An Overview of Monkeypox Virus Detection in Different Clinical Samples and Analysis of Temporal Viral Load Dynamics

Abstract

Mpox is a zoonotic disease caused by the Monkeypox virus (MPXV), and since May 2022, tens of thousands of cases have been reported in non-endemic countries. We aimed to evaluate the suitability of different sample types for mpox diagnostic and assess the temporal dynamics of viral load. We evaluated 1914 samples from 953 laboratory-confirmed cases. The positivity rate was higher for lesion (91.3%) and rectal swabs (86.1%) when compared with oropharyngeal swabs (69.5%) and urines (41.2%), indicating higher viral loads for the former. Supporting this, lesion and rectal swabs showed lower median PCR C_t values (C_t = 23 and C_t = 24), compared to oropharyngeal swabs and urines (C_t = 31). Stable MPXV loads were observed in swabs from lesions up to 30 days after symptoms onset, contrasting with a considerable decrease in viral load in rectal and oropharyngeal swabs. Overall, these results point to lesion swabs as the most suitable samples for detecting MPXV in the 2022-2023 multicountry outbreak and show comparable accuracy to rectal swabs up to 8 days after symptoms onset. These findings, together with the observation that about 5% of patients were diagnosed through oropharyngeal swabs while having negative lesions, suggest that multisite testing should be performed to increase diagnostic sensitivity.

Keywords: Ct values; Monkeypox virus; Portugal; clinical samples; positive rate; viral clearance; viral load.

Figure 1

Distribution of the number of samples analyzed and confirmed cases between May 17, 2022, and May 24, 2023.

Figure 2

(A) Positive rate of MPXV detection according to the type of samples from the 953 confirmed cases. (B) MPXV viral loads are given as C _t values, according to the type of samples. For this analysis, follow‐up positive samples were also considered. Results are presented as Violin plots, with median and interquartile ranges represented. The p values refer to the comparison between being MPXV positive or negative by type of sample. The indeterminate sample number was not considered. p values are accompanied by asterisks, depending on the degree of significance (i.e., from one asterisk for p values less than 0.05 to four asterisks for p values less than 0.0001).

Figure 3

Median time from symptoms onset until MPXV diagnostic according to sample type (A) and PCR C _t values (B–E). (B–E) Lesion swabs (n = 611), oropharyngeal swabs (n = 352), rectal swabs (n = 47), and urine (n = 10), respectively. Results are presented as box and whisker plots, with median and interquartile ranges represented in boxes, and range presented as whiskers. Dots at the end of the whiskers represent 5% of the data. C _t values are represented in the axis in decreasing order for better interpretation purposes.

Figure 4

Time to viral clearance in all patients. The dashed line represents the cumulative incidence calculated on the observed data. Parameters a, b, and c for each type of sample are shown.

Figure 5

PCR C _t values for MPXV in several lesion swabs (≥ two samples) from the same patient throughout the infection period since symptoms onset (Day 1) (n = 19). Grey lines connect samples from the same patient with three or more lesion swab samples (n = 6). The black continuous line estimates the time course of infection for visualization. The blue dashed line projects the curve before and after available data. The grey shadow indicates a 95% confidence interval. C _t values are represented in the C _t axis inverted for better interpretation purposes.

Figure 6

MPXV median C _t values of paired samples from different types, collected on the same day from the same patient, throughout the infection period since symptoms onset. Markers indicate PCR C _t values of lesion swabs (blue dots), oropharyngeal swabs (green squares), rectal swabs (red triangles) and urine samples (orange hexagons). (A1–F1) Black lines link PCR C _t values from different types of samples collected on the same day from the same patient. (A2)–(F2) represents the data of the (A1)–(F1) panels throughout the infection period since symptoms onset. Median C _t values with the upper and lower limits are displayed. The y‐axis scale is inverted for better interpretation purposes, as lower C _t values represent higher viral loads. p values are accompanied by asterisks, depending on the degree of significance (i.e., from one asterisk for p values less than 0.05 to four asterisks for p values less than 0.0001).