Search

Monkeypox virus (MPXV) was confirmed in May 2022 and designated a global health emergency by WHO in July 2022. MPX virions are big, enclosed, brick-shaped, and contain a linear, double-stranded DNA genome as well as enzymes. MPXV particles bind to the host cell membrane via a variety of viral-host protein interactions. As a result, the wrapped structure is a potential therapeutic target. DeepRepurpose, an artificial intelligence-based compound-viral proteins interaction framework, was used via a transfer learning setting to prioritize a set of FDA approved and investigational drugs which can potentially inhibit MPXV viral proteins. To filter and narrow down the lead compounds from curated collections of pharmaceutical compounds, we used a rigorous computational framework that included homology modeling, molecular docking, dynamic simulations, binding free energy calculations, and binding pose metadynamics. We identified Elvitegravir as a potential inhibitor of MPXV virus using our comprehensive pipeline., Competing Interests: Conflicts of Interest The authors declare that there are no conflicts of interest., (Copyright © 2023 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Background: In May 2022, the monkeypox virus (MPXV) spread into non-endemic countries and the global community was quick to test the lessons learned from the SARS-CoV-2 pandemic. Due to its symptomatic resemblance to other diseases, like the non-pox virus varicella zoster (chickenpox), polymerase chain reaction methods play an important role in correctly diagnosing the rash-causing pathogen. INSTAND quickly established a new external quality assessment (EQA) scheme for MPXV and orthopoxvirus (OPXV) DNA detection to assess the current performance quality of the laboratory tests., Methods: We analyzed quantitative and qualitative data of the first German EQA for MPXV and OPXV DNA detection. The survey included one negative and three MPXV-positive samples with different MPX viral loads. The threshold cycle (Ct) or other measures defining the quantification cycle (Cq) were analyzed in an assay-specific manner. A Passing Bablok fit was used to investigate the performance at laboratory level., Results: 141 qualitative datasets were reported by 131 laboratories for MPXV detection and 68 qualitative datasets by 65 laboratories for OPXV detection. More than 96% of the results were correctly identified as negative and more than 97% correctly identified as positive. An analysis of the reported Ct/Cq values showed a large spread of these values of up to 12 Ct/Cq. Nevertheless, there is a good correlation of results for the different MPXV concentrations at laboratory level. Only a few quantitative results in copies/mL were reported (MPXV: N = 5; OPXV: N = 2), but the results correlated well with the concentration differences between the EQA samples, which were to a power of ten each., Conclusion: The EQA results show that laboratories performed well in detecting both MPXV and OPXV. However, Ct/Cq values should be interpreted with caution when conclusions are drawn about the viral load as long as metrological traceability is not granted., Competing Interests: I have read the journal’s policy and the authors of this manuscript have the following competing interests: HZ declares that he was majority owner of GBD Gesellschaft fuer Biotechnologische Diagnostik mbH, Berlin, and owner and managing director of IQVD GmbH - Institut fuer Qualitaetssicherung, Berlin. HPG declares that he was minority owner of GBD Gesellschaft fuer Biotechnologische Diagnostik mbH, Berlin. HZ and HPG declare that they were managing directors of GBD Gesellschaft fuer Biotechnologische Diagnostik mbH, Berlin, during the study. This does not alter our adherence to PLoS One policies on sharing data and materials. All other authors have declared that no competing interests exist., (Copyright: © 2023 Vierbaum et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

During the 2022 monkeypox (MPX) outbreak, testing has been limited and results delayed, allowing ongoing transmission. Gold-standard quantitative polymerase chain reaction (qPCR) diagnostics are difficult to obtain. This research adapted the June 2022 CDC MPX qPCR assay for broad implementation. Validated using MPX stocks in a matrix with multiple sample types, MPX was detected with cycle threshold (Ct) values 17.46-35.59 and titer equivalents 8.01 × 106 to 2.45 × 100 plaque-forming unit (PFU)/mL. The detection limit was 3.59 PFU/mL. Sensitivity and specificity were both 100%. This qPCR assay can be quickly and broadly implemented in research and public health laboratories to increase diagnostic capacity amid the growing MPX outbreak., Competing Interests: Potential conflicts of interest. C. N. M. has received funding from the Centers for Disease Control and Prevention, World Health Organization, and the German Government; consulting fees from Curative Inc and Singletto Inc; and has stock options with Singletto Inc. All other authors report no potential conflicts. All authors have submitted the ICMJE Form for Disclosure of Potential Conflicts of Interest. Conflicts that the editors consider relevant to the content of the manuscript have been disclosed., (© The Author(s) 2022. Published by Oxford University Press on behalf of Infectious Diseases Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

The ongoing global pandemic caused by a variant of the monkeypox (or mpox) virus (MPXV) has prompted widespread concern. The MPXV DNA polymerase holoenzyme, consisting of F8, A22, and E4, is vital for replicating the viral genome and represents a crucial target for the development of antiviral drugs. However, the assembly and working mechanism for the DNA polymerase holoenzyme of MPXV remains elusive. Here, we present the cryo-electron microscopy (cryo-EM) structure of the DNA polymerase holoenzyme at an overall resolution of 3.5 Å. Unexpectedly, the holoenzyme is assembled as a dimer of heterotrimers, of which the extra interface between the thumb domain of F8 and A22 shows a clash between A22 and substrate DNA, suggesting an autoinhibition state. Addition of exogenous double-stranded DNA shifts the hexamer into trimer exposing DNA binding sites, potentially representing a more active state. Our findings provide crucial steps toward developing targeted antiviral therapies for MPXV and related viruses.

We describe the results of a prospective observational study of the clinical natural history of human monkeypox (mpox) virus (MPXV) infections at the remote L'Hopital General de Reference de Kole (Kole hospital), the rainforest of the Congo River basin of the Democratic Republic of the Congo (DRC) from March 2007 until August 2011. The research was conducted jointly by the Institute National de Recherche Biomedical (INRB) and the US Army Medical Research Institute of Infectious Diseases (USAMRIID). The Kole hospital was one of the two previous WHO Mpox study sites (1981-1986). The hospital is staffed by a Spanish Order of Catholic Nuns from La Congregation Des Soeurs Missionnaires Du Christ Jesus including two Spanish physicians, who were members of the Order as well, were part of the WHO study on human mpox. Of 244 patients admitted with a clinical diagnosis of MPXV infection, 216 were positive in both the Pan-Orthopox and MPXV specific PCR. The cardinal observations of these 216 patients are summarized in this report. There were three deaths (3/216) among these hospitalized patients; fetal death occurred in 3 of 4 patients who were pregnant at admission, with the placenta of one fetus demonstrating prominent MPXV infection of the chorionic villi. The most common complaints were rash (96.8%), malaise (85.2%), sore throat (78.2%), and lymphadenopathy/adenopathy (57.4%). The most common physical exam findings were mpox rash (99.5%) and lymphadenopathy (98.6%). The single patient without the classic mpox rash had been previously vaccinated against smallpox. Age group of less than 5 years had the highest lesion count. Primary household cases tended to have higher lesion counts than secondary or later same household cases. Of the 216 patients, 200 were tested for IgM & IgG antibodies (Abs) to Orthopoxviruses. All 200 patients had anti-orthopoxvirus IgG Abs; whereas 189/200 were positive for IgM. Patients with hypoalbuminemia had a high risk of severe disease. Patients with fatal disease had higher maximum geometric mean values than survivors for the following variables, respectively: viral DNA in blood (DNAemia); maximum lesion count; day of admission mean AST and ALT., Competing Interests: The authors have no competing interests with products or companies described in this article., (Copyright: This is an open access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. The work is made available under the Creative Commons CC0 public domain dedication.)

Monkeypox is a disease with pandemic potential. It is caused by the monkeypox virus (MPXV), a double-stranded DNA virus from the Poxviridae family, that replicates in the cytoplasm and must encode for its own RNA processing machinery including the capping machinery. Here, we present crystal structures of its 2'-O-RNA methyltransferase (MTase) VP39 in complex with the pan-MTase inhibitor sinefungin and a series of inhibitors that were discovered based on it. A comparison of this 2'-O-RNA MTase with enzymes from unrelated single-stranded RNA viruses (SARS-CoV-2 and Zika) reveals a conserved sinefungin binding mode, implicating that a single inhibitor could be used against unrelated viral families. Indeed, several of our inhibitors such as TO507 also inhibit the coronaviral nsp14 MTase., (© 2023. The Author(s).)

Mpox (previously known as monkeypox) is an infectious viral illness caused by the mpox virus (MPXV), an orthopoxvirus that belongs to the family Poxviridae . The symptoms of mpox in humans are similar to those of smallpox, although the mortality rate is lower. In recent years, the concern over a potential global pandemic has increased due to reports of mpox spreading across Africa and other parts of the world. Prior to this discovery, mpox was a rare zoonotic disease restricted to endemic regions of Western and Central Africa. The sudden emergence of MPXV cases in multiple regions has raised concerns about its natural evolution. This review aims to provide an overview of previously available information about MPXV, including its genome, morphology, hosts and reservoirs, and virus-host interaction and immunology, as well as to perform phylogenetic analysis on available MPXV genomes, with an emphasis on the evolution of the genome in humans as new cases emerge.

In 2003, the United States saw an epidemic of monkeypox that was later traced back to rodents of West Africa infected with the monkeypox virus (MPXV). Disease in the United States seemed less severe than the smallpox-like disease in the Democratic Republic of the Congo (DRC). In this study, researchers analyzed data from Central Africa: two distinct MPXV clades were confirmed by sequencing the genomes of MPXV isolates from Western Africa, the United States, and Central Africa. By comparing open reading frames across MPXV clades, scientists can infer which virus proteins might account for the observed variation in pathogenicity in humans. Monkeypox can be prevented and controlled with a better understanding of MPXV's molecular etiology and epidemiological and clinical features. In light of the current outbreaks worldwide, we provide updated information on monkeypox for medical professionals in this review.

Background: A large, worldwide outbreak of mpox (formerly referred to as monkeypox) involving mainly men who have sex with men commenced in May 2022. We evaluated the frequency of positivity for the causative agent, monkeypox virus (MPXV), in blood donations collected in August 2022, during the outbreak period in Southern England., Methods/materials: The sensitivity and specificity of an MPXV-specific PCR and a generic non-variola orthopoxvirus (NVO) PCR were evaluated using samples from mpox cases and synthetic DNA standards. Residual minipools from nucleic acid testing were obtained from 10,896 blood donors in Southern England, with 21% from London., Results: MPXV and NVO PCRs were both capable of detection of single copies of target sequence with calculated limits of detection (LOD) 90  s of 2.3 and 2.1 DNA copies and analytical sample sensitivities of 46 and 42 MPXV DNA copies/ml, respectively. 454 minipools produced from 10,896 unique donors were assayed for MPXV DNA by both methods. No positive minipools were detected by either PCR., Conclusions: Although blood donors are unrepresentative of the UK population in terms of MPXV infection risk, the uniformly negative MPXV DNA testing results provide reassurance that MPXV viraemia and potential transmission risk were rare or absent in donors during the outbreak period. Minipools from blood donors allow rapid implementation of large-scale population-based screening for emerging pathogens and represent an important resource for pandemic preparedness., (© 2023 The Authors. Transfusion published by Wiley Periodicals LLC on behalf of AABB.)

Monkeypox virus (MPXV) is a double-stranded DNA zoonotic virus of the Poxviridae family. Infected persons, animals, or inanimate items can all spread the virus to humans when they come into close contact. The first human-to-human transmission was reported in 1970 in the Democratic Republic of Congo. The outbreak emerged in May 2022 involved mostly men who had sex with men (MSM). Patients usually present with symptoms of rash along with fever, flu-like symptoms, and lesions in the genital and perineal region. A rising concern is ocular manifestations seen with MPVX like conjunctivitis, blepharitis, keratitis, and corneal lesions, especially in unvaccinated patients which might lead to blindness. Although it is self-limiting with supportive care, many patients benefited from tecovirimat. Combination therapy of brincidofovir and tecovirimat was also used for severe disease. Smallpox vaccinations will also play a vital role as unvaccinated patients had serious complications. Risk counseling should be carried out to prevent further spread amongst high-risk populations. Ophthalmologists should also remain aware of these ocular manifestations during the current outbreak and keep it as a differential diagnosis whenever they come across with aforementioned complaints that can be seen in MPVX illness., (Copyright © 2023 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Monkeypox virus (MPXV) is a double-stranded DNA virus belonging to the Poxviridae family of the genus Orthopoxvirus with two different clades known as West African and Congo Basin. Monkeypox (MPX) is a zoonosis that arises from the MPXV and causes a smallpox-like disease. The endemic disease status of MPX was updated to an outbreak worldwide in 2022. Thus, the condition was declared a global health emergency independent of travel issues, accounting for the primary reason for its prevalence outside Africa. In addition to identified transmission mediators through animal-to-human and human-to-human, especially sexual transmission among men who have sex with men came to prominence in the 2022 global outbreak. Although the severity and prevalence of the disease differ depending on age and gender, some symptoms are commonly observed. Clinical signs such as fever, muscle and headache pain, swollen lymph nodes, and skin rashes in defined body regions are standard and an indicator for the first step of diagnosis. By following the clinical signs, laboratory diagnostic tests like conventional polymerase chain reaction (PCR) or real-time PCR (RT-PCR) are the most common and accurate diagnostic methods. Antiviral drugs such as tecovirimat, cidofovir, and brincidofovir are used for symptomatic treatment. There is no MPXV-specific vaccine; however, currently available vaccines against smallpox enhance the immunization rate. This comprehensive review covers the MPX disease history and the current state of knowledge by assessing broad topics and views related to disease origin, transmission, epidemiology, severity, genome organization and evolution, diagnosis, treatment, and prevention., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier Ltd.. All rights reserved.)

We retrospectively screened oropharyngeal and rectal swab samples originally collected in California, USA, for Chlamydia trachomatis and Neisseria gonorrhoeae testing for the presence of monkeypox virus DNA. Among 206 patients screened, 17 (8%) had samples with detectable viral DNA. Monkeypox virus testing from mucosal sites should be considered for at-risk patients.

Background: In the 2022 mpox outbreak, several studies have explored longitudinal DNA shedding of mpox virus (MPXV) using PCR. However, there are fewer studies assessing infectivity in cell culture, and, by inference, MPXV transmissibility. Such information could help inform infection control and public health guidelines., Aims and Methods: The aim of this study was to correlate cell culture infectivity of clinical samples with viral loads in clinical samples. Between May to October 2022, clinical samples from different body sites sent to the Victorian Infectious Diseases Reference Laboratory in Melbourne, Australia for MPXV PCR detection were cultured in Vero cells as a surrogate for infectivity., Results: In the study period, 144 samples from 70 patients were tested by MPXV PCR. Viral loads in skin lesions were significantly higher than those in throat or nasopharyngeal samples (median Ct 22.0 vs 29.0, p = 0.0013 and median Ct 22.0 vs 36.5, p = 0.0001, respectively). Similarly, viral loads were significantly higher in anal samples compared to throat or nasopharyngeal samples (median Ct 20.0 vs. 29.0, p=<0.0001 and median Ct 20.0 vs. 36.5, p=<0.0001, respectively). Viral culture was successfully performed in 80/94 samples. Using logistic regression analysis, 50% of the samples were positive in viral culture at Ct 34.1 (95% confidence intervals 32.1-37.4)., Conclusions: Our data further validate recent findings showing that samples with a higher MPXV viral load are more likely to demonstrate infectivity in cell culture. Although the presence of infectious virus in cell culture may not directly translate with clinical transmission risk, our data may be used as an adjunct help inform guidelines on testing and isolation policies in individuals with mpox., Competing Interests: Declaration of Competing Interest We declare no competing interests. DAW, EPFC, and CKF are supported by Investigator Grants from the National Health and Medical Research Council (NHMRC) of Australia (GNT1174555, GNT1172873 and GNT1172900 respectively. This work was approved by the Royal Melbourne Hospital Human Research Ethics Committee (QA2022085 and HREC/79,322/MH-2021)., (Copyright © 2023 The Author(s). Published by Elsevier B.V. All rights reserved.)

Using data from 12 US health departments, we estimated mean serial interval for monkeypox virus infection to be 8.5 (95% credible interval 7.3-9.9) days for symptom onset, based on 57 case pairs. Mean estimated incubation period was 5.6 (95% credible interval 4.3-7.8) days for symptom onset, based on 35 case pairs.

Background: The current global mpox virus (MPXV) outbreak has been declared a Public Health Emergency of International Concern by WHO, with more than 80,000 cases confirmed across multiple continents. Diagnosis is confirmed by PCR of viral DNA from vesicle and other swabs., Objective: The aim of this study was to assess commercial RT PCR assays for Orthopoxvirus (OPX) and MPXV for analytical sensitivity, and percent agreements and compare them to primer/probe sets employed at the Victorian Infectious Diseases Reference Laboratory (VIDRL), Centers for Disease Control andPrevention (CDC) and US Army Medical Research Institute of Infectious Diseases (USAMRIID). Limits of detection (LOD), intra-run variability, cross-reactivity and performance on forty clinical samples was assessed on eleven commercial assays and five primer/probe combinations used at VIDRL, CDC and USAMRIID., Results: All assays were able to detect OPX and MPXV (LOD 57 to 14,495 copies/mL) with intra-run coefficients of variation between Cycle thresholds of 0.58 and 3.44, and there was no unexpected cross-reactivity. All assays demonstrated 100% negative percent agreement with clinical samples and all but one yielded 100% positive percent agreement., Conclusions: Variations in LOD between assays may be dependent on the platform used and sample type. Despite the overall comparable performance of the assays assessed, it is important that routine laboratories perform in-house validations before implementing RT PCR for OPX and/or MPXV as reliable and accurate laboratory diagnosis of MPXV and isolation is crucial to containing the spread of this current outbreak and informing public health interventions and response., Competing Interests: Declaration of Competing Interest The authors have no Conflicts of Interests to declare., (Crown Copyright © 2023. Published by Elsevier B.V. All rights reserved.)

While the world is still recovering from the Covid-19 pandemic, monkeypox virus (MPXV) awaits to cause another global outbreak as a challenge to all of mankind. However, the Covid-19 pandemic has taught us a lesson to speed up the pace of viral genomic research for the implementation of preventive and treatment strategies. One of the important aspects of MPXV that needs immediate insight is its evolutionary lineage based on genomic studies. Utilizing high-quality isolates from the GISAID (Global Initiative on Sharing All Influenza Data) database, primarily sourced from Europe and North America, we employed a SNP-based whole-genome phylogeny method and identified four major clusters among 628 MPXV isolates. Our findings indicate a distinct evolutionary lineage for the first MPXV isolate, and a complex epidemiology and evolution of MPXV strains across various countries. Further analysis of the host-pathogen interaction network revealed key viral proteins, such as E3, SPI-2, K7 and CrmB, that play a significant role in regulating the network and inhibiting the host's cellular innate immune system. Our structural analysis of proteins E3 and CrmB revealed potential disruption of stability due to certain mutations. While this study identified a large number of mutations within the new outbreak clade, it also reflected that we need to move fast with the genomic analysis of newly detected strains from around the world to develop better prevention and treatment methods.

The global spread of monkeypox virus has raised concerns over the establishment of novel enzootic reservoirs in expanded geographic regions. We demonstrate that although deer mice are permissive to experimental infection with clade I and II monkeypox viruses, the infection is short-lived and has limited capability for active transmission.

Background: Monkeypox is an emerging zoonosis endemic to Central and West Africa. Monkeypox virus (MPXV) is genetically structured in 2 major clades (clades 1 and 2/3), but its evolution is poorly explored., Methods: We retrieved MPXV genomes from public repositories and we analyzed geographic patterns using STRUCTURE. Molecular dating was performed using a using a Bayesian approach., Results: We show that the population transmitted in West Africa (clades 2/3) experienced limited drift. Conversely, clade 1 (transmitted in the Congo Basin) possibly underwent a bottleneck or founder effect. Depending on the model used, we estimated that the 2 clades separated ∼560-860 (highest posterior density: 450-960) years ago, a period characterized by expansions and contractions of rainforest areas, possibly creating the ecological conditions for the MPXV reservoir(s) to migrate. In the Congo Basin, MPXV diversity is characterized by 4 subpopulations that show no geographic structuring. Conversely, clades 2/3 are spatially structured with 2 populations located West and East of the Dahomey Gap., Conclusions: The distinct histories of the 2 clades may derive from differences in MPXV ecology in West and Central Africa., Competing Interests: Potential conflicts of interest. All authors: No reported conflicts of interest. All authors have submitted the ICMJE Form for Disclosure of Potential Conflicts of Interest., (© The Author(s) 2022. Published by Oxford University Press on behalf of the Infectious Diseases Society of America.)

Monkeypox virus (MPXV) is a dsDNA virus, belonging to Poxviridae family. The outbreak of monkeypox disease in humans is critical in European and Western countries, owing to its origin in African regions. The highest number of cases of the disease were found in the United States, followed by Spain and Brazil. Understanding the complete infection mechanism of diverse MPXV strains and their interaction with humans is important for therapeutic drug development, and to avoid any future epidemics. Using computational systems biology, we deciphered the genome-wide protein-protein interactions (PPIs) between 22 MPXV strains and human proteome. Based on phylogenomics and disease severity, 3 different strains of MPXV: Zaire-96-I-16, MPXV-UK&#95;P2, and MPXV&#95;USA&#95;2022&#95;MA001 were selected for comparative functional analysis of the proteins involved in the interactions. On an average, we predicted around 92,880 non-redundant PPIs between human and MPXV proteomes, involving 8014 host and 116 pathogen proteins from the 3 strains. The gene ontology (GO) enrichment analysis revealed 10,624 common GO terms in which the host proteins of 3 strains were highly enriched. These include significant GO terms such as platelet activation (GO:0030168), GABA-A receptor complex (GO:1902711), and metalloendopeptidase activity (GO:0004222). The host proteins were also significantly enriched in calcium signaling pathway (hsa04020), MAPK signaling pathway (hsa04010), and inflammatory mediator regulation of TRP channels (hsa04750). These significantly enriched GO terms and KEGG pathways are known to be implicated in immunomodulatory and therapeutic role in humans during viral infection. The protein hubs analysis revealed that most of the MPXV proteins form hubs with the protein kinases and AGC kinase C-terminal domains. Furthermore, subcellular localization revealed that most of the human proteins were localized in cytoplasm (29.22%) and nucleus (26.79%). A few drugs including Fostamatinib, Tamoxifen and others were identified as potential drug candidates against the monkeypox virus disease. This study reports the genome-scale PPIs elucidation in human-monkeypox virus pathosystem, thus facilitating the research community with functional insights into the monkeypox disease infection mechanism and augment the drug development., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Kataria, Kaur and Kaundal.)

The ongoing outbreak of the monkeypox, caused by monkeypox virus (MPXV), has been a public health emergency of international concern, indicating an urgent need for rapid and sensitive MPXV detection. Here, we designed a diagnostic test based on loop-mediated isothermal amplification (LAMP) and nanoparticle-based lateral flow biosensor(LFB)for diagnosis of MPXV infection, termed MPX-LAMP-LFB. A set of six LAMP primers was designed based the ATI gene of MPXV, and LAMP amplification of MPXV templates was performed at 63°C for only 40 min. The results were rapidly and visually decided using the LFB test within 2 min. The MPX-LAMP-LFB assay can specifically detect MPXV strains without cross-reaction with non-MPXV pathogens. The sensitivity of the MPX-LAMP-LFB assay is as low as 5 copies/μl of plasmid template and 12.5 copies/μl of pseudovirus in human blood samples. The whole process of the MPX-LAMP-LFB assay could be completed ~1 h, including rapid template preparation (15 min), LAMP reaction (40 min)and result reporting (<2 min). Collectively, MPX-LAMP-LFB assay developed here is a useful tool for rapid and reliable diagnosis of MPXV infection., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Huang, Xiao, Jia, Sun, Fu, Xu, Cui, Huang, Qu, Zhou and Wang.)

We performed anorectal testing in 18 cis-gender men who have sex with men with symptoms consistent with mpox virus (MPXV) infection. We found rectal MPXV DNA in 9/9 with and 7/9 without proctitis. Future study of anorectal testing is needed and may inform the diagnosis and pathogenesis of MPXV disease., Competing Interests: Potential conflicts of interest. P. F. R. reports grants or contracts from Merck, Finch, Summit, Artugen, Armata, Contrafect, and Janssen. All other authors report no potential conflicts. All authors have submitted the ICMJE Form for Disclosure of Potential Conflicts of Interest. Conflicts that the editors consider relevant to the content of the manuscript have been disclosed., (© The Author(s) 2022. Published by Oxford University Press on behalf of Infectious Diseases Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Human monkeypox has attracted attention recently. Monkeypox virus (MPXV) keeps evolving as it spreading around the world rapidly, which may threaten the health of more and more people. Here, we have developed a high order reference method based on digital PCR (dPCR) for MPXV detection, of which the limits of quantification (LoQ) and detection (LoD) are 38 and 6 copies/reaction, respectively. Pseudovirus reference materials (RM) containing the conserved F3L gene has been developed, and the homogeneity assessment showed that the RM was homogeneous. The reference value with its expanded uncertainty determined by the established dPCR is (2.74 ± 0.46) × 10 3 copies/μL. Six different MPXV test kits were accessed by the RM. Four out of six test kits cannot reach their claimed LoDs. The poor analytical sensitivity might cause false-negative results, which lead to incorrect diagnosis and treatment. The establishment of a high order reference method of dPCR and pseudovirus RM is very useful for improving the accuracy and reliability of MPXV detection., (© 2023. Springer-Verlag GmbH Germany, part of Springer Nature.)

The recent increase in monkeypox (MPX) cases has attracted attention of public health authorities due to its quick spread and transmission across non-endemic regions. This outbreak, unlike previous ones, displays different epidemiological features and transmission dynamics, which appear to be largely influenced by the newly divergent MPX lineages (B.1). Yet, the genomic characteristics driving the high dispersal and diversification of these lineages remain largely unknown. Herein, we sought to explore and characterize the genomic features and phylogenetic diversity of the B.1 lineages through a comparative genomic analysis inclusive of 1900 high quality complete MPXV genomes. Our analyses indicate that the current MPXV-2022 outbreak encompasses thirteen derived lineages with ten unique non-synonymous mutations in several genes linked to immune evasion, virulence factors and host recognition. Such mutations may translate in the rapid evolution and diversification of current MPXV lineages. Moreover, our analyses uncovered signals of genomic modifications suggestive of immune-modulatory enzymatic activity, such as APOBEC3 editing, which, as previously suggested could have favored evolutionary trends leading to the rapid spread of MPXV into non-endemic countries. Genomic surveillance continues to play a major role in unveiling the genomic signatures signaling potential adaptation of this emerging MPXV lineage and how it will continue to impact public health in the near future., Competing Interests: Declaration of competing interest Nothing to declare., (Copyright © 2023 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Given the paucity of antiviral treatments for monkeypox disease, caused by the Monkeypox virus (MPXV), there is a pressing need for the development/identification of new drugs to treat the infection. MPXV possesses a linear dsDNA genome that is replicated by a DNA replication complex of which DNA polymerase (DPol) forms an important component. Owing to the importance of DPol in the viral life cycle, identifying/designing small molecules abolishing its function could yield new antivirals. In this study, we first used the AlphaFold artificial intelligence program to model the 3D structure of the MPXV DPol; like the fold of DPol from other organisms, the MPXV DPol structure has the characteristic exonuclease, thumb, palm, and fingers sub-domains arrangement. Subsequently, we have identified several inhibitors through virtual screening of ZINC and antiviral libraries. Molecules with phenyl scaffold along with alanine-based and tetrazole-based molecules showed the best docking score of -8 to -10 kcal/mol. These molecules bind in the palm and fingers sub-domains interface region, which partially overlaps with the DNA binding path. The delineation of DPol/inhibitor interactions showed that majorly active site residues ASP549, ASP753, TYR550, ASN551, SER552, and ASN665 interact with the inhibitors. These compounds exhibit good Absorption, Distribution, Metabolism and Excretion properties., Competing Interests: Declaration of competing interest The authors declare no conflicts of interest., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Human mpox (monkeypox), a disease with similarities to smallpox, is endemic in Africa where it has persisted as a zoonosis with limited human-to-human spread. Unexpectedly, the disease expanded globally in 2022 driven by human-to-human transmission outside of Africa. It is not yet known whether the latter is due solely to behavioral and environmental factors or whether the mpox virus is adapting to a new host. Genome sequencing has revealed differences between the current outbreak strains, classified as clade IIb, and the prior clade IIa and clade I viruses, but whether these differences contribute to virulence or transmission has not been determined. We demonstrate that the wild-derived inbred castaneous mouse provides an exceptional animal model for investigating clade differences in mpox virus virulence and show that the order is clade I > clade IIa > clade IIb.1. The greatly reduced replication of the clade IIb.1 major outbreak strain in mice and absence of lethality at 100 times the lethal dose of a closely related clade IIa virus, despite similar multiplication in cell culture, suggest that clade IIb is evolving diminished virulence or adapting to other species.

Despite the proven value of applying genomic data for epidemiological purposes, commonly used high-throughput sequencing formats are not adapted to the response times required to intervene and finally control outbreaks. In this study, we propose a fast alternative to whole-genome sequencing (WGS) to track relevant microbiological strains: nanopore sequencing of multiple amplicons including strain marker single nucleotide polymorphisms (SNPs). As a proof a concept, we evaluated the performance of our approach to offer a rapid response to the most recent public health global alarm, the monkeypox virus (MPXV) global outbreak. Through a multisequence alignment, a list of 42 SNPs were extracted as signature makers for this outbreak. Twenty primer pairs were designed to amplify in a multiplex PCR the regions including 22 of these SNPs. Amplicon pools were sequenced in a MinION device, and SNPs were called in real time by an in-house bioinformatic pipeline. A total of 120 specimens (95 MPXV-PCR positive, Ct values from 14 to 39) were selected. In 67.37% of the positive subset, all 22 SNPs were called. After excluding low viral load specimens, in 92% of samples ≥11 outbreak SNPs were called. No false positives were observed in any of the 25 negative specimens. The total turnaround time required for this strategy was 5 hours, and the cost per sample was 14 euros. Nanopore sequencing of multiple amplicons harboring signature SNPs escapes the targeting limitations of strain-specific PCRs and offers a powerful alternative to systematic WGS, paving the way to real-time genomic epidemiology and making immediate intervention possible to finally optimize transmission control. IMPORTANCE Nanopore sequencing of multiple amplicons harboring signature single nucleotide polymorphisms (SNPs) escapes the targeting limitations of strain-specific PCRs and offers a powerful alternative to systematic whole-genome analysis, paving the way to real-time genomic epidemiology and making immediate intervention possible to finally optimize transmission control.

Monkeypox is a zoonotic viral infection that was first identified in humans in 1970 in the Democratic Republic of Congo. The cases seen again in early May 2022 have reached 78.000 as of today. On July 23, 2022, the World Health Organization decided that the monkeypox outbreak represents a public health emergency. For the early diagnosis and effective treatment of monkeypox, inter-individual transmission routes, disease symptoms, factors affecting the course of the disease, presence of another infection, prognosis, pharmacological agents used in the prophylactic treatment, and their effects, populations at risk, waste disposal protocol should be known. For this reason, our aim is to reveal the sources of transmission of the monkeypox virus from past to present, what are the signs and symptoms in patients after infection, ways of protection from the virus, the mutation status of the virus, and treatment approaches.

Monkeypox virus (MPXV) has gained interest because of a multicountry outbreak of mpox (formerly monkeypox) cases with no epidemiologic link to MPXV-endemic regions. We sequenced the complete genome of MPXV isolated from a patient in northern Mexico. Phylogenetic analysis grouped the virus with isolates from Germany.

Objectives: A global outbreak of monkeypox virus infections in human beings has been described since April 2022. The objectives of this study were to describe the clinical characteristics and complications of patients with a monkeypox infection., Methods: All consecutive patients with a polymerase chain reaction (PCR)-confirmed monkeypox infection seen in a French referral centre were included., Results: Between 21 May and 5 July 2022, 264 patients had a PCR-confirmed monkeypox infection. Among them, 262 (262/264, 99%) were men, 245 (245/259, 95%) were men who have sex with men, and 90 (90/216, 42%) practiced chemsex in the last 3 months. Seventy-three (73/256, 29%) patients were living with human immunodeficiency virus infection, and 120 (120/169, 71%) patients were taking pre-exposure prophylaxis against human immunodeficiency virus infection. Overall, 112 (112/236, 47%) patients had contact with a confirmed monkeypox case; it was of sexual nature for 95% of the contacts (86/91). Monkeypox virus PCR was positive on the skin in 252 patients, on the oropharyngeal sample in 150 patients, and on blood in eight patients. The majority of patients presented with fever (171/253, 68%) and adenopathy (174/251, 69%). Skin lesions mostly affected the genital (135/252, 54%) and perianal (100/251, 40%) areas. Overall, 17 (17/264, 6%) patients were hospitalized; none of them were immunocompromised. Complications requiring hospitalization included cellulitis (n = 4), paronychia (n = 3), severe anal and digestive involvement (n = 4), non-cardia angina with dysphagia (n = 4), blepharitis (n = 1), and keratitis (n = 1). Surgical management was required in four patients., Conclusion: The current outbreak of monkeypox infections has specific characteristics: it occurs in the men who have sex with men community; known contact is mostly sexual; perineal and anal areas are frequently affected; and severe complications include superinfected skin lesions, paronychia, cellulitis, anal and digestive involvement, angina with dysphagia, and ocular involvement., (Copyright © 2022 European Society of Clinical Microbiology and Infectious Diseases. Published by Elsevier Ltd. All rights reserved.)

In early May 2022, the first worldwide monkeypox virus (MPXV) outbreak was reported, with different clinical aspects from previously studied human monkeypox infections. Despite monkeypox medical importance, much of its biological aspects remain to be further investigated. In the present work, we evaluated ultrastructural aspects of MPXV asynchronous infections in Vero cells by transmission electron microscopy (TEM). The viral strain was isolated from a male patient infected during the 2022 outbreak. TEM analysis showed: (i) adhered intracellular mature virus particles before entry of the host cell; (ii) a reorganization of the rough endoplasmic reticulum cisternae into the so-called "mini-nuclei" structure associated with genome replication; and (iii) noticeably different sites within the viral factory presenting granular or fibrillar aspects. We also observed viral crescents, different MPXV particle morphotypes, and cellular alterations induced by infection, such as changes in the cytoskeleton structure and multimembrane vesicles abundance. Taken together, to the best of our knowledge, these results revealed for the first-time ultrastructural aspects of different steps of the MPXV cycle., (© 2023 Wiley Periodicals LLC.)

Background: In spring of 2022, an outbreak of monkeypox (mpox) spread worldwide. Here, we describe performance characteristics of monkeypox virus (MPXV)-specific and pan-orthopoxvirus qPCR assays for clinical use., Methods: We validated probe-based qPCR assays targeting MPXV-specific loci F3L and G2R (genes MPXVgp052/OPG065 and MPXVgp002 and gp190/OPG002, respectively) and a pan-orthopoxvirus assay targeting the E9L locus (MPXVgp057/OPG071). Clinical samples and synthetic controls were extracted using the Roche MP96 or Promega Maxwell 48 instrument. qPCR was performed on the AB7500 thermocycler. Synthetic control DNA and high concentration clinical samples were quantified by droplet PCR. Cross-reactivity was evaluated for camelpox and cowpox genomic DNA, vaccinia culture supernatant, and HSV- and VZV-positive clinical specimens. We also tested the performance of the F3L assay using dry swabs, Aptima vaginal and rectal swabs, nasopharyngeal, rectal, and oral swabs, cerebrospinal fluid, plasma, serum, whole blood, breastmilk, urine, saliva, and semen., Results: The MPXV-F3L assay is reproducible at a limit of detection (LoD) of 65.6 copies/mL of viral DNA in viral transport medium/universal transport medium (VTM/UTM), or 3.3 copies/PCR reaction. No cross-reactivity with herpesviruses or other poxviruses was observed. MPXV-F3L detects MPXV DNA in alternative specimen types, with an LoD ranging between 260-1000 copies/mL, or 5.7-10 copies/PCR reaction. In clinical swab VTM specimens, MPXV-F3L and MPXV-G2R assays outperformed OPXV-E9L by an average of 2.4 and 2.8 Cts, respectively. MPXV-G2R outperformed MPXV-F3L by 0.4 Cts, consistent with presence of two copies of G2R present in labile inverted terminal repeats (ITRs) of MPXV genome., Conclusions: MPXV is readily detected by qPCR using three clinically validated assays., Competing Interests: Declaration of Competing Interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: ALG reports contract testing from Abbott, Cepheid, Novavax, Pfizer, Janssen and Hologic and research support from Gilead and Merck, outside of the described work. Dr Kachikis reported serving as a research consultant for Pfizer and GlaxoSmithKline on maternal immunization-related projects in 2020 and as an unpaid consultant for GlaxoSmithKline in 2022 outside the submitted work. Dr Kachikis reported receiving grant support from Merck and Pfizer outside the submitted work., (Copyright © 2022 Elsevier B.V. All rights reserved.)

We report intrafamilial transmission of monkeypox virus to all members of a family (father, mother, and 2 children). Case reports in young children have been extremely rare during the 2022 mpox outbreak. Their clinical signs were mild, and clinical diagnosis would be difficult without knowledge of the father's monkeypox virus infection.

Monkeypox virus, a zoonotic Orthopox DNA virus was rarely reported outside of African regions until April 2022. Since then, thousands of cases have been reported worldwide. In order to cope with the increasing need for laboratory diagnosis, the availability of reliable commercial PCR assays is of paramount importance. In this study we compared the diagnostic performance of two commercial real-time (RT)-PCR assays, the Novaplex™ MPXV Assay and the Bio-Speedy® Monkeypox Virus qPCR Kit, for the detection of Monkeypox virus (MPXV) DNA from 154 human samples. These assays were compared to a recently published in-house assay that included a general MPXV target (G2T) and a West African specific target (genericWA). All assays demonstrated 100% specificity. While sensitivity of the Novpalex assay was 100% the sensitivity of the other assays was lower; 94% for the Bio-speedy assay and G2R assay and 88% for the genericWA assay. The sensitivity differences between the methods manifested almost entirely in those pharyngeal samples in which the Ct values were high (≥35). The Novaplex™ MPXV Assay showed higher Ct values compared with the other methods with a median of 27.1 compared with the Bio-Speedy assay (median 15.8, p < 0.001), the G2R assay (median 23.5, p < 0.001) and the genericWA assay (median 23.6, p < 0.001). For all 4 methods, the Ct values were higher in samples taken from oropharynx compared with samples from rectal and pustule swabs., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Competing Interests: We declare no competing interests.

Background: Mpox (formerly monkeypox) is a viral disease caused by the mpox virus (MPXV), endemic in Central and West Africa and currently causing a global outbreak of international concern. Much remains unknown about sample types most suited for mpox laboratory diagnosis. While it is established that high viral loads can be found in active skin lesions (currently the recommended mpox laboratory confirmation specimen type), WHO mpox testing guidelines encourage the use of oropharyngeal swabs as an additional sample type for mpox diagnosis and suggest investigating the value of other specimens like blood samples., Objective: In this study, we verified the value of select alternative specimen types for mpox laboratory confirmation., Methods: We included 25 patients with MPXV-confirmed skin lesions to compare diagnostic sensitivity of MPXV PCR testing on EDTA plasma and two upper respiratory specimens: oropharyngeal swabs and saliva., Results: In our patient cohort with MPXV-confirmed skin lesions, diagnostic sensitivity of MPXV PCR was 80% in EDTA plasma, 64% in oropharyngeal swabs, and 88% in saliva. MPXV viral loads were significantly higher in saliva compared to oropharyngeal swabs and EDTA plasma., Discussion: The WHO recommendation to collect oropharyngeal swabs as an additional specimen for mpox diagnosis might need to be revised to include saliva wherever feasible. We suggest investigating saliva as a diagnostic specimen in the absence of active skin lesions or during the phase preceding skin manifestations. Moreover, the relatively high MPXV DNA content of saliva warrants elucidating its potential role in disease transmission., Competing Interests: Declaration of Competing Interest None, (Copyright © 2022 The Author(s). Published by Elsevier B.V. All rights reserved.)

Monkeypox (mpox) is a zoonotic disease caused by Monkeypox virus (MPXV), an Orthopoxvirus; the wild mammalian reservoir species is not known. There are two genetic clades of MPXV: clade I and clade II (historically found in central and west Africa, respectively), with only Cameroon reporting both clades (1). Human cases have historically been reported from 1) mostly rural, forested areas in some central and west African countries; 2) countries reporting cases related to population migration or travel of infected persons; and 3) exposure to imported infected mammals (2). The annual number of cases in Africa has risen since 2014 and cumulatively surpassed reports from the previous 40 years for most countries. This reemergence of mpox might be due to a combination of environmental and ecological changes, animal or human movement, the cessation of routine smallpox vaccination since its eradication in 1980, improvements in disease detection and diagnosis, and genetic changes in the virus (2). This report describes the epidemiology of mpox since 1970 and during 2018-2021, using data from national surveillance programs, World Health Organization (WHO) bulletins, and case reports, and addresses current diagnostic and treatment challenges in countries with endemic disease. During 2018-2021, human cases were recognized and confirmed in six African countries, with most detected in the Democratic Republic of the Congo (DRC) and Nigeria. The reemergence and increase in cases resulted in its being listed in 2019 as a priority disease for immediate and routine reporting through the Integrated Disease Surveillance and Response strategy in the WHO African region.* In eight instances, patients with mpox were identified in four countries outside of Africa after travel from Nigeria. Since 2018, introductory and intermediate training courses on prevention and control of mpox for public health and health care providers have been available online at OpenWHO. † , § The global outbreak that began in May 2022 ¶ has further highlighted the need for improvements in laboratory-based surveillance and access to treatments and vaccines to prevent and contain the infection, including in areas of Africa with endemic mpox., Competing Interests: All authors have completed and submitted the International Committee of Medical Journal Editors form for disclosure of potential conflicts of interest. No potential conflicts of interest were disclosed.