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Adv Biomed Res 2023,  12:180

Human respiratory syncytial virus and coronavirus rates in Iranian elderly patients more than 60 years old with acute respiratory symptoms

1 Department of Microbiology, School of Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran
2 Infectious Diseases and Tropical Medicine Research Center, Shahid Beheshti University of Medical Sciences, Tehran, Iran

Date of Submission19-Nov-2022
Date of Acceptance31-Jan-2023
Date of Web Publication20-Jul-2023

Correspondence Address:
Dr. Ebrahim Faghihloo
Department of Microbiology, School of Medicine, Shahid Beheshti University of Medical Sciences, Tehran
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Source of Support: None, Conflict of Interest: None

DOI: 10.4103/abr.abr_395_22

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Background: Scientists have believed that a number of risk factors, especially viral infectious agents, can be related to respiratory diseases. Due to the pandemics in 2019, Human Respiratory Syncytial Virus and Coronavirus have attracted the attention of different kinds of research. In this study, we attempted to evaluate the prevalence of these viruses.
Materials and Methods: After extracting the RNA and DNA of these viruses, molecular tests were employed to report the rate of them in patients suffering from respiratory symptoms.
Results: Our results demonstrated that 31 samples were COVID-19 positive. Furthermore, two cases had Respiratory syncytial virus (RSV) subgroup A infections. However, no cases showed a coinfection of both viruses.
Conclusions: It seems that during the pandemic of COVID-19, RSV should not be ignored as it can be responsible for the respiratory syndrome.

Keywords: Coronavirus, epidemiology, RSV

How to cite this article:
Tavakolian S, Goudarzi H, Alavi Darazam I, Faghihloo E. Human respiratory syncytial virus and coronavirus rates in Iranian elderly patients more than 60 years old with acute respiratory symptoms. Adv Biomed Res 2023;12:180

How to cite this URL:
Tavakolian S, Goudarzi H, Alavi Darazam I, Faghihloo E. Human respiratory syncytial virus and coronavirus rates in Iranian elderly patients more than 60 years old with acute respiratory symptoms. Adv Biomed Res [serial online] 2023 [cited 2023 Sep 28];12:180. Available from:

  Introduction Top

Despite a progression in the health-care system, most scientists believe that respiratory diseases can cause high rates of mortality. This health problem is the third cause of death in developed countries, especially in European countries, comprising 440 000 in 2015.[1],[2] In the current age, greenhouse gasses, such as carbon dioxide emitted from numerous transportation systems, can progress this disease detrimentally. Unfortunately, some other risk factors can have the same effects on the public, ranging from smoking to tobacco smoking (including second-hand smoke) to allergens.[3]

However, infectious agents are other items that have been in the center of concentration. Take, for example, Respiratory syncytial virus (RSV) infection, which can account for one of the infant mortality (less than 1 year of age worldwide).[4] This virus belongs to the genus Orthopneumovirus, family Pneumoviridae, and order Mononegavirales.[5],[6],[7],[8] In terms of structure, this virus has an envelope and spherical capsid, packaging a negative-sense RNA genome.[9] A meta-analysis illustrated that approximately 160 000 kids (less than 5 years of age) died in 2015 due to this virus.[10] RSV usually leads to pneumonia and bronchiolitis when they are newborns[11]; nevertheless, elders need to be hospitalized in cold seasons because of this virus.[12] Two subgroups of RSV can be observed, including subgroups A and B, which can react with monoclonal antibodies directed against the G protein.[13]

Another virus that is suffering patients is the Coronavirus, and some health problems can be related to this virus, including severe acute respiratory syndrome (SARS), the common cold, and the Middle East respiratory syndrome (MERS).[14] This virus, which was less known before 2019, cast a shadow all around the world as it caused a pandemic called COVID-19. Around 620 000 and 27 000 individuals were infected and passed away, respectively, until April 15, 2020 in the United States.[15] Saliva, droplets, or discharges from the nose of an infected individual after coughing or sneezing can spread COVID-19. These viruses belong to Coronaviridae, and scientists classified them into four distinct genera, including Alpha coronavirus, Beta coronavirus, Delta coronavirus, and Gamma coronavirus. Nevertheless, only alpha and beta coronaviruses are able to infect mammals.[16] There is a spike (S) protein composed of more than 1300 amino acids that is responsible for interacting with cell receptors. Indeed, they can recognize zinc peptidase angiotensin-converting enzyme 2 (ACE2), and some other receptors.[17],[18] They usually enter the cells through endocytosis, and lysosomal proteases process their spike. Single-strand RNA accounts for virus reproduction since it uses the ribosome of the host cell to replicate polyproteins.[19] It should be mentioned that the non-structural genes of COVID-19 are located within the 5′-proximal two-thirds of the genome on gene 1; however, the 3′-proximal one-third of the genome (8–9 genes for coronaviruses) encodes the structural proteins.[20]

As both RSV and Coronavirus can spread quickly, causing high rates of death all around the world such as COVID-19, we attempted to evaluate the prevalence of these viruses in 2021, when there was a pandemic of Coronavirus.

  Materials and Methods Top


The ethical code of this article, which is approved by Shahid Beheshti University of Medical Sciences, is” IR.SBMU.MSP.REC.1399.164 (Grant no 23305). In this case-control study, we collected 79 samples of people in Loghman Hospital, Tehran, Iran, who had some symptoms, such as wheezing, coughing, and fever, between January and September 2021. All patients were more than 60 years old, and written consent was given from all of them for publications of this article. These throat swab specimens were obtained from patients with influenza-like illness and maintained in a viral transport medium. These swabs with ice packs were transported and stored at the Shahid Beheshti University of Medical Sciences. After collection, all samples were stored in =80°C in the Shahid Beheshti University of Medical Sciences Microbiology Department.

RNA extraction and cDNA synthesis for respiratory syncytial virus

RNA from throat swabs was extracted by the RSV RNA Extraction Kit (ViraGen, India), and we dissolved it in 50 ul of elution buffer. After that, 17.5 μL of extracted RNA was combined with 2.5 μL of dNTP, 1 μL of RT enzyme of moloney murine leukemia virus, 6 μL of RT buffer, 2.5 of μL random hexamer primers, and 0.5 μL of RNase inhibitor in a total volume of 30 μL. This mixture was incubated at 37°C for 45 min, according to the kit.

Primer sequences for respiratory syncytial virus

In the external step, forward primers were applied with GPB (nt 494-515, 5'-AAGATGATTACCATTTTGAAGT-3') and GPA (nt 511-530, 5'- GAAGTGTTCAACTTTGTACC-3') for subgroup B and subgroup A, respectively. We performed hemi-nested PCR step using nRSAG (nt 539-558, 5'- TATGCAGCAACAATCCAACC-3') as HRSV-A specific forward primer, and Nrsbg (nt 512-531, 5'-GTGGCAACAATCAACTCTGC-3') as HRSV-B specific forward primer. Common reverse primers for subgroups A and B called F1 (nt 3-22, 5'-CAACTCCATTGTTATTTGCC-3') in both external and hemi-nested polymerase chain reaction steps were employed.

External PCR for respiratory syncytial virus

27 ul of distilled water, 5 ul of 10X PCR buffer, 2 ul of dNTP, 2 ul of Mgcl2, 1 ul of Taq DNA polymerase, 1.5 ul of reverse primer, 1.5 ul of forward primer, and 10 ul of synthesized cDNA were mixed to perform PCR in the external step. The PCR schedule was 5 min at 94°C, which was followed by 35 cycles of 94°C for 30 sec, 50°C for 30 sec, 72°C for 30 sec, and finally 72°C for 10 min. The final product for external PCR was 450 bp in gel electrophoresis.

Hemi-nested PCR for respiratory syncytial virus

Hemi-nested PCR steps had a similar pattern to the external ones, yet with a 400-bp final product. We used the standard HRSV A2 strain as a positive control; moreover, negative control was utilized to confirm our clinical performances. We run our final products in 1.5% agarose gel electrophoresis under Ultraviolet light.

Real-time PCR test to detect SARS-COV2 RNA

We employed a one-Step RT-PCR COVID-19 kit from Pishtaz Teb Company. Two viral genes (namely RNA-dependent RNA polymerase (RdRp) and (N) Nucleocapsid) were targeted by this kit containing a primer and probe for the RNase P gene to prevent false negatives in the extraction steps. After amplification, we evaluated the internal control in the channel, ROX, and negative and positive controls in the FAM and HEX channels. The negative and positive controls had no fluorescence signal and a Ct ≤35 cycles, respectively. The positive threshold of this kit for both genes and the internal control should be Ct ≤40 with the sigmoid S curve.

  Results Top

We examined 79 samples (42 male and 37 female) of elders, including 23, 27, and 29 individuals who were in sixth, seventh, and eighth of their lives, suffering from respiratory diseases. Our results demonstrated that 31 samples were COVID-19 positive. Furthermore, two cases had RSV subgroup A infections, but no cases showed a coinfection of both viruses [Table 1] and [Table 2].
Table 1: Data about patients with positive Covid-19

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Table 2: Data about patients with positive SARS

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  Discussion Top

Unfortunately, health problems in some parts of the body, such as the sinuses, throat, airways, or lungs, can be responsible for the growing trend of death in the world.[21] Being a host to different kinds of infectious agents, which can occur because of exposure to animals, is likely to be a reason why some epidemics can be observed since a number of studies have reported this issue.[22],[23] Given the fact that these viruses are able to spread as easily as possible, evaluating their prevalence can give a new perspective to solve this problem. In terms of geography, we should compare a number of regions to understand the effects of vaccines, social distance, or other ways of virus transmission. In this study, we reported the incidence of two significant viruses in a Middle East country called Iran, which experienced fluctuations in Coronavirus death rates during the COVID-19 pandemic. The limitation of this study is the low number of samples, as it is difficult to collect them from the hospitals.

We found that among 79 patients suffering from respiratory diseases, two samples were RSV subgroup A infections positive. Some researchers have published the epidemiology of this infectious agent. For instance, a meta-analysis conducted on Iranian samples illustrated that 775 (18.7%) out of 4140 patients had RSV infections from 1996 to 2013.[24] Around 67% and 33% of 297 specimens from South Korean infants or children between 2008 and 2009 had HRSV-A and HRSV-B, respectively.[25] In Brazil, most RSV-positive people's mean age was lower than the negative ones, and researchers found this virus in 243 (23.1%) children. In this study, RSV was detected, with a predominance in Group B (209/227), and 10% of them suffered from bacterial infections.[26] Mathur and colleagues in a serologic study found that 32 patients had RSV, while 24 individuals had Influenza among 71 cases, and they reported that RSV can be significant in elders.[27] In 2018, 45.3% and 53.7% were subtypes A and B, respectively, and only 1% of hospitalized persons had both of these viruses. Moreover, they believe that the more age increases, the more chance of RSV-B infections can be observed.[28]

Another virus assessed in this study was the Coronavirus, and 31 samples were COVID-19 positive. In 2020, Desai and colleagues conducted a study to evaluate the outbreak of this virus. They found that 13.2% of patients with a mean age >50, but younger patients (mean age <50 years) illustrated a pooled prevalence of 9.0%.[29] A review in the USA showed that COVID-19 can take a heavy toll on elders, such as the highest mortality rates, hospitalizations, and hospitalizations in the intensive care unit.[30] During the COVID-19 pandemic, researchers suggested that some risk factors ranging from inherent immunity, temperature-humidity variation, to lower life expectancy can effect on the Coronavirus prevalence, which is the reason why in numerous countries we are likely to observe different rates of this virus.[31] It seems that this virus has a higher rate among cardiovascular and metabolic diseases in comparison with intensive care unit severe and non-ICU/severe patients.[32] Another study reported that patients with diabetes who suffered from COVID-19 had more severe symptoms.[33] More than 60% of patients hospitalized due to SARS-CoV-2 had post-COVID-19 symptoms.[34]

  Conclusion Top

It seems that during the pandemic of COVID-19, RSV should not be ignored as it can be responsible for the respiratory syndrome.

Financial support and sponsorship

The ethical code of this article, which is approved by Shahid Beheshti University of Medical Sciences, is” IR.SBMU.MSP.REC.1399.164 (Grant no 23305).

Conflicts of interest

There are no conflicts of interest.

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  [Table 1], [Table 2]


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