key: cord-326768-uo6482ah
authors: Hashem, Anwar M.; Al‐Subhi, Tagreed L.; Badroon, Nassrin A.; Hassan, Ahmed M.; Bajrai, Leena Hussein M.; Banassir, Talib M.; Alquthami, Khalid M.; Azhar, Esam I.
title: MERS‐CoV, influenza and other respiratory viruses among symptomatic pilgrims during 2014 Hajj season
date: 2019-02-20
journal: J Med Virol
DOI: 10.1002/jmv.25424
sha: 
doc_id: 326768
cord_uid: uo6482ah

More than two million Muslims visit Makkah, Saudi Arabia, annually to perform the religious rituals of Hajj where the risk of spreading respiratory infections is very common. The aim here was to screen symptomatic pilgrims for Middle East respiratory syndrome coronavirus (MERS‐CoV) and other viral etiologies. Thus, 132 nasopharyngeal samples were collected from pilgrims presenting with acute respiratory symptoms at the healthcare facilities in the holy sites during the 5 days of the 2014 Hajj season. Samples were tested using real‐time reverse transcription polymerase chain reactions and microarray. Demographic data including age, sex, and country of origin were obtained for all participants. While we did not detect MERS‐CoV in any of the samples, several other viruses were detected in 50.8% of the cases. Among the detected viruses, 64.2% of the cases were due to a single‐virus infection and 35.8% were due to the coinfections with up to four viruses. The most common respiratory virus was influenza A, followed by non‐MERS human coronaviruses, rhinoviruses, and influenza B. Together, we found that it was not MERS‐CoV but other respiratory viruses that caused acute respiratory symptoms among pilgrims. The observed high prevalence of influenza viruses underscores the need for more effective surveillance during the Hajj and adoption of stringent vaccination requirements from all pilgrims.

at the Jamaraat pillars in Mina. Finally, they finish their Hajj ritual by going back to the holy mosque in Makkah to perform "Tawaf".

Overcrowding of individuals in such confined settings leads to inevitable prolonged close contact and increases the risk of spreading and acquiring respiratory pathogens among pilgrims, which raises global and public health concerns due to the high potential of international spread of such pathogens. [1] [2] [3] [4] Another important driver of spreading and acquiring respiratory pathogens during Hajj is the great diversity of inbound viruses from around the world that can potentially spread among the immunologically naive hosts. In fact, acute respiratory infections are very common during Hajj and represent the leading cause of the most hospitalizations. [3] [4] [5] [6] [7] It has been suggested that more than one-third of pilgrims will suffer from respiratory symptoms during Hajj mostly due to the respiratory viruses. 2, [7] [8] [9] [10] [11] [12] Most commonly isolated viruses from symptomatic patients during Hajj were human rhinoviruses (hRVs), influenza virus and non-MERS human coronaviruses (hCoVs). 2, [9] [10] [11] [12] [13] [14] [15] [16] [17] [18] [19] [20] The emergence of the novel Middle East respiratory syndrome coronavirus (MERS-CoV) in Saudi Arabia, its endemicity, and high mortality rates (35%-40%) clearly represent another major public health concern, especially during Hajj. Since 2012, MERS-CoV caused more than 2250 confirmed cases in 27 countries in the Arabian Peninsula, Africa, Asia, Europe, and North America as of December 2018. 21, 22 Furthermore, multiple hospitals and household outbreaks have been reported mostly in the Saudi Arabia. 23 

RNA extraction was performed using the QIAamp Viral RNA Mini Kit (Qiagen, Hilden, Germany) according to manufacturerʼs instructions.

Extracted RNA from all samples was tested for MERS-CoV using realtime reverse transcription polymerase chain reactions targeting upstream region of the E-gene as described previously. 33 Positive and negative (no template) controls were included in all runs.

Remaining NP samples were used for complementary DNA (cDNA) synthesis and microarray testing as described previously. 34 Plus Analyzer (AutoGenomics Inc., Carlsbad, CA) according to manu-facturerʼs instructions and as previously described. 34 Samples were considered positive when the ratio between the virus and background signals was above the calculated threshold.

The data were analyzed using the Statistical Package for the Social Science Software (SPSS v20.0; SPSS Inc, Chicago, IL). The χ 2 and Fisher exact tests were used to compare the proportions and a twotailed probability value of P < 0.05 was considered statistically significant. Table 1 ). As shown in Table 1 , although a majority of positive patients were older than 60 years, this difference was not significant.

No statistical significance was found for comparisons of infection rates among males and females or the different age groups. One hundred and twenty-nine of the patients in this study were from 37 countries mostly from Asia and Africa followed by Europe and North America. Nationalities of three individuals were unknown. As (Table 3) . Coinfections with more than two viruses were not uncommon. In fact, four patients had triple concurrent infections, and one patient had a quadruple concurrent infection ( Table 3 ). The remaining nine coinfections were due to the unique combination of respiratory viruses (Table 3) . Not surprisingly, flu A, flu B, and hCoV OC43 were the most frequently detected viruses in the most age groups ( Coinfections with multiple viruses in symptomatic pilgrims were very common in our study and represented more than 35% of the positive cases and more than 18% of the total number of patients.

This rate is markedly higher than the previously reported. [10] [11] [12] [13] 17, 18, 38 Furthermore, while most of these reports have identified hRVs as the most coinfecting viruses during Hajj, our data showed hCoV OC43 as the most prevalent coinfecting virus followed by flu A and hRVs. [10] [11] [12] [13] 17, 18, 38 These marked differences most probably were due to technical and methodological differences. However, it is clear that enhanced surveillance using detection assays with high sensitivity and coverage such as microarray and multiplex PCR could enhance our understanding of pathogens involved in respiratory infections during mass gatherings and ultimately lead to better infection control. 12, 19 or hRV C, which could represent a significant number of potential rhinovirus infections among symptomatic pilgrim.

In conclusion, we investigated the etiology of acute respiratory infections in symptomatic pilgrims attending 2014 Hajj. While MERS-CoV was not detected in any of the patients, a variety of other respiratory viruses has been found in more than half of the patients with many coinfections with multiple viruses. Our observation as well as the previous reports from Hajj indicate that enhanced and active surveillance during Hajj seasons is critical to recognize the wide variety of pathogens that might be involved in Hajj epidemics and to implement proper infection control measures. Importantly, there is an evident risk of influenza infection among pilgrims underscoring the need for targeted, active and continuous surveillance for influenza viruses not only to monitor viral circulation but also to characterize circulating viruses to better understand vaccine effectiveness and to recognize the need to improve current influenza vaccination strategies during Hajj.

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High prevalence of common respiratory viruses and no evidence of Middle East respiratory syndrome coronavirus in Hajj pilgrims returning to Ghana

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Influenza and respiratory syncytial virus infections in British Hajj pilgrims

2 Patterns of reported respiratory symptoms and detected respiratory viruses during Hajj days. A, Bar graph showing the number of pilgrims presented with respiratory symptoms and tested for respiratory viruses with numbers of those tested positive or negative for any virus during daily. B, Bar graph showing the number of viruses detected from positive cases during the 5 days of Hajj. Day 1, Makkah then Mina in the afternoon; day 2, Arafat in the morning then Muzdalifah at night; day 3, Muzdalifah then Mina in the morning

Detection of respiratory viruses among pilgrims in Saudi Arabia during the time of a declared influenza A(H1N1) pandemic

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Hajj-part of the MERS-CoV surveillance program

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An opportunistic pathogen afforded ample opportunities: Middle East respiratory syndrome coronavirus

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MERS-CoV, influenza and other respiratory viruses among symptomatic pilgrims during 2014 Hajj season

The authors declared that there is no conflict of interests.

http://orcid.org/0000-0002-8471-7011
