![Ct values and its temporal trend curves by specimen types; the dots represent the Ct value and the viability of mpox virus at the time of viral culture [no viable virus (blue square), viable virus (red circle), not known (black cross)]; the first dotted vertical line (purple) indicates the estimated day of slope change, while the second one indicates the day of ending viable virus isolation; dotted horizontal line (red) is the optimal Ct cutoff for determining virus viability.](https://www.researchgate.net/profile/Masoud-Rahmati/publication/373139554/figure/fig2/AS:11431281188962680@1694818147056/Ct-values-and-its-temporal-trend-curves-by-specimen-types-the-dots-represent-the-Ct_Q320.jpg)
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© International Society of Travel Medicine 2023. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com
Journal of Travel Medicine, 2023, 1–10
https://doi.org/10.1093/jtm/taad111
Systematic Reviews
Systematic Reviews
Viral load dynamics and shedding kinetics of mpox
infection: a systematic review and meta-analysis
Hakyoung Kim, MD1,Rosie Kwon, MS2,3, Hojae Lee, MS2,3,
Seung Won Lee, MD, PhD4,Masoud Rahmati, PhD5,6,Ai Koyanagi, MD, PhD7,
Lee Smith, PhD8,Min Seo Kim, MD, PhD9, Guillermo F. López Sánchez, PhD10,
Dragioti Elena, PhD11 , 1 2,Seung Geun Yeo, MD, PhD13,Jae Il Shin , MD, PhD14,15,
Wonyoung Cho, PhD2and Dong Keon Yon , MD, FAAAAI, FACAAI, PhD2,3,16,*
1Department of Medicine, Kyung Hee University College of Medicine, Seoul 02447, South Korea,2Center for Digital
Health, Medical Science Research Institute, Kyung Hee University Medical Center, Kyung Hee University College of
Medicine, Seoul 02447, South Korea,3Department of Regulatory Science, Kyung Hee University, Seoul 02447, South
Korea,4Department of Precision Medicine, Sungkyunkwan University School of Medicine, Suwon 16419, South Korea,
5Department of Physical Education and Sport Sciences, Faculty of Literature and Human Sciences, Lorestan University,
Khoramabad 6815144316, Iran, 6Department of Physical Education and Sport Sciences, Faculty of Literature and
Humanities, Vali-E-Asr University of Rafsanjan, Rafsanjan 7718897111, Iran,7Research and Development Unit, Parc
Sanitari Sant Joan de Deu, Barcelona 08830, Spain,8Centre for Health, Performance and Wellbeing, Anglia Ruskin
University, Cambridge CB1 1PT, UK,9Cardiovascular Disease Initiative, Broad Institute of MIT and Harvard, Cambridge,
MA 02142, USA,10Division of Preventive Medicine and Public Health, Department of Public Health Sciences, School of
Medicine, University of Murcia, Murcia 30120, Spain,11Department of Medical and Health Sciences, Pain and
Rehabilitation Centre, Linköping University, Linköping 581 83, Sweden,12Research Laboratory Psychology of Patients,
Families, and Health Professionals, Department of Nursing, School of Health Sciences, University of Ioannina, Ioannina
45221, Greece, 13Department of Otolaryngology - Head & Neck Surgery, Kyung Hee University Medical Center, Kyung Hee
University College of Medicine, Seoul 02447, South Korea,14Department of Pediatrics, Yonsei University College of
Medicine, Seoul 03722, South Korea,15Severance Underwood Meta-research Center, Institute of Convergence Science,
Yonsei University, Seoul 03722, South Korea and 16Department of Pediatrics, Kyung Hee University Medical Center, Kyung
Hee University College of Medicine, Seoul 02447, South Korea
*To whom correspondence should be addressed. Tel: +82-2-228-2050; Fax: +82-2-393-9118; Email: yonkkang@gmail.com
Submitted 11 June 2023; Revised 28 July 2023; Editorial Decision 9 August 2023; Accepted 9 August 2023
Abstract
Background: Viral load dynamics and shedding kinetics are critical factors for studying infectious diseases.
However, evidence on the viral dynamics of mpox remains limited and inconclusive. Thus, we aimed to provide
a comprehensive understanding of the viral load and viability of the re-emerged mpox virus since 2022.
Methods: For this systematic review and meta-analysis, we searched PubMed/MEDLINE, Embase and Google
Scholar for published articles that are related to mpox viral dynamics up to April 2023.
Results: From 19 studies, 880 samples and 1477 specimens were collected. The pooled median Ct values appeared
in the following order: skin lesion [Ct value 21.7 (IQR 17.8–25.5)], anorectal [22.3 (16.9–27.6)], saliva [25.9 (22.5–31.1)],
oral [29.0 (24.5–32.8)], semen [29.6 (25.9–33.4)], urine [30.5 (24.6–36.4)], pharyngeal [31.9 (26.5–37.3)], urethra [33.0
(28.0–35.0)] and blood [33.2 (30.4–36.1)]. People living with human immunodeficiency virus (HIV) have a lower Ct
value in the skin [skin HIV+, 19.2 (18.3–20.0) vs skin HIV−, 25.4 (21.2–29.0)]. From the Ct values and test day since
symptom onset, we identified temporal trends of viral load for each specimen type. Changes in the trend were
observed at 4 days in saliva, 5 days in blood, 6 days in skin, 7 days in anorectal, urine, semen and pharyngeal
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2Journal of Travel Medicine, 2023, Vol. 30, 5
and 8 days in the urethra. We determined optimal Ct cutoff values for anorectal (34.0), saliva (27.7) and urethra
(33.0) specimens, where a Ct value above each cutoff suggests minimal viral viability. Using these cutoff values, we
derived the duration of viable viral isolation in each specific specimen type (anorectal 19 days, saliva 14 days and
urethra 14 days).
Conclusion: Skin lesion, anorectal and saliva samples contained the highest viral load. The peak viral load manifests
within 4–8 days after symptom onset, and viable virus detection was presumed to cease within 14–19 days from
symptom onset in anorectal, saliva and urethral samples.
Key words: Mpox, virus, viral load, viral dynamics
Introduction
Although initially recognized as an endemic disease limited to
certain areas of Africa,1,2the global outbreaks of mpox have been
unprecedented since 2022.3,4This outbreak exhibits unique char-
acteristics in terms of severity, sex distribution and symptoms,
including a notable transmission route through sexual contact
among men who have sex with men (MSM).5Based on recent
genomic analysis, the current outbreak is more likely caused by
superspreading events with a single infection source, wherein
infected individuals had extensive contacts within a short period,
combined with global travel patterns.6–8The low genomic vari-
ability of circulating strains supports this hypothesis.6–8
Several studies have investigated the viral dynamics of mpox.
Quantitative polymerase chain reaction (PCR) was commonly
used to measure viral loads in various specimens, and some
studies also employed viral culture to confirm viability.9The
patterns and duration of viral clearance were assessed. Although
viral load or culture positivity does not directly correlate with
mpox infectivity,10 these factors are considered as indicators
when estimating viral infectivity.11,12
While advancements have been made in developing more
sensitive and specific assays for measuring viral load, previous
studies have been limited in their focus on specific sites and
small sample sizes. Therefore, in this systematic review and meta-
analysis, we aimed to provide a comprehensive understanding
of the viral load, viability, transmission and shedding dynamics
of the mpox virus. By examining specific site characteristics and
viral shedding patterns, we proposed a plausible mechanism and
suggested reference values for the development of policies aimed
at preventing and controlling the outbreak of mpox.
Methods
For this systematic review and meta-analysis, articles on
PubMed/MEDLINE, Embase and Google Scholar were searched
using the keywords ((“monkeypox” or “MPX” or “MPOX”)
and (“DNA” or (“viral” and (“load” or “shedding” or
“dynamics” or “kinetics”)))) up to 21 April 2023. This study
included cohort studies, cross-sectional surveys, case series
and case reports that depicted the clinical features of mpox
and the viral load for each specimen type. The selection
process of the studies included in our systematic review
and meta-analysis is illustrated in the Preferred Reporting
Items for Systematic Reviews and Meta-Analysis (PRISMA)
2020 diagram.13 Our systematic review and meta-analysis
protocol were registered with PROSPERO (registration number
CRD42023421311).
Literature search strategy
Two independent researchers (HK and WC) conducted separate
searches and reviews of the titles, abstracts and full texts of
published studies in accordance with the PRISMA 2020 guide-
lines.13 Another researcher (DKY) participated and confirmed
any disagreements in screening.
We evaluated all the existing related observational stud-
ies, including retrospective or prospective cohort studies, cross-
sectional studies, case series and case reports. Duplicate records,
review articles, meta-analyses and studies that omitted the Ct
value or reported it in a different unit were excluded. The study
selection process according to the PRISMA 2020 guidelines is
summarized in Figure S1.
Eligibility checkpoints
This systematic review and meta-analysis collected and analysed
cohort studies, cross-sectional studies and case-series/reports that
included the following characteristics: age, sex, men who had
MSM, history of human immunodeficiency virus (HIV) infection,
smallpox vaccination status and viral load of mpox virus in
different types of specimens. Specimens were collected from the
skin, anus and rectum, saliva, mouth, urine, semen, pharynx
(oropharynx or nasopharynx), urethra and blood.
Mpox viral load was inferred from the PCR cycle threshold
(Ct) value, which indicates the number of PCR cycles required to
obtain a positive result. The Ct value is approximately inversely
proportional to the quantity of viral DNA. Generally, a lower Ct
value indicates a higher viral load, as there is a higher quantity
of viral DNA in the specimen. We referred to the trend in viral
load as changes in Ct values over a period of time and analysed
the daily Ct value of the subjects from the onset of symptoms to
track the temporal trend of viral load in each type of specimen.
Data extraction and analysis
We extracted the following data from each of the collected
studies: first author, publication year, country, sample size, study
date, study design (cohort, cross-sectional, case series and case
report), characteristics (age, sex, MSM, history of HIV infection,
smallpox vaccination status, assay type and DNA extraction
tools), test day since symptom onset and mpox viral load.
The median and interquartile ranges were recalculated by
combining the Ct values and days since symptom onset for each
specimen in the included studies.14 To identify non-linear trends
in Ct values over time, we employed local polynomial regression,
a non-parametric statistical method.15 This technique involves
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Journal of Travel Medicine, 2023, Vol. 30, 5 3
estimating the relationship between the variables, Ct values
and days since symptom onset by fitting separate polynomial
functions to local cluster of data points. The size of each cluster
or neighbourhood, was determined by a chosen window width
or fraction of the data, which dictated the number of nearby data
points used in the estimation process. In our study, we selected
a fraction of two-thirds. Subsequently, we conducted weighted
linear least squares regression on each subset, systematically
moving through the data point by point. This iterative process
was repeated until each individual data point had been accounted
for.
We applied a mathematical method, called an optimization
algorithm technique, to track the optimal Ct value to classify
viability from the Ct values of specimens with confirmed viral
culture. We iteratively scanned the Ct value as a variable and
found the point where the accuracy in the confusion matrix was
maximized. This specific Ct value was then defined as the Ct
cut-off value. Additionally, we examined the receiver operating
characteristic curve (ROC) curve to observe how sensitivity and
specificity values changed with varying Ct values. The area
under the ROC curve (AUC) was calculated for providing an
indication of the model’s ability to differentiate between negative
and positive groups.
We defined the day of ending isolation as the point at which
virus viability was stochastically low at the intersection of the Ct
cut-off value and the Ct value curve over time. We assumed that
this marked reduction in viral viability corresponds to a notable
decrease in the probability of transmitting the virus to others.
All statistical analyses and generated figures were performed
using Excel (version 2021; Microsoft, Redmond, WA, USA)
and Python 3 (version 3.10.6; Python Software Foundation,
Wilmington, DE, USA), with Pandas (version 1.5.1), Statsmodels
(version 0.14.0), Scikit-learn (version 1.0.2), Seaborn (version
0.12.1) and Matplotlib (version 3.6.0).
Assessment of the methodological quality of
included studies
For each cross-sectional and cohort study, the adapted Newcas-
tle–Ottawa scale was utilized for quality assessment in Tabl e S 1.5
The Joanna Briggs quality evaluation instrument was reviewed
for each case report and series in Tab l e S 2.5
Patient and public involvement
No patients were engaged in establishing the research question or
outcome measures, nor in developing plans for the study’s design
or implementation. No patients were asked to provide feedback
on the interpretation or writing of the results. Nonetheless, if
requested, we intend to convey the findings of the study to
relevant groups.
Results
Study characteristics
Our reference studies had three study designs: 68.4% case-
series/report (13/19), 26.3% cross-sectional studies (5/19) and
5.3% cohort study (1/19). The studies were conducted in eight
countries (Italy, Spain, Israel, France, Australia, Belgium, USA
and India) across four continents (Europe, Asia, Australia and
North America) all within or during 2022 (Tab l e s 1 and 2). All
studies used real-time PCR (RT-PCR) as a mean of viral load
assay.
The 19 reference studies included 880 samples and 1477 spec-
imens, participants consisted of 559 males (63.5%), 11 females
(1.3%) and 310 unknown (35.2%). A total of 441 participants
(94.0%) out of 469 respondents were MSM, 192 participants
(37.8%) out of 508 respondents were living with HIV and 64
participants (18.1%) out of 353 respondents had a history of
smallpox vaccination. The median age of the participants was
36.1 years [standard deviation (SD) 2.1 years]. Their clinical
presentations are depicted in the table, with rash being the most
common (95.8%) symptom (Tab l e 2 ).
Key findings
We presented the mpox viral load and viability of the samples
from case reports, classified by specimen types. As the lower Ct
value was related to higher viral DNA in the specimen, the pooled
median Ct values and test day since symptom onset appeared
in the following order in the individuals: skin lesion [Ct value
21.7 (IQR 17.8–25.5); test day since symptom onset 6.0 (IQR
4.0–10.0)], anorectal [22.3 (16.9–27.6); 7.0 (5.0–12.0)], saliva
[25.9 (22.5–31.1); 6.0 (4.0–8.0)], oral [29.0 (24.5–32.8); 3.5
(3.3–3.8)], semen [29.6 (25.9–33.4); 8.0 (6.0–12.8)], urine [30.5
(24.6–36.4); 6.0 (4.0–10.0)], pharyngeal [31.9 (26.5–37.3); 6.0
(4.0–9.0)], urethra [33.0 (28.0–35.0); 8.0 (6.5–13.0)] and blood
[33.2 (30.4–36.1); 5.0 (4.0–8.0)] (Tab l e 3 ).
The skin lesion, anorectal and saliva contained the most viral
DNA and the specimens taken from the pharyngeal, urethra and
blood had relatively low levels of viral DNA. Among samples
with information on viability, samples from anorectal (27 out of
37) and saliva (23 out of 40) had a higher yield of viable virus.
By contrast, the virus contained in pharyngeal samples (4 out of
53) was mostly non-viable (Tabl e 3 and Figure 1).
From the Ct values and test day since symptom onset, we
found temporal trends of viral load for each specimen. The
changes in the trend of viral load were observed over a period
of 6 days in skin, 7 days in anorectal, 4 days in saliva, 7 days
in urine, 7 days in semen, 7 days in pharyngeal, 8 days in
urethra and 5 days in blood (Figure S2 to S4). No significant
differences were observed in the temporal trends of any specimen
across different age groups (Tab l e S 3 and Figure S5). A compar-
ison of viral shedding between HIV-positive and HIV-negative
individuals across various specimens revealed notable disparity
only in the skin samples (Tabl e S 4 and Figure S6). Skin samples
from people with HIV showed significantly low Ct value [19.2
(18.3–20.0)] compared to those from people without HIV [25.4
(21.2–29.0)].
We established the optimal Ct cutoff values for anorectal
(34.0), saliva (27.7) and urethra samples (33.0) based on the
relationship between Ct values and viral culture. Samples with
Ct values exceeding these cutoffs were deemed unlikely to yield
successful culture results. Subsequently, we calculated the dura-
tion of isolation required for each specimen type, resulting in
19.0 days for anorectal samples, 14.0 days for saliva samples
and 14.0 days for urethra samples.
Different types of specimens collected showed different
trends in viral load. In the curve of skin samples, the Ct value
remained below 20 for ∼6 days before the curve gradually
declined to around 30 by Day 20. Conversely, the anorectal curve
exhibited a lower overall viral load compared to skin samples
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4Journal of Travel Medicine, 2023, Vol. 30, 5
Ta b l e 1 . Summary of included studies
Author, year Country Sample, nStudy date Study design Number of
specimen types
Specimen type
Allan-Blitz et al., 202223 USA 22 2022 Case-series 1 Saliva
Antinori et al., 202234 Italy 42022.05.17–2022.05.22 Case-series 6 NPS, skin, anorectal, blood,
saliva, semen
Baetselier et al., 202235 Belgium 42022.05 Cross-sectional 1Anorectal
Colavita et al., 202336 Italy 32022 Case-series 5 OPS, anorectal, blood, saliva,
semen
Coppens et al., 202322 Belgium 25 2022 Case-series 3 OPS, blood, saliva
Elbaz et al., 202337 Israel 215 2022.01.01–2022.08.10 Cross-sectional 3OPS, skin, anorectal
Gaspari et al., 202238 Italy 30 2022.06.20–2022.08.10 Case-series 5 OPS, skin, anorectal, blood, urine
Hernaez et al., 202324 Spain 44 2022.06.07–2022.06.26 Cross-sectional 2OPS, saliva
Lapa et al., 202239 Italy 12022.05 Case report 3Skin, blood, semen
Lim et al., 202340 Australia 70 2022.05.19–2022.10.19 Cross-sectional 6NPS, skin, anorectal, blood,
urine, oral
Loconsole et al., 202225 Italy 10 2022.06.01–2022.08.01 Case-series 3 NPS, skin, blood,
Mileto et al., 202241 Italy 12022.05.24 Case report 5OPS, skin, anorectal, blood,
semen
Moschese et al., 202242 Italy 33 2022.05–2022.07 Case-series 2 Anorectal, urethra
Ouafi et al., 202343 France 138 2022.05.23–2022.08.18 Cross-sectional 3OPS, skin, anorectal
Palich et al., 202344 France 50 2022.05.20–2022.06.13 Case-series 6 PSa, skin, anorectal, blood, urine,
semen
Paran et al., 202245 Israel 32 2022 Case-series 3 OPS, skin, anorectal
Peiro-Mestres et al., 202246 Spain 12 2022.05–2022.06 Case-series 7 NPS, skin, anorectal, urine,
saliva, semen, oral
Relhan et al., 202347 India 5 2022.06.23–2022.08.12 Case-series 5 OPS, NPS, skin, blood, urine
Tarín-Vicente et al., 202230 Spain 181 2022.05.11–2022.06.29 Cohort 3OPS, skin, anorectal
NPS, nasopharyngeal; OPS, oropharyngeal; PS, pharyngeal.
aIn this study, pharyngeal referred to both oropharyngeal and nasopharyngeal.
and demonstrated a decrease after reaching its peak on Day 7.
The saliva curve maintained a relatively constant Ct value of
around 25, with a small peak observed on Day 4. Both urine and
semen samples showed peak Ct values of around 28 on Day 7.
The pharyngeal and urethra curves remained relatively stable,
ranging between Ct values of 30–35, with minimal changes until
Day 20. The blood curve followed a similar Ct value range but
wasplottedupto∼10 days (Figure 2).
Discussion
Findings of our study
Our results showed that mpox viral load was found to be high
in skin lesion, anorectal and saliva and low in semen, urine,
pharynx, urethra and blood, in decreasing order. While there
are limited data on viability in skin lesion samples, a substantial
portion of samples from both the anorectal and saliva sources
were identified to contain viable viruses.
Consistent with our analysis, previous reports have identified
skin, anorectum and saliva as the primary sources of mpox
transmission. Given that anogenital lesions were reported to be
the most common location of mucocutaneous lesions16,17 and
that many isolated lesions presented at the exact site of sexual
contact,18 these findings strengthen the idea that local contact,
encompassing intimate contact in sexual intercourse, anal or oral
sex and kissing, was the main mode of transmission in the 2022
outbreak.
No case or evidence supporting the transmission through
urine, semen, urethra or blood has been reported thus far.19
Nevertheless, the potential for sufficient infection through urine
or semen remains plausible. There were research findings of
viable virus isolation in seminal fluid,20 and data of viable virus
isolated from urine samples were also included in our data.
Notably, the diagnostic value of the pharyngeal specimen
seemed to be lower than previously understood. The median
Ct value for oropharyngeal specimens was 32.4, which is close
to the minimum of detection, considering that the US Centers
for Disease Control and Prevention recommend immediate re-
extraction and re-testing for mpox specimen samples with a Ct
value of ∼34 or above.21 In addition, among 53 oropharyngeal
specimens with information on viability, 49 (92.5%) were found
to be non-viable. These results suggest that oropharyngeal spec-
imens may contribute little to diagnostic accuracy and raise the
need for a revision of the WHO recommendation that oropha-
ryngeal swabs be taken as an additional test specimen along with
skin lesion swabs.22 Blood samples showed the highest Ct values,
which was consistent with the WHO recommendation that blood
samples were not suitable for diagnosing mpox.
We outlined the temporal profile of viral load across different
specimen types using local polynomial regression and observed
that peak viral load typically occurs within 4–8 days after symp-
tom onset, followed by a gradual decline. This being established,
infectivity was presumed to be at its highest within the first
week after symptom occurrence. The viral load of the skin lesion
exhibited a plateau during the early phase of infection, aligning
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Journal of Travel Medicine, 2023, Vol. 30, 5 5
Ta b l e 2 . Baseline characteristics of the reference studies
Characteristics Included study, k(%) Sample, n(%) Specimen, n(%)
Tot a l 19 (100) 880 (100) 1477 (100)
Age, mean year (SD) 14 (73.7) 36.1 [2.1]
Sex
Male 16 (84.2) 559 (63.5)
Female 16 (84.2) 11 (1.3)
Not known 3 (15.8) 310 (36.8)
MSMa13 (68.4) 441/469 (94.0)
People living with HIVa11 (57.9) 192/508 (37.8)
Smallpox vaccinationa8 (42.1) 64/353 (18.1)
Assay type
RT-PCR 19 (100) 880 (100) 1477 (100)
DNA extraction
Qiagen 8 (42.1) 150 (17.0) 352 (23.8)
BioMérieux 2 (10.5) 265 (30.1) 246 (16.7)
Roche 2 (10.5) 150 (17.0) 314 (21.3)
Promega 1 (5.3) 44 (3.8) 86 (5.8)
RealStar 1 (5.3) 33 (2.5) 64 (4.3)
Thermo Fisher Scientific 1 (5.3) 22 (2.6) 15 (1.0)
Abbott Molecular 1 (5.3) 4 (0.5) 4 (0.3)
ELITechGroup 1 (5.3) 1 (0.1) 14 (0.9)
Not recorded 2 (10.5) 211 (24.0) 382 (25.9)
Country
Italy 7 (36.8) 82 (9.3) 223 (15.1)
Spain 3 (15.8) 237 (26.9) 486 (32.9)
Israel 2 (10.5) 247 (28.1) 144 (9.7)
France 2 (10.5) 188 (21.4) 362 (24.5)
Belgium 2 (10.5) 29 (3.3) 62 (4.2)
Australia 1 (5.3) 70 (8.0) 144 (9.7)
USA 1 (5.3) 22 (2.5) 15 (1.0)
India 1 (5.3) 5 (0.6) 41 (2.8)
Study design
Case-series/report 13 (68.4) 228 (25.9) 622 (42.1)
Cross-sectional 5 (26.3) 471 (53.5) 552 (37.4)
Cohort 1 (5.3) 181 (20.6) 303 (20.5)
Clinical presentationsa
Rash 11 (57.9) 345/360 (95.8)
Fever 9 (47.4) 93/169 (55.0)
Lymphadenopathy 7 (36.8) 211/315 (67.0)
Fatigue 7 (36.8) 83/141 (58.9)
Myalgia 7 (36.8) 85/167 (50.9)
Headache 5 (26.3) 60/133 (45.1)
Eating disorder 4 (21.1) 24/57 (42.1)
Proctitis 4 (21.1) 82/287 (28.6)
Sore throat 4 (21.1) 20/121 (16.5)
Rectal pain 3 (15.8) 2/13 (15.4)
Pharyngitis 3 (15.8) 28/232 (12.1)
Genital swelling 2 (10.5) 17/186 (9.1)
Malaise 2 (10.5) 8/13 (61.5)
Back pain 2 (10.5) 2/30 (6.7)
Chills sweats 1 (5.3) 9/22 (40.9)
Dysuria 1 (5.3) 2/5 (40.0)
Chest pain 1 (5.3) 1/5 (20.0)
Diarrhoea 1 (5.3) 1/22 (4.5)
MSM, men who sex with men; RT-PCR, real-time polymerase chain reaction.
aSubjects who responded or had obvious medical record were counted only.
with previous reports that skin lesions are among the initial
symptoms. Following the peak, both skin lesion and anorectal
specimen viral loads displayed a steep decrease, suggesting a
rapid decline in infectivity from these sources after ∼6and
7 days, respectively. These findings underscored the importance
of implementing timely preventive measures and interventions,
within a week of symptom occurrence, to curb the transmission
of the virus.
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6Journal of Travel Medicine, 2023, Vol. 30, 5
Ta b l e 3 . Ct value and median test day since symptom onset by specimen types
Specimen
type
Included
studies
Total
specimen
Number of
viable/non-viable virusa
Test day since symptom
onset (IQR)b
Ct value Days of viral
load changea
Days of ending
isolationa
Median (IQR) Cut-off
Tot a l 19 1477 79/150
Skin 13 575 3/- 6.0 (4.0–10.0) 21.7 (17.8–25.5) 6.0
Anorectal 13 243 27/10 7.0 (5.0–12.0) 22.3 (16.9–27.6) 34.0 7.0 19.0
Saliva 6101 23/17 6.0 (4.0–8.0) 25.9 (22.5–31.1) 27.7 4.0 14.0
Oral 2 14 -3.5 (3.3–3.8) 29.0 (24.5–32.8)
Urine 548 -6.0 (4.0–10.0) 30.5 (24.6–36.4) 7.0
Semen 6 43 1/5 8.0 (6.0–12.8) 29.6 (25.9–33.4) 7.0
Pharyngeal 15 353 4/49 6.0 (4.0–9.0) 31.9 (26.5–37.3) 7.0
OPS 10 273 4/49 5.0 (4.0–8.0) 32.4 (27.9–36.9) 7.0
NPS 444 -8.5 (6.0–12.0) 32.4 (29.9–36.1)
Urethra 131 17/14 8.0 (6.5–13.0) 33.0 (28.0–35.0) 33.0 8.0 14.0
Blood 10 69 -/6 5.0 (4.0–8.0) 33.2 (30.4–36.1) 5.0
IQR, interquartile range.
aCalculations were based on the information available in the included studies. bGiven as a median value with an interquartile range.
Saliva demonstrated an earlier and more pronounced peak in
viral load compared to blood. What was particularly intriguing
was that, unlike other specimen types, the majority of saliva
samples containing viable viruses were collected within a week
of symptom onset. As no viable virus had been detected in blood
samples thus far19, it was plausible to propose that the high
viral load observed in saliva is predominantly a result of direct
oral infection through activities like kissing or oral sex, while
shedding of the non-viable viral particles from the bloodstream
to saliva may also play a certain role in contributing to the viral
load. This notion was supported by the detection of positive
saliva PCR results in asymptomatic cases before the appearance
of a rash or skin lesions at the time of testing.23
While the high viral load and presence of viable virus in saliva
suggested the potential for transmission through respiratory
droplets or aerosols, it was unlikely to be the main mechanism
of spread in the 2022 outbreak. One previous study reported a
high proportion of mpox DNA in saliva (85%).24 However, the
study found viable virus in only two out of 45 mask samples
(4.4%) and no viable virus in 44 air filters (0%).25 Furthermore,
there had been no documented cases of human-to-human aerosol
transmission in mpox.
Our evaluation of the mpox virus revealed that pharyngeal
samples, including the oropharynx and nasopharynx, had a
low viral load and a minimal likelihood of viability. Studies
examining the viral load of mpox have found that a notable
percentage of patients (43.6%) only test positive for the virus
in skin lesions.25 Additionally, in most observed cases, the first
lesion to appear was a rash in the genital and perianal areas.
These findings suggested that the respiratory epithelium was not
the primary site of viral replication during the 2022 outbreak.
Therefore, although saliva contains a high viral load with via-
bility, it is less likely that inhaled respiratory droplets play a
significant role as a source of infection and transmission. Instead,
we propose that a viable virus presented in saliva primarily
initiates transmission and infection when it enters non-intact skin
or mucosa through direct contact.
Through the analysis of the relationship between viral load
and the viability of the mpox virus, we proposed an optimal Ct
value cutoff for anorectal, saliva, and urethra for determining
virus viability. Samples exceeding these cutoffs demonstrated
limited success in culture. Combined with this cutoff, we were
able to calculate the duration of viable virus isolation for each
specific specimen type: 19 days for anorectal samples, 14 days for
saliva and 14 days for the urethra. After this duration of culture
viability, based on our assumption, the likelihood of infection
through that particular source is expected to be very low.9,26
Therefore, conducting additional tests to assess infectivity from
that specific source may not be necessary for patients with Ct
values above the cutoff or after the specified duration.
The impact of comorbidity with HIV on mpox infection is
widely recognized, although there is limited evidence supporting
different mpox viral dynamics. Most studies examining this
impact included people with well-controlled HIV and high CD4
counts; however, Mitjà et al. reported the greatest disease sever-
ity, hospitalization and mortality among individuals with low
CD4 counts and high HIV viral load.27 Our results demonstrate
a higher viral load only in skin specimens from people with
HIV, which aligns with previous findings reporting widespread,
large and severe skin lesions in individuals with lower
CD4 counts.27–29
Plausible mechanism
The WHO had designated two major clades of mpox virus: Clade
I (Congo Basin) and Clade II (West African), with the latter
encompassing subclades IIa and IIb.17 The 2022 global outbreak
primarily involved Clade IIb (sublineage: A.2, B.1).17,24 In the
current outbreak, the anogenital area is the most common site
of skin rash presentation.
The genital and anorectal epithelium have a lower level
of keratinization and a higher presence of antigen-presenting
cells like macrophages and dendritic cells compared to other
parts of the skin. Thus, these areas are more susceptible to
pathogen acquisition.17,30 Moreover, possible mucosal abrasion
and damage acquired during sexual intercourse may facilitate
the inoculation of the mpox virus. Consequently, from a clinical
standpoint, the highest transmission rate was observed through
sexual contact, despite the potential for infection in other areas.30
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Journal of Travel Medicine, 2023, Vol. 30, 5 7
Figure 1. Viral load and virus viability of mpox by specimen types; box height, interquartile range; box cap, minimum and maximum; line inside box,
median; white triangle, mean Ct value.
This observation supports the notion that, while mpox could
theoretically be transmitted via respiratory droplets, engaging in
oral sex is likely to pose a higher risk of transmission. Neverthe-
less, further studies are required to support our speculations and
to acquire a fuller understanding of the mode of transmission in
mpox virus.
Policy implications
Despite its initial identification in humans in 1970, the endemic
nature of mpox across several Central and West African
countries has resulted in a limited amount of research on the
virus, particularly regarding the Clade IIb that caused the 2022
outbreak.2,17,31 As mpox continues to spread to new regions via
travel, it might be perceived as a novel disease in each area.
There is an imperative need for political efforts to establish
protocols for diagnosis, treatment and, most importantly,
prevention. Policymakers should be able to decide specimen
selection and collection timing for diagnosis based on current
findings.
Strengths and limitations
To the best of our knowledge, the present systematic review
and meta-analysis is the first to examine the viral load dynam-
ics of mpox comprehensively and numerically across different
specimen types. Our review extended to the temporal profiles,
time-varying viability of the mpox virus and duration of viable
virus isolation. These findings provide indirect insights into
its mode of transmission and carry implications for preventive
measures.
Our study benefits from a large sample size and various speci-
men types, enabling a thorough understanding of viral dynamics.
By establishing the timeframe for peak viral load and the dura-
tion of viable virus isolation for different types of specimens,
our findings can assist in the development of social restriction
protocols for confirmed or suspected individuals, particularly for
international travellers. Furthermore, when selecting specimens
and determining the optimal collection timing for diagnosis, our
research outcomes should be considered as valuable guidance.
Moreover, our analysis explored the relationship between viral
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8Journal of Travel Medicine, 2023, Vol. 30, 5
Figure 2. Ct values and its temporal trend curves by specimen types; the dots represent the Ct value and the viability of mpox virus at the time of viral
culture [no viable virus (blue square), viable virus (red circle), not known (black cross)]; the first dotted vertical line (purple) indicates the estimated
day of slope change, while the second one indicates the day of ending viable virus isolation; dotted horizontal line (red) is the optimal Ct cutoff for
determining virus viability.
load and viability, leading to the identification of an optimal Ct
value cutoff for predicting the presence of viable viruses.
Nevertheless, our study contains several limitations. First, we
have limited specific demographic information for each sample,
and there are also instances of partial disclosure of this informa-
tion. As a result, we were unable to compare the viral dynamics
between subgroups based on factors, such as MSM status, HIV
infection and smallpox vaccination. These differences could be
estimated through references from individual studies. Second,
the available results correlating clinical features with viral load
are insufficient. We can only indirectly compare these findings
with our previous paper. Third, there is limited information on
viable cultures. However, based on the available information,
some assessment of viability can still be made. Moreover, it is
important to consider potential variability between laboratories,
as the Ct value can be influenced by multiple factors beyond viral
load. These factors include sample quality, PCR assay efficiency,
probe design and instrument variability. However, it is worth
noting that real-time PCR technology has evolved throughout
the COVID-19 pandemic, resulting in little variation.32,33 In
addition, we conducted separate analyses for each manufacturer
to ensure comparability between different DNA extraction plat-
forms (Tab l e S 5 and Figure S7). Nevertheless, the lack of clear
standardization of procedures across laboratories may introduce
bias in Ct values favouring studies with larger sample sizes. To
improve future studies, it would be advantageous to establish
guidelines for the standardization process between laboratories.
Fourth, due to the limitations of literature so far, only 5%
of cohort study was included in our study. Finally, the direct
correlation between mpox infectivity and viral load or culture
viability remains uncertain. The clinical significance of our find-
ings and their implications are based on inferences, and further
evaluations through prospective studies are necessary.
Conclusion
The present systematic review and meta-analysis offer an insight
into the viral dynamics of the mpox virus. This work provides
clinicians and researchers with essential reference values, such as
Ct value cutoffs and days of ending isolation, to aid in managing
precautions, guiding the selection of diagnostic specimens and
conducting follow-up observations.
Supplementary data
Supplementary data are available at JTM online.
Funding
This research was supported by grants from the Korea
Health Technology R&D Project through the Korea Health
Industry Development Institute (KHIDI), funded by the Ministry
of Health and Welfare, Republic of Korea (grant number:
HV22C0233), from the National Research Foundation of Korea
(NRF; grant number: RS-2023-00248157) and from the Min-
istry of Food and Drug Safety (grant number: 21153MFDS601)
in 2023. The funders played no role in the study design, data
collection, analysis, interpretation, or writing of the manuscript.
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Journal of Travel Medicine, 2023, Vol. 30, 5 9
Acknowledgements
None.
Author contribution statement
D.K.Y. and W.C. had full access to all the data in the study and
took responsibility for the integrity of the data and the accuracy
of the data analysis. All authors approved the final version
before submission. Study concept and design: W.C. and D.K.Y.;
acquisition, analysis or interpretation of data: W.C. and D.K.Y.;
drafting of the manuscript: H.K. and W.C.; critical revision of
the manuscript for important intellectual content: all authors;
statistical analysis: W.C. and D.K.Y. and study supervision: J.I.S.,
W.C. and D.K.Y. D.K.Y. supervised the study and is guarantor for
this study. D.K.Y. is a senior author. J.I.S., W.C., and D.K.Y con-
tributed equally as co-corresponding authors. The corresponding
author attests that all listed authors meet authorship criteria and
that no others meeting the criteria have been omitted.
Authors’ contributions
Hakyoung Kim (Conceptualization, Formal analysis, Investiga-
tion, Writing—original draft [equal]), Rosie Kwon (Writing—
review & editing [equal]), Hojae Lee (Writing—review & editing
[equal]), Seung Won Lee (Writing—review & editing [equal]),
Masoud Rahmati (Writing—review & editing [equal]), Ai Koy-
anagi (Writing—review & editing [equal]), Lee Smith (Writing—
review & editing [equal]), Min Seo Kim (Writing—review &
editing [equal]), Guillermo F. López Sánchez (Writing—review
& editing [equal]), Dragioti Elena (Writing—review & editing
[equal]), Seung Geun Yeo (Writing—review & editing [equal]),
Jae Il Shin (Supervision [equal]), Wonyoung Cho (Writing—
review & editing [equal]), and Dong Keon Yon (Conceptualiza-
tion, Data curation, Formal analysis, Funding acquisition, Inves-
tigation, Methodology, Project administration, Resources, Soft-
ware, Supervision, Validation, Visualization, Writing—original
draft, Writing—review & editing [equal])
Conflict of interest: None declared.
Data availability
All data are provided in the article and in the appendix. The study
protocol, statistical code and data set are available from DKY
(Email: yonkkang@gmail.com).
Ethics statement
This systematic review article does not require Institutional
Review Board approval. Our systematic review and meta-anal-
ysis protocol was registered with PROSPERO (registration num-
ber CRD42023421311).
Informed consent
Not applicable.
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