Case presentation and laboratory diagnosis
On November 4, 2021, a 9-month-old male infant presented a two-day fever
and maculopapular rash, clinically suspected of measles virus infection,
was admitted to Dashti Healthcare Center, Bushehr, Iran. He has not
received the MMR vaccine, as the first shot of the vaccine administers
to children who are 12 months old. There was no information available
regarding the past medical or travelling history of the patient. Upon
admission, a nasopharyngeal swab, urine and serum specimen were
collected which the first two samples, where placed in VTM (viral
transport media). These specimens were sent at a low temperature (4℃) to
the National Measles and Rubella Center for measles virus serological,
cellular and molecular examination according to the CDC and WHO
protocols [11] . At first, Anti-Measles virus IgM Elisa test
(EUROIMMUN,Germany) was performed on the serum specimen in which the
Elisa test was positive. For further investigation, the nasopharyngeal
sample was cultured in Vero-hSLAM cell line, to see the specific
cytopathic effect (CPE) of measles virus –the syncytia. Then the
molecular detection was performed, as the following: total genomic
ribonucleotide acid was extracted using High Pure Viral Nucleic acid kit
(Roche Diagnostics GMbH, Germany) according to the manufacturer’s
instructions. To confirm the serology result, the extracted RNA was
tested for measles virus using the 1-step RT-PCR Master mix
(Biotechrabbit, Germany) by using specific primers (MeV214 and MeV216).
Gel electrophoresis of the PCR product showed an approxiamtely 610 bp
band which were assessed by sanger sequencing using Genetic analyzer
(3130, Applied Biosystems, USA). Nucleotide analysis confirmed indicated
that the measles virus belonged to the B3 strain (based on C-terminal
of the nucleocapsid protein ), which is currently circulating in Iran.
Concurrently, we investigated the nasopharyngeal swab for influenza
virus and SARS-CoV-2 to determine whether the patient had a
co-infection. For influenza virus and SARS-CoV-2 detection, two
separated real-time RT-PCR reactions using specific primers and probes
and Invitrogen (USA) master mix were used. The results showed that, the
patient was positive for influenza A virus with a cycle threshold of 32,
and was negative for SARS-CoV-2. To find out the subtype of this
influenza virus strain, another qRT-PCR (Invitrogen,USA) targeting the
HA gene was performed, which results in influenza virus A/H3N2
identification.