12. Sampling and Diagnosis of Mpox virus
The PCR test can detect the monkeypox virus (MPXV), which causes
monkeypox. Personal protective equipment (PPE) must be worn while
working within 1 metre of a suspected monkeypox case in order to secure
the safety of healthcare workers. This includes droplet and contact
precautions, as well as eye protection, if spills are possible during
diagnostic procedures such as pharynx biopsies or lesion samples. When
gathering a sample for diagnostic testing, it is essential to obtain a
viral biopsy from an exposed lesion or the surface of a vesicle (a small
fluid-filled abscess). If there are multiple incisions, it is crucial
that the sample be obtained from a vesicle, an ulcer, or a vesicle with
a crust. Rub the probe gently over the lesion and position it in a tube
containing viral culture medium or viral transport medium. Additionally,
a pharynx sample should be taken if pharyngeal lesions are present. As
unlabeled tubes cannot be processed, designate the collection tube with
the patient’s name, date of birth, and location of the sample. When
high-risk contacts of a confirmed or highly probable case of monkeypox
develop systemic symptoms but lack a rash or lesions suitable for
sampling, a viral pharynx swab can be taken. Notably, even if the throat
biopsy yields a negative result, individuals must continue to monitor
their symptoms, adhere to isolation protocols as directed by their local
health protection team, and be reevaluated and sampled if additional
symptoms develop. It is crucial to package the samples separately when
investigating other infections, such as sexually transmitted infections.
Samples should ideally be sent to a nearby laboratory for examination.
However, if MPXV testing is not available locally, the laboratory should
send the samples to the other advance laboratory for MPXV PCR, while
continuing to follow their normal local procedures for testing other
infections such as HSV, varicella-zoster virus (VZV), or syphilis. For
monitoring confirmed or highly probable cases of monkeypox, additional
samples for follow-up testing are required. These samples should consist
of a lesion swab and a pharynx swab, both of which should be stored in
viral transport medium. In addition to obtaining a blood sample using an
EDTA tube, a urine sample should be collected in a universally sterile
container. It is crucial to submit these samples to a facility capable
of conducting MPXV testing. By incorporating these samples into the
monitoring procedure, healthcare providers can collect valuable
information to evaluate clinical deterioration or determine if a patient
should be transferred from isolation to an inpatient setting. In
summary, appropriate sampling techniques are essential for accurate
diagnostic testing in cases of monkeypox. After collecting the
specimens, it is important to store them appropriately for testing.
Place the specimens in sterile leak-proof containers, ensuring
durability and suitability for shipping and temperature conditions. It
is recommended to refrigerate the specimens (2-8°C) or freeze them
(-20°C or lower) if testing cannot occur promptly after collection.
Specimens such as dry swabs, swabs in viral transport medium (VTM), or
lesion crusts stored at 2-8°C can be tested within 7 days from
collection. If stored frozen (-20°C or lower), swab specimens in VTM and
lesion crusts can be tested within 30 days from collection, while dry
swabs stored frozen can be tested within 60 days from collection. When
shipping specimens for laboratory testing, it is important to follow
certain guidelines. The ongoing Mpox outbreak has been linked to Clade
II of the mpox virus, which is now classified as the West African clade.
To ship specimens suspected or confirmed to contain Clade II of the Mpox
virus, they should be labeled and categorized as Biological Substance.
It is recommended to individually bag the specimens to prevent the
rejection of all specimens in case of a leak. To maintain the required
temperature, specimens should be conveyed on dry ice whenever feasible.
It is not recommended to ship specimens at room temperature, as
specimens received outside the permissible temperature range will be
denied. Even if there are temperature fluctuations because of the
refrigerant used, both the primary container and secondary packaging
should maintain their integrity. The packaging of dry ice must prevent
pressure accumulation and container rupture while allowing for gas
discharge. The specimens should only be packaged and shipped by
personnel trained in hazardous transportation requirements. The outer
package must be marked and labelled with the sender’s name and address,
the recipient’s name and address, a biological substance label, the
correct shipping name (Biological substance), an identification number,
and the name and telephone number of the shipper’s responsible party.
Following these instructions ensures the safe and correct shipment of
mpox samples for laboratory testing.
To confirm Mpox virus infection, diagnostic tests, and clinical and
epidemiological data must be correlated. The diagnosis of Mpox(Table 2) virus infection is based on clinical signs,
laboratory tests, and medical history2. By checking
for lymphadenopathy at the prodromal stage of the illness, it is
possible to clinically distinguish Mpox from chickenpox or smallpox53. The specimen samples are collected and forwarded
to the lab to verify the diagnosis in compliance with WHO criteria. PCR
is the most widely used laboratory diagnostic method to identify Mpox
due to its accuracy and sensitivity 54. The fluid that
is produced by vesicles and pustules as well as dry crusts are the best
samples to use in the diagnosis of Mpox. When a lesion sample needs to
be kept cool and kept in a dry, sterile tube without a viral transport
medium, a biopsy appears to be an alternative in some circumstances. PCR
blood tests are often inconclusive and should not be routinely obtained
from patients because viremia typically lasts for a short period
compared to the time the material is taken after symptoms start55. The recommended specimen type for diagnosing
monkeypox is skin lesion material, including swabs of lesion exudate,
roofs from multiple lesions, or lesion crusts. Testing should be offered
to individuals meeting the suspected case definition, considering both
clinical and epidemiological factors. It can be challenging to
differentiate monkeypox based solely on clinical presentation, and other
potential causes of skin lesions should be considered. Safety
procedures, including proper personal protective equipment and handling
of specimens, must be followed. The collection and storage of specimens,
including skin lesion material and additional types for research
purposes, are needed. Nucleic acid amplification testing (NAAT),
particularly polymerase chain reaction, is recommended for confirming
monkeypox infection. Reagents, controls, waste disposal, and biosafety
measures should be appropriately managed during testing. Serological
testing and genetic sequencing are additional tools for diagnosis and
surveillance. Overall, adherence to biosafety measures and risk
assessments are crucial in handling specimens and conducting laboratory
testing for monkeypox. For confirmation of monkeypox virus (MPXV)
infection, clinical and epidemiological data should be considered.
Positive detection using an orthopoxvirus (OPXV) polymerase chain
reaction (PCR) assay, followed by MPXV confirmation via PCR and/or
sequencing, or in suspected cases, positive detection using an
MPXV-specific PCR, confirms MPXV infection. Although MPXV-specific
confirmatory testing is preferred, a positive OPXV PCR assay result is
sufficient for laboratory confirmation. In instances where clinical
presentation and epidemiology indicate MPXV infection despite negative
PCR results, serological testing can be utilised to investigate prior
infection. False-negative results can be influenced by factors such as
specimen quality, handling, transportation, and technical issues.
Genetic sequence data (GSD), obtained through sequencing positive MPXV
specimens, can provide valuable information on the virus’s origins,
epidemiology, and characteristics. WHO encourages countries and
laboratories to share GSD, including raw data, through public access
databases to enhance understanding and collaboration. Sequencing can be
performed using Sanger or next-generation sequencing methods.
Laboratories must adhere to national reporting regulations. Positive or
negative test findings should be immediately reported to national
authorities. States Parties to the IHR are reminded of their
responsibility to communicate relevant public health information with
WHO for occurrences that they have notified WHO. Access to quick and
reliable laboratory testing of samples from cases under investigation is
critical for diagnosing and monitoring this developing virus. All
nations should have access to reliable testing, either locally or by
referral to laboratories in other countries willing and capable of
diagnosing OPXV or MPXV. WHO’s Regional Offices can help Member States
gain access to testing through referral. Inactivation of samples at the
local laboratory, where suitable and securely conducted, may improve
referral and alleviate logistical issues. Countries are invited to
exchange their sequencing data so that the present outbreak can be
better understood.