- Email: [email protected]
Molecular dynamics of Staphylococcus aureus nasal carriage in Hajj pilgrims
Accepted Manuscript Molecular dynamics of Staphylococcus aureus nasal carriage in Hajj pilgrims Paul O. Verhoeven, Philippe Gautret, Cyrille H. Haddar, Samir Benkouiten, Julie Gagnaire, Khadidja Belhouchat, Florence Grattard, Rémi Charrel, Bruno Pozzetto, Tassadit Drali, Frédéric Lucht, Philippe Brouqui, Ziad A. Memish, Philippe Berthelot, Elisabeth Botelho-Nevers PII:
S1198-743X(15)00384-5
DOI:
10.1016/j.cmi.2015.03.020
Reference:
CMI 224
To appear in:
Clinical Microbiology and Infection
Received Date: 18 January 2015 Revised Date:
22 March 2015
Accepted Date: 26 March 2015
Please cite this article as: Verhoeven PO, Gautret P, Haddar CH, Benkouiten S, Gagnaire J, Belhouchat K, Grattard F, Charrel R, Pozzetto B, Drali T, Lucht F, Brouqui P, Memish ZA, Berthelot P, BotelhoNevers E, Molecular dynamics of Staphylococcus aureus nasal carriage in Hajj pilgrims, Clinical Microbiology and Infection (2015), doi: 10.1016/j.cmi.2015.03.020. This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
ACCEPTED MANUSCRIPT Molecular dynamics of Staphylococcus aureus nasal carriage in Hajj pilgrims
Paul O. Verhoeven1,2, Philippe Gautret 3, Cyrille H. Haddar 2, Samir Benkouiten3, Julie Gagnaire1, Khadidja Belhouchat3, Florence Grattard1,2, Rémi Charrel3, Bruno Pozzetto1,2,
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Tassadit Drali3, Frédéric Lucht1,4, Philippe Brouqui3, Ziad A. Memish5 , Philippe Berthelot1,2,4 and Elisabeth Botelho-Nevers1,4
GIMAP EA 3064 (Groupe Immunité des Muqueuses et Agents Pathogènes), University of
SC
1
Lyon, 42023 Saint-Etienne, France
Laboratory of Infectious Agents and Hygiene, University Hospital of Saint-Etienne, 42055
Saint-Etienne Cedex 02, France 3
M AN U
2
Aix Marseille Université, URMITE, UM63, CNRS 7278, IRD 198, Inserm 1095 and Institut
Hospitalo-Universitaire Méditerranée Infection, Marseille, France
Infectious Diseases Department, University Hospital of Saint-Etienne, 42055 Saint-Etienne
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4
Cedex 02, France 5
Public Health Directorate, Saudi Ministry of Health, World Health Organization
EP
Collaborating Center for Mass Gathering Medicine and College of Medicine, Alfaisal
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University, Riyadh, Kingdom of Saudi Arabia
Running title: Staphylococcus aureus carriage in Hajj Keywords: Staphylococcus aureus carriage; carriage acquisition; Hajj; overcrowding; respiratory viruses.
Corresponding author:
E. Botelho-Nevers: [email protected] Tél: + (33) 4 77 12 77 89 Fax: + (33) 4 77 12 78 24
ACCEPTED MANUSCRIPT ABSTRACT
During the 2012 Hajj season, the acquisition risk of Staphylococcus aureus nasal carriage in a
RI PT
cohort of French pilgrims was 22.8%, statistically associated with the acquisition of viral respiratory pathogens (P =0.03). The carriage of S. aureus belonging to the emerging clonal
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complex 398 significantly increased following the pilgrimage (P < 0.05).
ACCEPTED MANUSCRIPT 1
Staphylococcus aureus nasal carriage is common and virtually all individuals could become carrier during their live [1]. Intercontinental travels play a significant role in the
3
spread of specific S. aureus clones such as previously shown for the CA-MRSA USA300
4
clone [2]. The Hajj Muslim pilgrimage to Mecca is the largest annual mass gathering in the
5
world; the overcrowding is major, leading to a high risk of transmission of pathogens between
6
pilgrims [3]. Transmission of S. aureus has been largely described in household and hospital
7
settings associated with overcrowding [4]. To our knowledge, only one study examined S.
8
aureus carriage in the Hajj context, showing a low rate of MRSA carriage [5].
SC
RI PT
2
The aim of the study was to assess the S. aureus nasal carriage dynamics and
10
genotypic diversity among a cohort of French pilgrims departing from Marseille, France, to
11
Mecca, during the 2012 Hajj season.
12
M AN U
9
Nasal swabs were systematically taken from each participant in the month before departing for the Kingdom of Saudi Arabia (KSA) (named pre-Hajj samples) and in the 3
14
days before leaving the KSA (named Hajj samples). In a number of cases, additional nasal
15
samples were taken in the KSA at the onset of respiratory symptoms (also named Hajj
16
samples). More details were described previously [6]. For this study, only pilgrims that had at
17
least two samples including the pre-Hajj sample were included. The total nucleic acids were
18
extracted from nasal swabs as previously described [6] and frozen at -20°C until analysis. No
19
bacterial culture was performed. DNA templates were tested for S. aureus by quantitative
20
PCR targeting the spa gene [7]. Positive nasal samples were sequenced, and spa types defined
21
using BioNumerics software v 6.6 (Applied Maths, Sint-Martens-Latem, Belgium). Overall
22
acquisition of S. aureus nasal carriage during the course of the pilgrimage was defined (i) by
23
negative pre-Hajj and at least one positive Hajj sample or (ii) participants considered to have
24
acquired a new strain (harboring different S. aureus spa types in pre-Hajj and Hajj samples).
25
Detection of respiratory viruses was performed using specific real-time RT-PCR [6].
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13
ACCEPTED MANUSCRIPT 26
Categorical variables were compared using non parametric tests (IBM SPSS V20.0 (Chicago,
27
Illinois). P values < 0.05 were considered significant.
28
Of the 169 participants enrolled in the cohort, 158 (93.5%) were included into the study. The mean age of participants was 59.6 +/- 12.2 years. The sex ratio (male/female) was
30
0.65. Ninety-two pilgrims (58.2%) reported suffering from at least one chronic disease [6],
31
such as diabetes mellitus (n=43). Eighty-five participants received antibiotics for respiratory
32
symptoms during their stay [6]. The carriage rate was 46.2% (n=73) at departure and 36.7%
33
(n=58) at return time. Ninety-eight participants were carriers (Figure 1) : (i) 40 participants
34
were carriers at pre-Hajj sample and subsequently non-carriers; intake of antibiotics in this
35
group was similar to that of other participants (P = 0.82); (ii) 25 participants had a negative
36
pre-Hajj sample and subsequently acquired S. aureus carriage during the course of the
37
pilgrimage ; among them, 4 participants had been sampled twice during the pilgrimage and
38
acquired S. aureus with different spa types; (iii) 33 participants were positive for both pre-
39
and Hajj samples; among them, 21 participants had similar spa types of S. aureus in pre- and
40
Hajj samples and one-third (n=11) had different spa types in pre- and Hajj samples (Figure
41
1). In one case, comparison of spa types from pre- and Hajj samples was not possible as one
42
sequence was incomplete. The overall acquisition of S. aureus was therefore 22.8% (36 out of
43
158 pilgrims). The acquisition of S. aureus was statistically associated with the acquisition of
44
respiratory viruses (10/25 vs 26/133, P= 0.03). Molecular analysis of S. aureus spa gene in
45
pre-Hajj and Hajj samples is summarized in Figure 2. S. aureus acquired belonged mainly to
46
clonal complexes (CC) 45 (21.6%), CC398 (13.5%), CC6 (10.8%) and CC5 (8.1%). CC398
47
was significantly more prevalent in S. aureus strains acquired during the pilgrimage compared
48
to CCs already present at departure (5/37 vs 2/73 RR= 4.93, CI 95%:[1.01-24.2], P=0.04)
49
(Figure 2B).
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29
ACCEPTED MANUSCRIPT 50
This is the first prospective analysis addressing the genetic dynamics of S. aureus nasal carriage at the Hajj where overcrowding provides ideal conditions for person-to-person
52
transmission of infectious agents [3, 6].
53
The overall rate of S. aureus nasal carriage was nearly 60%, a high rate usually observed in
54
longitudinal studies [8]. At a given time (e.g at departure or at return time), the S. aureus
55
observed carriage rate was concordant to literature [1] including in Hajj setting [5].
56
In this study about one-third of participants acquired S. aureus nasal carriage and one-third of
57
carriers, based on spa typing results, acquired a new strain of S. aureus. This overall
58
acquisition rate of carriage was similar to the one recently reported in long-term care facilities
59
in France where overcrowding is often present [9]. Acquisition of S. aureus carriage was
60
found associated with acquisition of respiratory viruses, in accordance with recent data
61
showing the role of pathogen interactions in S. aureus carriage [10]. Dispersal of S. aureus
62
has been shown to be significantly increased after experimental rhinovirus infection [11],
63
rhinovirus being the most frequent virus found in our cohort [6].
SC
M AN U
TE D
64
RI PT
51
We developed an original protocol allowing the identification of S. aureus at the spa type level directly from total nucleic acids purified form the nasal specimens. The typing
66
results revealed that participation to the Hajj increased carriage of the CC398 clone that is
67
emerging, notably in France [12]. This suggests that travels and gatherings may contribute to
68
the spread of this clone. The other spa types identified in pilgrims who acquired S. aureus
69
belonged to various clonal complexes already described in this geographical area [13].
AC C
70
EP
65
Limitations to our study may be considered. First, data about the persistence of S.
71
aureus carriage after the return to France are lacking. We may hypothesize that pilgrims who
72
acquired carriage may become persistent ones as half of them exhibited high bacterial load in
73
nasal samples (not shown) [1]. Second, strains were not available for antimicrobial
ACCEPTED MANUSCRIPT 74
susceptibility testing to better describe the epidemiology of S. aureus acquired during the
75
Hajj.
76
In conclusion, this preliminary study documents a high acquisition rate of S. aureus carriage in pilgrims following participation in the 2012 Hajj, involving notably the emerging
78
S. aureus CC398. The results of this small-scale study illustrate the influence of mass travels
79
in the spread of infectious agents that colonize humans.
80
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77
Conflict of interest: All the authors declare to have no conflict of interest.
82
Acknowledgement. We thank Mr Philippe Michelucci, for English editing of the manuscript.
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ACCEPTED MANUSCRIPT 83
FIGURE 1
87 88 89 90 91 92 93 94 95 96
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86
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85
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FIGURE 2
98 99
B
Spa-type
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Clonal complex (CC) or sequence type (ST)*
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A
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CC1 t127, t693 CC182 t493 CC22 t005, t223, t309, t8506, t852 CC30 t012, t10307 CC398 t5635, t571, Unknown CC45 t015, t026, t031, t095, t230, t331, t550, t6100 CC5 t010, t2164, t688 CC59 t216 CC6 t701 CC7 t091 CC8 t008, t037 CC80 t044 ST126 t605 ST707 t1537 Unknown t11928, t6704, t8099, t8457, t9276, Unknown All * clonal complexes or sequences type were deducted form the spa-type.
100 101
No.(%) of S. aureus isolated in France KSA 2 (2.9) 1 (2.8) 1 (1.4) 0 7 (10.1) 2 (5.7) 6 (8.7) 2 (5.7) 2 (2.9) 5 (14.3) 14 (20.3) 8 (22.8) 1 (1.4) 3(8.6) 1(1.4) 0 1(1.4) 2 (5.7) 1(1.4) 0 4 (5.8) 1 (2.8) 2 (2.9) 2 (5.7) 2 (2.9) 0 1 (1.4) 2 (5.7) 24 (34.8) 7 (20) 69 35
ACCEPTED MANUSCRIPT 102
LEGEND OF FIGURES
103 Figure 1. Flow chart of nasal carriage state in Hajj pilgrims.
105
1
At least one sample positive for S. aureus
106
2
Participants who acquired S. aureus during the course of pilgrimage i.e negative at pre-Hajj
107
sampling and subsequently positive during or after the Hajj.
108
Among these participants, 4 have been sampled twice in KSA and harbored each one
109
different clonal complex at each sampling (i.e: (CC1-CC80; CC6-CC5; CC6-CC398; CC1-
110
undetermined)
111
3
112
spa type recovered from the pre-Hajj sample was undetermined.
113
Figure 2. Dynamics of Staphylococcus aureus nasal carriage in a cohort of pilgrims from
114
France to the Kingdom of Saudi Arabia (Hajj pilgrimage 2012).
115
A. Minimum spanning tree based on the spa repeats of 104 S. aureus DNA including S.
116
aureus acquired during the course of the pilgrimage (red) and S. aureus already carried by
117
pilgrims at departure (blue). Spa-types are indicated in the bubbles. Clusters surrounded in
118
grey correspond to the main clonal complexes (CCs) deducted from the spa-types. The bubble
119
sizes are proportional to the number of S. aureus spa types according to the scale given on the
120
Figure. The distance between S. aureus spa types corresponds to the number of different
121
repeats: bold line (1), line (2), dotted line (3), grey dotted line (4), light grey dotted line (>4).
122
* This bubble includes one sequence belonging to the spa-type t095. ** This bubble includes
123
one sequence belonging to the spa-type t5635. *** This bubble includes one sequence
124
belonging to the spa-type t6704. Six S. aureus spa types with incomplete sequence were not
125
included in the Figure.
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104
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The sequence of the S. aureus DNA recovered in the Hajj sample was incomplete and the
ACCEPTED MANUSCRIPT 126
B. Distribution of clonal complexes or sequence types deducted from the spa-types identified
127
in pre-Hajj (France) and Hajj samples (KSA) on the spa repeats of 104 S. aureus DNA.
128
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129
ACCEPTED MANUSCRIPT 130
REFERENCES
131
Reference List
132 (1) Verhoeven PO, Gagnaire J, Botelho-Nevers E, et al. Detection and clinical relevance of Staphylococcus aureus nasal carriage: an update. Expert Rev Anti Infect Ther 2014; 12: 75-89.
136 137
(2) Zhou YP, Wilder-Smith A, Hsu LY. The role of international travel in the spread of methicillin-resistant Staphylococcus aureus. J Travel Med 2014; 21: 272-81.
138 139
(3) Ahmed QA, Arabi YM, Memish ZA. Health risks at the Hajj. Lancet 2006; 367:100815.
140 141 142
(4) Clements A, Halton K, Graves N, et al. Overcrowding and understaffing in modern health-care systems: key determinants in meticillin-resistant Staphylococcus aureus transmission. Lancet Infect Dis 2008; 8: 427-34.
143 144 145
(5) Memish ZA, Balkhy HH, Almuneef MA, Al-Haj-Hussein BT, Bukhari AI, Osoba AO. Carriage of Staphylococcus aureus among Hajj pilgrims. Saudi Med J 2006; 27: 136772.
146 147
(6) Benkouiten S, Charrel R, Belhouchat K, et al. Circulation of respiratory viruses among pilgrims during the 2012 Hajj pilgrimage. Clin Infect Dis 2013; 57:992-1000.
148 149 150
(7) Stegger M, Andersen PS, Kearns A, et al. Rapid detection, differentiation and typing of methicillin-resistant Staphylococcus aureus harbouring either mecA or the new mecA homologue mecA(LGA251). Clin Microbiol Infect 2012; 18:395-400.
151 152 153
(8) Muthukrishnan G, Lamers RP, Ellis A, et al. Longitudinal genetic analyses of Staphylococcus aureus nasal carriage dynamics in a diverse population. BMC Infect Dis 2013; 13:221.
154 155 156
(9) Couderc C, Jolivet S, Thiebaut AC, et al. Fluoroquinolone use is a risk factor for methicillin-resistant Staphylococcus aureus acquisition in long-term care facilities: a nested case-case-control study. Clin Infect Dis 2014; 59: 206-15.
157 158 159 160
(10) Rodrigues F, Foster D, Nicoli E, et al. Relationships between rhinitis symptoms, respiratory viral infections and nasopharyngeal colonization with Streptococcus pneumoniae, Haemophilus influenzae and Staphylococcus aureus in children attending daycare. Pediatr Infect Dis J 2013; 32: 227-32.
161 162 163
(11) Bassetti S, Bischoff WE, Walter M, et al. Dispersal of Staphylococcus aureus into the air associated with a rhinovirus infection. Infect Control Hosp Epidemiol 2005; 26:196-203.
164 165 166
(12) Valour F, Tasse J, Trouillet-Assant S, et al. Methicillin-susceptible Staphylococcus aureus clonal complex 398: high prevalence and geographical heterogeneity in bone and joint infection and nasal carriage. Clin Microbiol Infect 2014; 20: O772-5.
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133 134 135
ACCEPTED MANUSCRIPT 167 168 169 170
(13) Monecke S, Ehricht R, Slickers P, et al. Microarray-based genotyping of Staphylococcus aureus isolates from camels. Vet Microbiol 2011; 150 :309-14.
171
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172 173 174
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175 176
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S1198-743X(15)00384-5
DOI:
10.1016/j.cmi.2015.03.020
Reference:
CMI 224
To appear in:
Clinical Microbiology and Infection
Received Date: 18 January 2015 Revised Date:
22 March 2015
Accepted Date: 26 March 2015
Please cite this article as: Verhoeven PO, Gautret P, Haddar CH, Benkouiten S, Gagnaire J, Belhouchat K, Grattard F, Charrel R, Pozzetto B, Drali T, Lucht F, Brouqui P, Memish ZA, Berthelot P, BotelhoNevers E, Molecular dynamics of Staphylococcus aureus nasal carriage in Hajj pilgrims, Clinical Microbiology and Infection (2015), doi: 10.1016/j.cmi.2015.03.020. This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
ACCEPTED MANUSCRIPT Molecular dynamics of Staphylococcus aureus nasal carriage in Hajj pilgrims
Paul O. Verhoeven1,2, Philippe Gautret 3, Cyrille H. Haddar 2, Samir Benkouiten3, Julie Gagnaire1, Khadidja Belhouchat3, Florence Grattard1,2, Rémi Charrel3, Bruno Pozzetto1,2,
RI PT
Tassadit Drali3, Frédéric Lucht1,4, Philippe Brouqui3, Ziad A. Memish5 , Philippe Berthelot1,2,4 and Elisabeth Botelho-Nevers1,4
GIMAP EA 3064 (Groupe Immunité des Muqueuses et Agents Pathogènes), University of
SC
1
Lyon, 42023 Saint-Etienne, France
Laboratory of Infectious Agents and Hygiene, University Hospital of Saint-Etienne, 42055
Saint-Etienne Cedex 02, France 3
M AN U
2
Aix Marseille Université, URMITE, UM63, CNRS 7278, IRD 198, Inserm 1095 and Institut
Hospitalo-Universitaire Méditerranée Infection, Marseille, France
Infectious Diseases Department, University Hospital of Saint-Etienne, 42055 Saint-Etienne
TE D
4
Cedex 02, France 5
Public Health Directorate, Saudi Ministry of Health, World Health Organization
EP
Collaborating Center for Mass Gathering Medicine and College of Medicine, Alfaisal
AC C
University, Riyadh, Kingdom of Saudi Arabia
Running title: Staphylococcus aureus carriage in Hajj Keywords: Staphylococcus aureus carriage; carriage acquisition; Hajj; overcrowding; respiratory viruses.
Corresponding author:
E. Botelho-Nevers: [email protected] Tél: + (33) 4 77 12 77 89 Fax: + (33) 4 77 12 78 24
ACCEPTED MANUSCRIPT ABSTRACT
During the 2012 Hajj season, the acquisition risk of Staphylococcus aureus nasal carriage in a
RI PT
cohort of French pilgrims was 22.8%, statistically associated with the acquisition of viral respiratory pathogens (P =0.03). The carriage of S. aureus belonging to the emerging clonal
AC C
EP
TE D
M AN U
SC
complex 398 significantly increased following the pilgrimage (P < 0.05).
ACCEPTED MANUSCRIPT 1
Staphylococcus aureus nasal carriage is common and virtually all individuals could become carrier during their live [1]. Intercontinental travels play a significant role in the
3
spread of specific S. aureus clones such as previously shown for the CA-MRSA USA300
4
clone [2]. The Hajj Muslim pilgrimage to Mecca is the largest annual mass gathering in the
5
world; the overcrowding is major, leading to a high risk of transmission of pathogens between
6
pilgrims [3]. Transmission of S. aureus has been largely described in household and hospital
7
settings associated with overcrowding [4]. To our knowledge, only one study examined S.
8
aureus carriage in the Hajj context, showing a low rate of MRSA carriage [5].
SC
RI PT
2
The aim of the study was to assess the S. aureus nasal carriage dynamics and
10
genotypic diversity among a cohort of French pilgrims departing from Marseille, France, to
11
Mecca, during the 2012 Hajj season.
12
M AN U
9
Nasal swabs were systematically taken from each participant in the month before departing for the Kingdom of Saudi Arabia (KSA) (named pre-Hajj samples) and in the 3
14
days before leaving the KSA (named Hajj samples). In a number of cases, additional nasal
15
samples were taken in the KSA at the onset of respiratory symptoms (also named Hajj
16
samples). More details were described previously [6]. For this study, only pilgrims that had at
17
least two samples including the pre-Hajj sample were included. The total nucleic acids were
18
extracted from nasal swabs as previously described [6] and frozen at -20°C until analysis. No
19
bacterial culture was performed. DNA templates were tested for S. aureus by quantitative
20
PCR targeting the spa gene [7]. Positive nasal samples were sequenced, and spa types defined
21
using BioNumerics software v 6.6 (Applied Maths, Sint-Martens-Latem, Belgium). Overall
22
acquisition of S. aureus nasal carriage during the course of the pilgrimage was defined (i) by
23
negative pre-Hajj and at least one positive Hajj sample or (ii) participants considered to have
24
acquired a new strain (harboring different S. aureus spa types in pre-Hajj and Hajj samples).
25
Detection of respiratory viruses was performed using specific real-time RT-PCR [6].
AC C
EP
TE D
13
ACCEPTED MANUSCRIPT 26
Categorical variables were compared using non parametric tests (IBM SPSS V20.0 (Chicago,
27
Illinois). P values < 0.05 were considered significant.
28
Of the 169 participants enrolled in the cohort, 158 (93.5%) were included into the study. The mean age of participants was 59.6 +/- 12.2 years. The sex ratio (male/female) was
30
0.65. Ninety-two pilgrims (58.2%) reported suffering from at least one chronic disease [6],
31
such as diabetes mellitus (n=43). Eighty-five participants received antibiotics for respiratory
32
symptoms during their stay [6]. The carriage rate was 46.2% (n=73) at departure and 36.7%
33
(n=58) at return time. Ninety-eight participants were carriers (Figure 1) : (i) 40 participants
34
were carriers at pre-Hajj sample and subsequently non-carriers; intake of antibiotics in this
35
group was similar to that of other participants (P = 0.82); (ii) 25 participants had a negative
36
pre-Hajj sample and subsequently acquired S. aureus carriage during the course of the
37
pilgrimage ; among them, 4 participants had been sampled twice during the pilgrimage and
38
acquired S. aureus with different spa types; (iii) 33 participants were positive for both pre-
39
and Hajj samples; among them, 21 participants had similar spa types of S. aureus in pre- and
40
Hajj samples and one-third (n=11) had different spa types in pre- and Hajj samples (Figure
41
1). In one case, comparison of spa types from pre- and Hajj samples was not possible as one
42
sequence was incomplete. The overall acquisition of S. aureus was therefore 22.8% (36 out of
43
158 pilgrims). The acquisition of S. aureus was statistically associated with the acquisition of
44
respiratory viruses (10/25 vs 26/133, P= 0.03). Molecular analysis of S. aureus spa gene in
45
pre-Hajj and Hajj samples is summarized in Figure 2. S. aureus acquired belonged mainly to
46
clonal complexes (CC) 45 (21.6%), CC398 (13.5%), CC6 (10.8%) and CC5 (8.1%). CC398
47
was significantly more prevalent in S. aureus strains acquired during the pilgrimage compared
48
to CCs already present at departure (5/37 vs 2/73 RR= 4.93, CI 95%:[1.01-24.2], P=0.04)
49
(Figure 2B).
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29
ACCEPTED MANUSCRIPT 50
This is the first prospective analysis addressing the genetic dynamics of S. aureus nasal carriage at the Hajj where overcrowding provides ideal conditions for person-to-person
52
transmission of infectious agents [3, 6].
53
The overall rate of S. aureus nasal carriage was nearly 60%, a high rate usually observed in
54
longitudinal studies [8]. At a given time (e.g at departure or at return time), the S. aureus
55
observed carriage rate was concordant to literature [1] including in Hajj setting [5].
56
In this study about one-third of participants acquired S. aureus nasal carriage and one-third of
57
carriers, based on spa typing results, acquired a new strain of S. aureus. This overall
58
acquisition rate of carriage was similar to the one recently reported in long-term care facilities
59
in France where overcrowding is often present [9]. Acquisition of S. aureus carriage was
60
found associated with acquisition of respiratory viruses, in accordance with recent data
61
showing the role of pathogen interactions in S. aureus carriage [10]. Dispersal of S. aureus
62
has been shown to be significantly increased after experimental rhinovirus infection [11],
63
rhinovirus being the most frequent virus found in our cohort [6].
SC
M AN U
TE D
64
RI PT
51
We developed an original protocol allowing the identification of S. aureus at the spa type level directly from total nucleic acids purified form the nasal specimens. The typing
66
results revealed that participation to the Hajj increased carriage of the CC398 clone that is
67
emerging, notably in France [12]. This suggests that travels and gatherings may contribute to
68
the spread of this clone. The other spa types identified in pilgrims who acquired S. aureus
69
belonged to various clonal complexes already described in this geographical area [13].
AC C
70
EP
65
Limitations to our study may be considered. First, data about the persistence of S.
71
aureus carriage after the return to France are lacking. We may hypothesize that pilgrims who
72
acquired carriage may become persistent ones as half of them exhibited high bacterial load in
73
nasal samples (not shown) [1]. Second, strains were not available for antimicrobial
ACCEPTED MANUSCRIPT 74
susceptibility testing to better describe the epidemiology of S. aureus acquired during the
75
Hajj.
76
In conclusion, this preliminary study documents a high acquisition rate of S. aureus carriage in pilgrims following participation in the 2012 Hajj, involving notably the emerging
78
S. aureus CC398. The results of this small-scale study illustrate the influence of mass travels
79
in the spread of infectious agents that colonize humans.
80
RI PT
77
Conflict of interest: All the authors declare to have no conflict of interest.
82
Acknowledgement. We thank Mr Philippe Michelucci, for English editing of the manuscript.
AC C
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SC
81
ACCEPTED MANUSCRIPT 83
FIGURE 1
87 88 89 90 91 92 93 94 95 96
EP
86
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85
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ACCEPTED MANUSCRIPT 97
FIGURE 2
98 99
B
Spa-type
EP
Clonal complex (CC) or sequence type (ST)*
TE D
M AN U
SC
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A
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CC1 t127, t693 CC182 t493 CC22 t005, t223, t309, t8506, t852 CC30 t012, t10307 CC398 t5635, t571, Unknown CC45 t015, t026, t031, t095, t230, t331, t550, t6100 CC5 t010, t2164, t688 CC59 t216 CC6 t701 CC7 t091 CC8 t008, t037 CC80 t044 ST126 t605 ST707 t1537 Unknown t11928, t6704, t8099, t8457, t9276, Unknown All * clonal complexes or sequences type were deducted form the spa-type.
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No.(%) of S. aureus isolated in France KSA 2 (2.9) 1 (2.8) 1 (1.4) 0 7 (10.1) 2 (5.7) 6 (8.7) 2 (5.7) 2 (2.9) 5 (14.3) 14 (20.3) 8 (22.8) 1 (1.4) 3(8.6) 1(1.4) 0 1(1.4) 2 (5.7) 1(1.4) 0 4 (5.8) 1 (2.8) 2 (2.9) 2 (5.7) 2 (2.9) 0 1 (1.4) 2 (5.7) 24 (34.8) 7 (20) 69 35
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LEGEND OF FIGURES
103 Figure 1. Flow chart of nasal carriage state in Hajj pilgrims.
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1
At least one sample positive for S. aureus
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2
Participants who acquired S. aureus during the course of pilgrimage i.e negative at pre-Hajj
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sampling and subsequently positive during or after the Hajj.
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Among these participants, 4 have been sampled twice in KSA and harbored each one
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different clonal complex at each sampling (i.e: (CC1-CC80; CC6-CC5; CC6-CC398; CC1-
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undetermined)
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3
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spa type recovered from the pre-Hajj sample was undetermined.
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Figure 2. Dynamics of Staphylococcus aureus nasal carriage in a cohort of pilgrims from
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France to the Kingdom of Saudi Arabia (Hajj pilgrimage 2012).
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A. Minimum spanning tree based on the spa repeats of 104 S. aureus DNA including S.
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aureus acquired during the course of the pilgrimage (red) and S. aureus already carried by
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pilgrims at departure (blue). Spa-types are indicated in the bubbles. Clusters surrounded in
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grey correspond to the main clonal complexes (CCs) deducted from the spa-types. The bubble
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sizes are proportional to the number of S. aureus spa types according to the scale given on the
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Figure. The distance between S. aureus spa types corresponds to the number of different
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repeats: bold line (1), line (2), dotted line (3), grey dotted line (4), light grey dotted line (>4).
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* This bubble includes one sequence belonging to the spa-type t095. ** This bubble includes
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one sequence belonging to the spa-type t5635. *** This bubble includes one sequence
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belonging to the spa-type t6704. Six S. aureus spa types with incomplete sequence were not
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included in the Figure.
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The sequence of the S. aureus DNA recovered in the Hajj sample was incomplete and the
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B. Distribution of clonal complexes or sequence types deducted from the spa-types identified
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in pre-Hajj (France) and Hajj samples (KSA) on the spa repeats of 104 S. aureus DNA.
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REFERENCES
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Reference List
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(3) Ahmed QA, Arabi YM, Memish ZA. Health risks at the Hajj. Lancet 2006; 367:100815.
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(4) Clements A, Halton K, Graves N, et al. Overcrowding and understaffing in modern health-care systems: key determinants in meticillin-resistant Staphylococcus aureus transmission. Lancet Infect Dis 2008; 8: 427-34.
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(5) Memish ZA, Balkhy HH, Almuneef MA, Al-Haj-Hussein BT, Bukhari AI, Osoba AO. Carriage of Staphylococcus aureus among Hajj pilgrims. Saudi Med J 2006; 27: 136772.
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(12) Valour F, Tasse J, Trouillet-Assant S, et al. Methicillin-susceptible Staphylococcus aureus clonal complex 398: high prevalence and geographical heterogeneity in bone and joint infection and nasal carriage. Clin Microbiol Infect 2014; 20: O772-5.
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(13) Monecke S, Ehricht R, Slickers P, et al. Microarray-based genotyping of Staphylococcus aureus isolates from camels. Vet Microbiol 2011; 150 :309-14.
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