais , Países Baixos/epidemiologia , Lactente , Masculino , Feminino , Infecções Respiratórias/epidemiologia , Infecções Respiratórias/virologia , Infecções Respiratórias/imunologia , Prevalência , Pessoa de Meia-Idade , Adulto , Citocinas/metabolismo , Idoso , Pré-Escolar , Idoso de 80 Anos ou mais , Viroses/epidemiologia , Viroses/virologia , Viroses/imunologia , Vírus/isolamento & purificação , Vírus/classificação , Vírus/imunologia
13.
Curr Opin Virol ; 66: 101411, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38718574

RESUMO

Virus infection activates specific pattern recognition receptors and immune signal transduction, resulting in pro-inflammatory cytokine production and activation of innate immunity. We describe here the molecular organization of early signaling pathways downstream of viral recognition, including conformational changes, post-translational modifications, formation of oligomers, and generation of small-molecule second messengers. Such molecular organization allows tight regulation of immune signal transduction, characterized by swift but transient responses, nonlinearity, and signal amplification. Pathologies of early immune signaling caused by genomic mutations illustrate the fine regulation of the immune transduction cascade.


Assuntos
Imunidade Inata , Transdução de Sinais , Viroses , Humanos , Animais , Viroses/imunologia , Viroses/virologia , Viroses/metabolismo , Vírus/genética , Vírus/imunologia , Processamento de Proteína Pós-Traducional , Interações Hospedeiro-Patógeno , Receptores de Reconhecimento de Padrão/metabolismo , Citocinas/metabolismo
14.
J Infect ; 88(6): 106169, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38697269

RESUMO

Gastroenteritis viruses are the leading etiologic agents of diarrhea in children worldwide. We present data from thirty-three (33) eligible studies published between 2003 and 2023 from African countries bearing the brunt of the virus-associated diarrheal mortality. Random effects meta-analysis with proportion, subgroups, and meta-regression analyses were employed. Overall, rotavirus with estimated pooled prevalence of 31.0 % (95 % CI 24.0-39.0) predominated in all primary care visits and hospitalizations, followed by norovirus, adenovirus, sapovirus, astrovirus, and aichivirus with pooled prevalence estimated at 15.0 % (95 % CI 12.0-20.0), 10 % (95 % CI 6-15), 4.0 % (95 % CI 2.0-6.0), 4 % (95 % CI 3-6), and 2.3 % (95 % CI 1-3), respectively. Predominant rotavirus genotype was G1P[8] (39 %), followed by G3P[8] (11.7 %), G9P[8] (8.7 %), and G2P[4] (7.1 %); although, unusual genotypes were also observed, including G3P[6] (2.7 %), G8P[6] (1.7 %), G1P[6] (1.5 %), G10P[8] (0.9 %), G8P[4] (0.5 %), and G4P[8] (0.4 %). The genogroup II norovirus predominated over the genogroup I-associated infections (84.6 %, 613/725 vs 14.9 %, 108/725), with the GII.4 (79.3 %) being the most prevalent circulating genotype. In conclusion, this review showed that rotavirus remains the leading driver of viral diarrhea requiring health care visits and hospitalization among under-five years children in Africa. Thus, improved rotavirus vaccination in the region and surveillance to determine the residual burden of rotavirus and the evolving trend of other enteric viruses are needed for effective control and management of cases.


Assuntos
Gastroenterite , Humanos , Gastroenterite/virologia , Gastroenterite/epidemiologia , Pré-Escolar , Lactente , África/epidemiologia , Prevalência , Diarreia/virologia , Diarreia/epidemiologia , Rotavirus/genética , Rotavirus/isolamento & purificação , Rotavirus/classificação , Recém-Nascido , Genótipo , Viroses/epidemiologia , Viroses/virologia , Infecções por Rotavirus/epidemiologia , Infecções por Rotavirus/virologia , Vírus/classificação , Vírus/genética , Vírus/isolamento & purificação
15.
Environ Res ; 252(Pt 4): 119070, 2024 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-38710431

RESUMO

Mangrove wetlands, as one of the natural ecosystems with the most ecological services, have garnered widespread attention about their microbial driven biogeochemical cycling. Urbanization have led to different spatial patterns of environmental conditions and microbial communities in mangroves. However, viruses, as the pivotal drivers of biogeochemical cycling in mangroves, remain inadequately explored in terms of how their ecological potential and complex interactions with host respond to functional zonings. To address this knowledge gap, we conducted a comprehensive investigation on the structural and functional properties of temperate and lytic viruses in mangrove wetlands from different functional zonings by jointly using high-throughput sequencing, prokaryotic and viral metagenomics. Multiple environmental factors were found to significantly influence the taxonomic and functional composition, as well as lysogen-lysis decision-making of mangrove viruses. Furthermore, enriched auxiliary metabolic genes (AMGs) involved in methane, nitrogen and sulfur metabolism, and heavy metal resistance were unveiled in mangrove viruses, whose community composition was closely related to lifestyle and host. The virus-host pairs with different lifestyles were also discovered to react to environmental changes in different ways, which provided an empirical evidence for how virus and bacteria dynamics were specific to viral lifestyles in nature. This study expands our comprehension of the intricate interactions among virus, prokaryotic host and the environment in mangrove wetlands from multiple perspectives, including viral lifestyles, virus-host interactions, and habitat dependence. Importantly, it provides a new ecological perspective on how mangrove viruses are adapted to the stress posed by urbanization.


Assuntos
Áreas Alagadas , Vírus/genética , Ecossistema , Metagenômica
16.
Int J Biol Macromol ; 270(Pt 2): 132254, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38729501

RESUMO

Therapeutic proteins have been employed for centuries and reached approximately 50 % of all drugs investigated. By 2023, they represented one of the top 10 largest-selling pharma products ($387.03 billion) and are anticipated to reach around $653.35 billion by 2030. Growth hormones, insulin, and interferon (IFN α, γ, and ß) are among the leading applied therapeutic proteins with a higher market share. Protein-based therapies have opened new opportunities to control various diseases, including metabolic disorders, tumors, and viral outbreaks. Advanced recombinant DNA biotechnology has offered the production of therapeutic proteins and peptides for vaccination, drugs, and diagnostic tools. Prokaryotic and eukaryotic expression host systems, including bacterial, fungal, animal, mammalian, and plant cells usually applied for recombinant therapeutic proteins large-scale production. However, several limitations face therapeutic protein production and applications at the commercial level, including immunogenicity, integrity concerns, protein stability, and protein degradation under different circumstances. In this regard, protein-engineering strategies such as PEGylation, glycol-engineering, Fc-fusion, albumin conjugation, and fusion, assist in increasing targeting, product purity, production yield, functionality, and the half-life of therapeutic protein circulation. Therefore, a comprehensive insight into therapeutic protein research and findings pave the way for their successful implementation, which will be discussed in the current review.


Assuntos
Peptídeos , Humanos , Peptídeos/química , Peptídeos/uso terapêutico , Animais , Viroses/tratamento farmacológico , Viroses/prevenção & controle , Proteínas Recombinantes/uso terapêutico , Engenharia de Proteínas/métodos , Antivirais/uso terapêutico , Vírus
17.
Biomed Pharmacother ; 175: 116608, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38703502

RESUMO

Recent advances in metagenomic testing opened a new window into the mammalian blood virome. Comprised of well-known viruses like human immunodeficiency virus, hepatitis C virus, and hepatitis B virus, the virome also includes many other eukaryotic viruses and phages whose medical significance, lifecycle, epidemiology, and impact on human health are less well known and thus regarded as commensals. This review synthesizes available information for the so-called commensal virome members that circulate in the blood of humans considering their restriction to and interaction with the human host, their natural history, and their impact on human health and physiology.


Assuntos
Viroma , Humanos , Viroma/genética , Animais , Vírus/genética , Vírus/isolamento & purificação , Metagenômica/métodos , Viroses/virologia , Viroses/sangue
18.
J Clin Microbiol ; 62(6): e0034524, 2024 Jun 12.
Artigo em Inglês | MEDLINE | ID: mdl-38757981

RESUMO

Viral enrichment by probe hybridization has been reported to significantly increase the sensitivity of viral metagenomics. This study compares the analytical performance of two targeted metagenomic virus capture probe-based methods: (i) SeqCap EZ HyperCap by Roche (ViroCap) and (ii) Twist Comprehensive Viral Research Panel workflow, for diagnostic use. Sensitivity, specificity, and limit of detection were analyzed using 25 synthetic viral sequences spiked in increasing proportions of human background DNA, eight clinical samples, and American Type Culture Collection (ATCC) Virome Virus Mix. Sensitivity and specificity were 95% and higher for both methods using the synthetic and reference controls as gold standard. Combining thresholds for viral sequence read counts and genome coverage [respectively 500 reads per million (RPM) and 10% coverage] resulted in optimal prediction of true positive results. Limits of detection were approximately 50-500 copies/mL for both methods as determined by ddPCR. Increasing proportions of spike-in cell-free human background sequences up to 99.999% (50 ng/mL) did not negatively affect viral detection, suggesting effective capture of viral sequences. These data show analytical performances in ranges applicable to clinical samples, for both probe hybridization metagenomic approaches. This study supports further steps toward more widespread use of viral metagenomics for pathogen detection, in clinical and surveillance settings using low biomass samples. IMPORTANCE: Viral metagenomics has been gradually applied for broad-spectrum pathogen detection of infectious diseases, surveillance of emerging diseases, and pathogen discovery. Viral enrichment by probe hybridization methods has been reported to significantly increase the sensitivity of viral metagenomics. During the past years, a specific hybridization panel distributed by Roche has been adopted in a broad range of different clinical and zoonotic settings. Recently, Twist Bioscience has released a new hybridization panel targeting human and animal viruses. This is the first report comparing the performance of viral metagenomic hybridization panels.


Assuntos
Metagenômica , Sensibilidade e Especificidade , Vírus , Humanos , Metagenômica/métodos , Metagenômica/normas , Vírus/genética , Vírus/isolamento & purificação , Vírus/classificação , Viroses/diagnóstico , Viroses/virologia , Padrões de Referência , Técnicas de Diagnóstico Molecular/métodos , Técnicas de Diagnóstico Molecular/normas , Limite de Detecção , Hibridização de Ácido Nucleico/métodos , Viroma
19.
Environ Res ; 255: 119156, 2024 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-38759773

RESUMO

Comprehensive data on bacterial and viral pathogens of diarrhea and studies applying culture-independent methods for examining antibiotic resistance in wastewater are lacking. This study aimed to simultaneously quantify antibiotic resistance genes (ARGs), class 1 integron-integrase (int1), bacterial and viral pathogens of diarrhea, 16S rRNA, and other indicators using a high-throughput quantitative PCR (HT-qPCR) system. Thirty-six grab wastewater samples from a wastewater treatment plant in Japan, collected three times a month between August 2022 and July 2023, were centrifuged, followed by nucleic acid extraction, reverse transcription, and HT-qPCR. Fourteen targets were included, and HT-qPCR was performed on the Biomark X9™ System (Standard BioTools). For all qPCR assays, R2 was ≥0.978 and the efficiencies ranged from 90.5% to 117.7%, exhibiting high performance. Of the 36 samples, 20 (56%) were positive for Norovirus genogroup II (NoV-GII), whereas Salmonella spp. and Campylobacter jejuni were detected in 24 (67%) and Campylobacter coli in 13 (36%) samples, with mean concentrations ranging from 3.2 ± 0.8 to 4.7 ± 0.3 log10 copies/L. NoV-GII detection ratios and concentrations were higher in winter and spring. None of the pathogens of diarrhea correlated with acute gastroenteritis cases, except for NoV-GII, suggesting the need for data on specific bacterial infections to validate bacterial wastewater-based epidemiology (WBE). All samples tested positive for sul1, int1, and blaCTX-M, irrespective of season. The less explored blaNDM-1 showed a wide prevalence (>83%) and consistent abundance ranging from 4.3 ± 1.0 to 4.9 ± 0.2 log10 copies/L in all seasons. sul1 was the predominant ARG, whereas absolute abundances of 16S rRNA, int1, and blaCTX-M varied seasonally. int1 was significantly correlated with blaCTX-M in autumn and spring, whereas it showed no correlation with blaNDM-1, questioning the applicability of int1 as a sole indicator of overall resistance determinants. This study exhibited that the HT-qPCR system is pivotal for WBE.


Assuntos
Águas Residuárias , Águas Residuárias/microbiologia , Águas Residuárias/virologia , Japão , Bactérias/genética , Bactérias/efeitos dos fármacos , Bactérias/isolamento & purificação , Reação em Cadeia da Polimerase em Tempo Real/métodos , Resistência Microbiana a Medicamentos/genética , Farmacorresistência Bacteriana/genética , Genes Bacterianos , RNA Ribossômico 16S/análise , RNA Ribossômico 16S/genética , Vírus/genética , Vírus/efeitos dos fármacos , Vírus/isolamento & purificação , Microfluídica/métodos
20.
Biologicals ; 86: 101771, 2024 May.
Artigo em Inglês | MEDLINE | ID: mdl-38763040

RESUMO

In the past few years NGS has become the technology of choice to replace animal-based virus safety methods and this has been strengthened by the recent revision to the ICHQ5A virus safety chapter. Here we describe the validation of an NGS method using an agnostic analysis to detect and identify RNA virus and actively replicating DNA virus contaminants in cell banks. We report the results of the validation of each step in the sequencing process that established quality criteria to ensure consistent sequencing data. Furthermore, the validation of the analysis algorithm designed to identify virus specific sequences is described along with steps undertaken to ensure the integrity of the sequencing data from generation to analysis. Lastly, the validated sequencing and analysis systems were used to establish a limit of detection (LOD) for model viruses in cells that are commonly used in biomanufacturing. The LOD from these studies ranged from 1E+03 to 1E+04 genome copies and were dependent on the virus type with little variability between the different cell types. Thus, the validation of the NGS method for adventitious agent testing and the establishment of a general LOD for cell-based samples provides a suitable alternative to traditional virus detection methods.


Assuntos
Sequenciamento de Nucleotídeos em Larga Escala , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Animais , Humanos , Linhagem Celular , Limite de Detecção , Vírus/genética , Vírus/isolamento & purificação , Vírus de RNA/genética , Vírus de RNA/isolamento & purificação , Vírus de DNA/genética
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