# Environmental surface sampling of respiratory and gastrointestinal pathogens during the 2022–2023 Grand Magal of Touba

Ihssane Ouaddane, Coumba Diouf, Georges Diatta, Cheikh Sokhna, Philippe Gautret

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## To cite this version:

Ihssane Ouaddane, Coumba Diouf, Georges Diatta, Cheikh Sokhna, Philippe Gautret. Environmental surface sampling of respiratory and gastrointestinal pathogens during the 2022–2023 Grand Magal of Touba. Journal of Infection and Public Health, 2025, 18 (5), 102710 [5 p.]. 10.1016/j.jiph.2025.102710. hal-05032297

HAL Id: hal-05032297

https://amu.hal.science/hal-05032297v1

Submitted on 26 Nov 2025

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Original article

# Environmental surface sampling of respiratory and gastrointestinal pathogens during the 2022–2023 Grand Magal of Touba

<figure><img src="image_3.png" /></figure>

Ihssane Ouaddane a,b, Coumba Diouf a,b,c, Georges Diatta c, Cheikh Sokhna a,b,c,
Philippe Gautret a,b,*

## ARTICLE INFO

**Article history:**

Received 2 July 2024
Received in revised form 12 February 2025
Accepted 16 February 2025

**Keywords:**

Environmental sampling
PCR
Magal
Respiratory pathogens
Gastro-intestinal pathogens
Surveillance

## ABSTRACT

**Background:** The Grand Magal of Touba (GMT) pilgrimage is known to be associated with risks of infection, but no studies on the role microbial contamination of the environment has been conducted yet.

**Materials and methods:** We sampled pilgrims and surfaces in the houses where they were accommodated during the 2022–2023 GMT. Respiratory and gastrointestinal pathogens were tested by qPCR.

**Results:** Of the 152 surfaces sampled, 10.5% tested positive for *Streptococcus pneumoniae*, 5.9% for *Staphylococcus aureus*, 16.4% for SARS-CoV-2 (Severe acute respiratory syndrome Coronavirus 2), 3.9% for *Klebsiella pneumoniae*, and 0.6% for rhinovirus. Only *Escherichia coli* was positive out of all the gastrointestinal pathogens tested, with enteropathogenic *E. coli* (EPEC) (23.7%), and enteroaggregative *E. coli* (EAEC) (22.4%) being the most frequent. Of 71 pilgrims tested, 66.2% acquired a respiratory pathogen, including 16.9% *Haemophilus* spp., 7.0% SARS-CoV-2, 11.3% *Streptococcus pneumoniae*, 5.6% *Staphylococcus aureus*, 1.4% rhinovirus, and 2.8% influenza virus. A further 23.9% acquired a gastrointestinal pathogen, including EAEC (18.3%), EPEC (2.8%), *Giardia lamblia* (5.6%) and EIEC/Shigella (2.8%).

**Conclusion:** These preliminary results showed a good correlation between results of surface sampling and human infection for SARS-CoV-2 and EAEC. SARS-CoV-2 surface sampling could be used as a surveillance tool during the GMT. Further larger-scale studies are needed to confirm these preliminary findings.

© 2025 The Author(s). Published by Elsevier Ltd on behalf of King Saud Bin Abdulaziz University for Health Sciences. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).

## Introduction

The Grand Magal of Touba (GMT) is a Muslim religious event celebrated in Senegal on the 18th of Safar, according to the Islamic calendar. It is estimated that between four and five million pilgrims attend the event each year. Pilgrims come from all over Senegal but also from neighboring countries and the diaspora [1]. The two central religious events of the GMT are visits to the mausoleum of Cheikh Ahmadou Bamba, the founder of Mouridism and the Great Mosque of Touba. Pilgrims also visit their personal Mouride spiritual guides, or “marabouts”, who receive their followers in their personal residence in the city. Recitation of the Quran and spiritual inspection complete these religious activities [2].

Pilgrims are usually accommodated in families or in marabout homes, since there is no hotel in Touba, and hundreds of individuals can be accommodated at each location.

Food is prepared and eaten collectively by the pilgrims, and also served in the streets and along the roads to Touba. The slaughtering of animals (mostly cattle) is mostly carried out in a rudimentary way in the streets, in front of residential buildings. In this context, respiratory tract and gastrointestinal infections are frequent among pilgrims, resulting mostly from interhuman transmission in overcrowded conditions [2].

Ndiaw Goumballa et al. did a study in 2022 on the respiratory and gastrointestinal infections at the 2017–2021 GMT on pilgrims from two villages in the south of Senegal, Dielmo and Ndiop. Of the 535 pilgrims tested, 18.4% acquired respiratory viruses, notably rhinovirus (10.1%) and coronaviruses (5.6%) and 39.9% bacteria, notably *Haemophilus* spp. (18.9%) and *Streptococcus pneumoniae* (14.1%). The acquisition of gastrointestinal pathogens was lower, with enteroaggregative *Escherichia coli* (EAEC) (18.9%) and

<footnote>a Aix Marseille Univ, AP-HM, SSA, RITMES, Marseille, France</footnote>

<footnote>b IHU-Méditerranée Infection, Marseille, France</footnote>

<footnote>c MINES, Campus International IRD-UCAD de L'IRD, Dakar, Senegal</footnote>

<footnote>* Correspondence to: RITMES, Institut Hospitalo-Universitaire Méditerranée Infection, 19-21 Boulevard Jean Moulin, Marseille Cedex 05 13385, France.
E-mail address: philippe.gautret@ap-hm.fr (P. Gautret).</footnote>
enteropathogenic *E. coli* (EPEC) (10.5 %) being the most frequent. A decrease was observed in the acquisition rates of pathogens in 2020–2021 GMT [3].

Previous investigations conducted at another religious mass gathering, the Hajj in Saudi Arabia, provided evidence of the presence of nucleic acids of various respiratory pathogens on surfaces frequently touched by pilgrims [4,5]. However, no similar study has been carried out to date on the GMT. To close this gap and provide new findings, we evaluated the presence of respiratory and gastrointestinal pathogens in environmental surfaces during the GMT by sampling surfaces at various houses where pilgrims were accommodated in 2022 and 2023. Pilgrims from Ndiop and Dielmo, hosted in these houses were also sampled.

## Materials and methods

### Location

This study was conducted in Touba, Senegal before and after the GMT that took place on 15 September 2022 and 3–4 September 2023. The same 8 houses were investigated in the two years. Three houses (houses A, B and C) were located in the Soura neighborhood, and surface samples were obtained on 14 September 2022 and on 4 September 2023. Pilgrims from Ndiop were accommodated in these three houses. Three other houses (houses D and E) were investigated in the Darou Minan and Marche Okass neighborhood on 3 September 2023. Pilgrims of Dielmo were hosted in these houses. The investigation of houses (F and G) took place in the Dianatou Mahwa neighborhood on September 16, 2022.

### Sampling strategy

The sampling strategy for this study involved collecting human samples from pilgrims originating from the villages of Dielmo and Ndiop. These two villages were chosen as they benefit from ongoing epidemiological monitoring, providing a unique context for data collection. In Touba, environmental samples were collected from the houses where the same pilgrims from Dielmo and Ndiop were accommodated during the Grand Magal. Both selected houses and pilgrims represent therefore a convenience sample.

### Surface sampling

Surface swabbing was conducted using Sigma Transwabs. Swabbing was performed on 25 cm² (5 × 5 cm) surfaces that were frequently touched by pilgrims, such as door handles, kitchen and bathroom surfaces, and food preparation and storage areas. Swabs were stored at -80 °C before transport to Marseille on dry ice.

### Pilgrim sampling

Throat, nasal and stool swabs were collected between one and three days before leaving Ndiop and Dielmo (pre-Magal samples) and between three and six days following their return from Touba (post-Magal samples). Demographics, chronic medical conditions and respiratory and gastrointestinal symptoms on return were documented using an ad hoc questionnaire.

The procedure was conducted using Sigma Transwabs. Swabs were kept at 4 °C before being transported to the Dakar laboratory for storage in a -80 °C freezer and subsequently transferred to Marseille on dry ice for processing.

### Identification of pathogens

DNA and RNA were extracted using KingFisher™ Flex. All quantitative real-time PCR tests were performed using a C1000 Touch™

Thermal Cycle (Bio-Rad, Hercules, CA, USA). Real-time PCR amplifications were carried out using the LightCycler® 480 Probes Master kit (Roche diagnostics, France) according to the manufacturer's recommendations. Negative controls (PCR mix) and positive controls (DNA from bacterial or parasite strains or RNA from viral strains) were included in each run. A cycle threshold (CT) value of ≤ 35 was used to assess positive results of bacteria, parasites, or virus amplification.

The choice of pathogens that were tested was based on their frequency in respiratory and gastrointestinal samples obtained from GMT pilgrims during previous cohort studies [1].

#### a. Respiratory pathogens

Common human coronaviruses (HCoV) were detected by one-step duplex quantitative RT-PCR amplifications of HCoV/HPIV-R Gene Kit (REF: 71-045, BioMérieux, Marcy l'Etoile, France), according to the manufacturer's recommendations. Influenza A, Influenza B, Human Rhinovirus (HRV), Metapneumovirus (MPV), Respiratory Syncytial Virus (RSV), and SARS-CoV-2 (Severe acute respiratory syndrome Coronavirus 2) were also detected using the multiplex RNA Virus Master Kit (Roche Diagnostics, France) by RT-PCR amplifications. The SHD gene of *Haemophilus* spp., the phoE gene of *Klebsiella pneumoniae*, the nucA gene of *Staphylococcus aureus*, the lytA-CDC gene of *S. pneumoniae*, and the copB gene of *Moraxella catarrhalis* were amplified (Supplementary Table 1).

#### b. Gastrointestinal pathogens

The Multiplex RNA Virus Master Kit (Roche Diagnosis, France) was used to detect the Adenovirus. The mapA gene of *Campylobacter jejuni*, the ipaH gene of *Shigella* spp./EIEC (enteroinvasive *E. coli*), the invA gene of *Salmonella* spp., the stx1 and stx2 genes of EHEC (enterohemorrhagic *E. coli*), the EAF and eae genes of EPEC (enteropathogenic *E. coli*), the pCVD432 gene of EAEC (enteroaggregative *E. coli*), and the whi2 gene for *Tropheryma whipplei* were detected. The 18S gene was used to detect *Entamoeba histolytica* and *Giardia lamblia*. Finally, the Hsp70 gene was amplified to test *Cryptosporidium* spp. (Supplementary Table 2).

## Results

### Environmental samples

In 2022–2023, we collected 152 surface samples in houses where pilgrims were accommodated, including 25 from house A, 33 from house B, 30 from house C, 27 from house D, 22 from house E, 4 from house F and 11 from house G. A total of 67 samples (44.1 %) were positive for at least one pathogen (Table 1), with houses B and C having the highest positivity rates (39.4 % and 63.3 %, respectively).

With regard to respiratory viruses, samples tested were positive only for HRV and SARS-CoV-2, with 0.6 % samples testing positive for HRV and 16.4 % for SARS-CoV-2. Regarding bacteria, 10.5 % samples were positive for *S. pneumoniae*, 5.9 % for *S. aureus* and 3.9 % for *K. pneumoniae*.

Of all the gastrointestinal pathogens tested, only *E. coli* produced positive results, including EIEC/Shigella (0.6 %), EHEC (1.9 %), EPEC (23.7 %), and EAEC (22.4 %) (Table 1).

### Human samples

In 2022 and 2023, a total of 71 pilgrims were sampled, including 19 pilgrims accommodated in house A, 25 in house B, 6 in house C, 13 in house D and 8 in house E. The sociodemographic characteristics of selected pilgrims attending the Grand Magal of Touba reveal a relatively young population, with a mean age of 29.8 years, ranging from 1 year to 74 years. A proportion of 17.7 % of the pilgrims were children aged 0–15 years, while 21.3 % were aged 45 years or older, reflecting a significant presence of both younger and older individuals. In terms of gender distribution, women represent the
<table><caption>**Table 1**
Respiratory and gastrointestinal pathogens on surfaces in houses in Touba during the Grand Magal of Touba.</caption><thead><tr><th>Pathogens</th><th>House A N = 25</th><th>House B N = 33</th><th>House C N = 30</th><th>House D N = 27</th><th>House E N = 22</th><th>House F N = 4</th><th>House G N = 11</th><th>Total N = 152</th></tr></thead><tbody><tr><td>At least one pathogen</td><td>11 (44%)</td><td>13 (39.4%)</td><td>19 (63.3%)</td><td>9 (33.3%)</td><td>7 (31.8%)</td><td>3 (75%)</td><td>5 (45.4%)</td><td>67 (44.1%)</td></tr><tr><td>Respiratory pathogens</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>Influenza A</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td></tr><tr><td>Influenza B</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td></tr><tr><td>HRV</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>1 (9.1%)</td><td>1 (0.6%)</td></tr><tr><td>HCoV</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td></tr><tr><td>RSV</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td></tr><tr><td>MPV</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td></tr><tr><td>SARS-CoV-2</td><td>6 (24%)</td><td>5 (15.1%)</td><td>5 (16.7%)</td><td>6 (22.2%)</td><td>3 (13.6%)</td><td>0 (0%)</td><td>0 (0%)</td><td>25 (16.4%)</td></tr><tr><td>Streptococcus pneumoniae</td><td>1 (4%)</td><td>3 (9.1%)</td><td>9 (30%)</td><td>2 (7.4%)</td><td>1 (4.5%)</td><td>0 (0%)</td><td>0 (0%)</td><td>16 (10.5%)</td></tr><tr><td>Staphylococcus aureus</td><td>1 (4%)</td><td>2 (6.1%)</td><td>5 (16.7%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>1 (9.1%)</td><td>9 (5.9%)</td></tr><tr><td>Klebsiella pneumoniae</td><td>0 (0%)</td><td>1 (3.0%)</td><td>2 (6.7%)</td><td>0 (0%)</td><td>0 (0%)</td><td>1 (25%)</td><td>2 (18.2%)</td><td>6 (3.9%)</td></tr><tr><td>Haemophilus spp.</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td></tr><tr><td>Moraxella catarrhalis</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td></tr><tr><td>Gastro-intestinal pathogens</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>Adenovirus</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td></tr><tr><td>EIEC/Shigella</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>1 (25%)</td><td>0 (0%)</td><td>1 (0.6%)</td></tr><tr><td>EHEC</td><td>1 (4%)</td><td>1 (3.0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>1 (9.1%)</td><td>3 (1.9%)</td></tr><tr><td>EAEC</td><td>5 (20%)</td><td>7 (21.2%)</td><td>3 (10%)</td><td>16 (59.2%)</td><td>1 (4.5%)</td><td>1 (25%)</td><td>1 (9.1%)</td><td>34 (22.4%)</td></tr><tr><td>EPEC</td><td>6 (24%)</td><td>9 (27.3%)</td><td>7 (23.3%)</td><td>12 (44.4%)</td><td>0 (0%)</td><td>1 (25%)</td><td>1 (9.1%)</td><td>36 (23.7%)</td></tr><tr><td>Salmonella</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td></tr><tr><td>Tropheryma whipplei</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td></tr><tr><td>Campylobacter jejuni</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td></tr><tr><td>Giardia lamblia</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td></tr><tr><td>Cryptosporidium spp.</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td></tr><tr><td>Entamoeba histolytica</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td></tr></tbody></table>

HRV: Human Rhinovirus, HCoV: Human Coronavirus, RSV: Respiratory Syncytial Virus, MPV: Metapneumovirus, SARS-CoV-2: Severe acute respiratory syndrome Coronavirus 2, EIEC: Enteroinvasive *E. coli*, EHEC: Enterohemorrhagic *E. coli*, EAEC: Enteroaggregative *E. coli*, EPEC: Enteropathogenic *E. coli*.

majority at 53.4%, while men account for 46.5%. Overall, 3/71 (4.2%) pilgrims reported a chronic medical condition, all of which were cases of hypertension.

About 60.6% pilgrims reported respiratory symptoms on return from Touba while 14.1% reported gastrointestinal symptoms (Supplementary Table 3). Most common respiratory symptoms were rhinitis (43.7%), cough (40.8%) and dyspnea (21.1%). Of note, 19.7% pilgrims were febrile.

Almost half of the pilgrims tested positive for at least one respiratory pathogen before leaving for Touba (Table 2), with, *S. aureus* and *Haemophilus* spp. (22.5%, respectively) (22.5%), *M. catarrhalis* (14.1%) and *S. pneumoniae* (11.3%) being found most frequently. The proportion of pilgrims testing positive for at least one respiratory pathogen upon their return from Touba increased to 67.6%, with the highest acquisition rates for *Haemophilus* spp. (16.9%), rhinovirus (12.7%), and *S. pneumoniae* and *M. catarrhalis* (11.3%, respectively).

About half of the pilgrims tested positive for at least one gastrointestinal pathogen before leaving for Touba (Table 2), with almost all cases found to be due to EAEC (54.9%). The proportion of pilgrims testing positive for at least one pathogen upon their return from Touba increased to 60.6%, with the highest acquisition rate for EAEC (18.3%).

No marked differences were seen among pilgrims from Ndiop or Dielmo and according to houses where they were accommodated (Supplementary Tables 4 to 11).

## Discussion

Demographic characteristics of pilgrims in this study were almost similar to those observed in previous studies [3]. Respiratory tract and, to a lesser extent, gastrointestinal infection symptoms are common during the GMT, with rates of 55% and 13%, respectively being reported [3]. In this study, the prevalence of respiratory symptoms in pilgrims was slightly higher than that in the pre-COVID

<table><caption>**Table 2**
Respiratory and gastrointestinal pathogens in pilgrims before and after participating in the Grand Magal of Touba.</caption><thead><tr><th></th><th>Pre-Magal<br>N = 71</th><th>Post-Magal<br>N = 71</th><th>Acquisition<br>N = 71</th></tr></thead><tbody><tr><td>Respiratory pathogens</td><td></td><td></td><td></td></tr><tr><td>At least one pathogen</td><td>41 (57.7%)</td><td>48 (67.6%)</td><td>47 (66.2%)</td></tr><tr><td>Influenza A</td><td>1 (1.4%)</td><td>2 (2.8%)</td><td>2 (2.8%)</td></tr><tr><td>Influenza B</td><td>0 (0%)</td><td>1 (1.4%)</td><td>1 (1.4%)</td></tr><tr><td>HRV</td><td>0 (0%)</td><td>9 (12.7%)</td><td>9 (12.7%)</td></tr><tr><td>HCoV</td><td>2 (2.8%)</td><td>0 (0%)</td><td>0 (0%)</td></tr><tr><td>RSV</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td></tr><tr><td>MPV</td><td>0 (0%)</td><td>1 (1.4%)</td><td>1 (1.4%)</td></tr><tr><td>SARS-CoV-2</td><td>1 (1.4%)</td><td>5 (7.0%)</td><td>5 (7.0%)</td></tr><tr><td>Streptococcus pneumoniae</td><td>8 (11.3%)</td><td>14 (19.7%)</td><td>8 (11.3%)</td></tr><tr><td>Staphylococcus aureus</td><td>16 (22.5%)</td><td>16 (22.5%)</td><td>4 (5.6%)</td></tr><tr><td>Klebsiella pneumoniae</td><td>3 (4.2%)</td><td>1 (1.4%)</td><td>1 (1.4%)</td></tr><tr><td>Haemophilus spp.</td><td>16 (22.5%)</td><td>31 (43.7%)</td><td>12 (16.9%)</td></tr><tr><td>Moraxella catarrhalis</td><td>10 (14.1%)</td><td>17 (23.9%)</td><td>8 (11.3%)</td></tr><tr><td>Gastro-intestinal pathogens</td><td></td><td></td><td></td></tr><tr><td>At least one pathogen</td><td>42 (59.1%)</td><td>43 (60.6%)</td><td>17 (23.9%)</td></tr><tr><td>Adenovirus</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td></tr><tr><td>EIEC/Shigella</td><td>7 (9.8%)</td><td>2 (2.8%)</td><td>2 (2.8%)</td></tr><tr><td>EHEC</td><td>1 (1.4%)</td><td>3 (4.2%)</td><td>3 (4.2%)</td></tr><tr><td>EAEC</td><td>39 (54.9%)</td><td>32 (45.7%)</td><td>13 (18.3%)</td></tr><tr><td>EPEC</td><td>2 (2.8%)</td><td>2 (2.8%)</td><td>2 (2.8%)</td></tr><tr><td>Salmonella</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td></tr><tr><td>Tropheryma whipplei</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td></tr><tr><td>Campylobacter jejuni</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td></tr><tr><td>Giardia lamblia</td><td>2 (2.8%)</td><td>5 (7.0%)</td><td>4 (5.6%)</td></tr><tr><td>Cryptosporidium spp.</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td></tr><tr><td>Entamoeba histolytica</td><td>0 (0%)</td><td>0 (0%)</td><td>0 (0%)</td></tr></tbody></table>

HRV: Human Rhinovirus, HCoV: Human Coronavirus, RSV: Respiratory Syncytial Virus, MPV: Metapneumovirus, SARS-CoV-2: Severe acute respiratory syndrome Coronavirus 2, EIEC: Enteroinvasive *E. coli*, EHEC: Enterohemorrhagic *E. coli*, EAEC: Enteroaggregative *E. coli*, EPEC: Enteropathogenic *E. coli*.
period. Of note, that of dyspnea in 2023 was three times higher than previously reported and we have no clear explanation for this. Further studies in following years will be of interest to this respect. Acquisition rates of respiratory and gastro-intestinal pathogens have been documented in pilgrims from Ndiop and Dielmo in 2017–2021 [3] and we found similar results in 2022 with the notable exception of an acquisition of SARS-CoV-2 that had not been observed in previous years [6].

We found low rates of viral contamination of surfaces with less than 1% of samples testing positive for rhinovirus, but 16% testing positive for SARS-CoV-2. Higher rates were observed for bacterial contamination, with 24% of surfaces testing positive for EPEC, 22% for EAEC, 10% for S. pneumoniae, 6% for S. aureus, 4% for K. pneumoniae, 2% for EHEC and less than 1% for EIEC/Shigella. No surface tested positive for intestinal protozoa.

SARS-CoV-2 and EAEC were both relatively frequent on surfaces and in pilgrim samples. In addition, in houses A and B where the number of surfaces sampled was high and where greatest numbers of pilgrims were sampled, a relative correlation was observed between results on surface and in clinical samples for these two pathogens. A contrario, some pathogens like rhinovirus, Haemophilus spp. and M. catarrhalis, were not or rarely found on surfaces although they were frequent in pilgrims. Finally, some pathogens like EPEC were relatively frequently observed on surfaces while their prevalence in clinical samples was low.

Most studies addressing the role of surfaces in the transmission of pathogens have been conducted in the health care setting. Most gram-positive bacteria, including *Enterococcus* spp., *S. aureus*, and *Streptococcus pyogenes*, can survive for months on dry surfaces [7]. Many gram-negative bacteria, including *Acinetobacter* spp., *E. coli*, *Klebsiella* spp., *Pseudomonas aeruginosa*, *Serratia marcescens*, and *Shigella* spp. can also survive for several months [7]. In contrast, *H. influenzae* persists only for a few days, as do respiratory viruses such as coronaviruses, rhinoviruses, and influenza viruses [7]. Many studies reported the identification of SARS-CoV-2 on surfaces, however there is a lack of evidence that these viruses were viable [8]. One study conducted on aircraft surfaces suggests that SARS-CoV-2 only survives for a few hours [9].

To our knowledge, only two studies evaluating the presence of pathogens in surface samples have been conducted in the context of a mass gathering. Both were conducted at the Hajj in Saudi Arabia, an event which is associated with a high prevalence of respiratory tract infections among pilgrims [10]. Memish et al. conducted a study in 2013, on highly touched surfaces in the international airport of Jeddah at the time of the pilgrims' departure after participating in the Hajj [4]. Using a multiplex PCR test, seven of 40 surface samples tested positive including three for adenovirus, three for coronavirus, and one each for *H. influenzae* and *M. catarrhalis* [4]. In 2016 and 2018, Hoang et al. tested frequently touched surfaces in hotels in Mecca and Medina, commercial centers and restaurants in Mecca, tents and ablution facilities in Mina and Arafat, and the interior of a bus travelling from Mina to Mecca. Of 142 samples, 70 tested positive by PCR. Overall, the prevalence of positive surfaces was 28% for *K. pneumoniae*, 16% for rhinovirus, 6% for S. pneumoniae, 5% for S. aureus, 4% for *H. influenzae*, and 1% for coronavirus [5].

Our results at the GMT are unique. They reveal relatively high rates of SARS-CoV-2 and EAEC contamination of surfaces that are also frequently found in pilgrims. The role of surface contamination as a possible source of infection in humans is difficult to establish although it has been suggested for both S. pneumoniae and EAEC [11,12]. The variation in Gregorian dates for the Grand Magal, and thus the seasons, likely influences microbiological findings. Seasonality significantly impacts pathogen circulation, with respiratory viruses like influenza and RSV peaking in colder months, while gastrointestinal pathogens are more common in warmer seasons.

These variations may affect the types and prevalence of pathogens detected.

On the other hand, SARS-CoV-2 surveillance of wastewater has been successfully used for tracking the virus at the population level [13].

Unfortunately, there is no wastewater collection system in Touba to conduct such experiments. Surface sampling could therefore be used as a potential tool for early detection of the virus before it spreads in the community during the GMT.

Our study has some limitations. First, the use of a convenience sample may have induced a selection bias. Secondly, the use of a questionnaire may have induced recall bias and some questions may have been misunderstood. However, our results are relatively similar over years suggesting that answers to questionnaires are reliable. Also, we sampled a small number of surfaces. Thirdly, we tested only 12 respiratory pathogens and 11 gastrointestinal pathogens (chosen based on their higher frequency in pilgrims). Also, because the detection was based on molecular techniques only, we cannot conclusively prove that the detected pathogens were alive and potentially infective. Finally, this study lacks advanced molecular techniques like PFGE, MLST, or genomic analysis, limiting insights into strain persistence and global relevance. Finally, the local nature of the Grand Magal and the non-generalizability of the results should be acknowledged. However, the study provides useful insights that could inform strategies for managing similar mass gatherings in Africa.

## Conclusion

Our study highlights the presence of significant surface contamination by SARS-CoV-2 and EAEC during the Grand Magal of Touba, with these pathogens also frequently detected in pilgrims. This suggests potential interactions between surface contamination and human infection, particularly for pathogens like SARS-CoV-2, which has been effectively monitored through environmental surveillance in other contexts. Our findings underscore the value of surface sampling as a complementary tool for pathogen detection, especially in settings lacking wastewater surveillance systems, and emphasize the need for further research to better understand the role of surface contamination in pathogen transmission during mass gatherings.

## List of abbreviations

**COVID:** CoronaVirus Disease - 2019

**CT:** Cycle Threshold

**DNA:** Deoxyribonucleic Acid

**EAEC:** Enteroaggregative *E. coli*

**EHEC:** Enterohemorrhagic *E. coli*

**EIEC:** Enteroinvasive *E. coli*

**EPEC:** Enteropathogenic *E. coli*

**GMT:** Grand Magal of Touba

**HCoV:** Human Coronavirus

**HPIV:** Human Parainfluenza Viruses

**MLST:** Multilocus Sequence Typing

**PCR:** Polymerase Chain Reaction

**PFGE:** Pulsed-Field Gel Electrophoresis

**qPCR:** Quantitative Polymerase Chain Reaction

**RNA:** Ribonucleic acid

**RT-PCR:** Real-time Polymerase Chain Reaction

**SARS-CoV-2:** Severe acute respiratory syndrome Coronavirus 2

## Ethics approval and consent to participate

Pilgrims were invited to participate on a voluntary basis. Participants (or their parents when they were minors) were asked to
sign a written consent form. The protocol was approved by the
Senegalese National Ethics Committee for Health Research (SEN17/
62) and was carried out in accordance with the good clinical prac-
tices recommended by the Declaration of Helsinki and its amend-
ments.

The transport of samples to France was approved by the
Senegalese Ministry of the Environment and Sustainable
Development (No. 1270 DPN/PF-APA).

Consent for publication

The authors understand and agree that the data provided may be
included in this paper and published.

Funding

This study was supported by the Institut Hospitalo-Universitaire (IHU) Méditerranée Infection, the French National Research Agency under the "Investissements d'avenir" programme, reference ANR-10-IAHU-03.

Author contributions statement

PG and CS designed the study and coordinated the work. IO, CD and GD collected the samples. IO and CD conducted the qPCR technique. IO wrote the draft. CS critically reviewed the manuscript. All authors contributed to and approved the current version of the manuscript.

Data availability

Data and materials available on request.

Declaration of Competing Interest

The authors declare that they have no known competing fi-
nancial interests or personal relationships that could have appeared
to influence the work reported in this paper.

Acknowledgements

Not applicable.

**Appendix A. Supporting information**

Supplementary data associated with this article can be found in
the online version at doi:10.1016/j.jiph.2025.102710.

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