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PLOS One logoLink to PLOS One
. 2023 May 18;18(5):e0285879. doi: 10.1371/journal.pone.0285879

Long lasting anti-IgG chikungunya seropositivity in the Mayotte population will not be enough to prevent future outbreaks: A seroprevalence study, 2019

Giuseppina Ortu 1,2,*, Gilda Grard 3,4, Fanny Parenton 5, Marc Ruello 1, Marie-Claire Paty 1, Guillaume André Durand 4, Youssouf Hassani 5, Henriette De Valk 1, Harold Noël 1; Unono Wa Maore group
Editor: Joel Mossong6
PMCID: PMC10194921  PMID: 37200250

Abstract

Chikungunya is an arboviral disease causing arthralgia which may develop into a debilitating chronic arthritis. In Mayotte, a French overseas department in the Indian Ocean, a chikungunya outbreak was reported in 2006, affecting a third of the population. We aimed at assessing the chikungunya seroprevalence in this population, after over a decade from that epidemic. A multi-stage cross sectional household-based study exploring socio-demographic factors, and knowledge and attitude towards mosquito-borne disease prevention was carried out in 2019. Blood samples from participants aged 15–69 years were taken for chikungunya IgG serological testing. We analyzed associations between chikungunya serological status and selected factors using Poisson regression models, and estimated weighted and adjusted prevalence ratios (w/a PR). The weighted seroprevalence of chikungunya was 34.75% (n = 2853). Seropositivity for IgG anti-chikungunya virus was found associated with living in Mamoudzou (w/a PR = 1.49, 95%CI: 1.21–1.83) and North (w/a PR = 1.41, 95%CI: 1.08–1.84) sectors, being born in the Comoros islands (w/a PR = 1.30, 95%CI: 1.03–1.61), being a student or unpaid trainee (w/a PR = 1.35, 95%CI: 1.01–1.81), living in precarious housing (w/a PR = 1.30, 95%CI: 1.02–1.67), accessing water streams for bathing (w/a PR = 1.72, 95%CI: 1.1–2.7) and knowing that malaria is a mosquito-borne disease (w/a PR = 1.42, 95%CI: 1.21–1.83). Seropositivity was found inversely associated with high education level (w/a PR = 0.50, 95%CI: 0.29–0.86) and living in households with access to running water and toilets (w/a PR = 0.64, 95%CI: 0.51–0.80) (n = 1438). Our results indicate a long-lasting immunity from chikungunya exposure. However, the current population seroprevalence is not enough to protect from future outbreaks. Individuals naïve to chikungunya and living in precarious socio-economic conditions are likely to be at high risk of infection in future outbreaks. To prevent and prepare for future chikungunya epidemics, it is essential to address socio-economic inequalities as a priority, and to strengthen chikungunya surveillance in Mayotte.

Introduction

Chikungunya virus (CHIKV) is an RNA virus from the Togaviridae family transmitted by Aedes spp mosquitoes. Chikungunya disease (CHIK) is responsible for acute febrile illness which can be misdiagnosed as dengue, with which it shares the same main vectors. Classical clinical presentation includes headache, nausea, fatigue, muscle pain, rash and joint swelling, but the most relevant is arthralgia that may disappear within two weeks, (may) persist for months or develop into debilitating chronic arthritis [13]. Atypical and severe forms of CHIKV infection with neurological implications have been reported in hospitalized patients [4], as well as in newborns subsequent to peripartum mother-to-infant transmission [5]. Immunity develops after infection, and protects the individual from future infections [6]. To date, no licensed vaccines are available, and treatment is only symptomatic.

CHIK outbreaks have been reported mainly in Asia, Africa, and recently, in the Americas and even in Southern Europe, hence becoming a relevant emerging threat in the northern hemisphere [1]. In Africa, where presumably CHIKV originated [7], outbreaks have been reported since 1952, and specifically in the west and central regions of the continent. In 2005–2006, a CHIKV epidemic that started in Kenya spread to the islands of the Indian Ocean, [811], among them, Mayotte, a densely populated French overseas department, where 37.2% of the population were affected [12].

After the 2005–2006 massive outbreak, a surveillance system for chikungunya and dengue viruses was set up in 2008 in Mayotte [13], based on the systematic screening of patients with dengue-like syndromes for malaria by rapid diagnostic test [14] and chikungunya, dengue and leptospirosis by PCR. Although a dedicated surveillance system has been in place in the last decade, no autochthonous CHIKV infections have been reported in Mayotte since 2008 [8, 1517].

Though immunity in those with past infection is long-lasting [6] and maybe lifelong, CHIKV foci of infection or large scale outbreaks are becoming likelier to occur as the proportion of immune people declines [18]. Indeed, in the last decade, Mayotte has seen a fast and dynamic population growth due to high birthrates and to immigration, to a lesser extent [18]. Because of this context of migrations, the level of immunity in the population might have changed.

After the last epidemic in 2006 [12], no further studies to assess seroprevalence were performed. Considering the dynamic changes of the population in the recent years, a seroprevalence study was carried out to assess the level of herd immunity in the population after more than a decade.

Santé Publique France set up in 2018 a general population survey Unono Wa Maore (“About the Mahoran”) to assess the population health needs and set up appropriate prevention and health promotion campaigns. For this study, blood samples were collected, and made available for a wide range of analyses, providing the opportunity to assess the seroprevalence of various arbovirus infections, among them CHIKV. Alongside biological sample data collection, participants were interviewed to collect socio-demographic information, and information on attitude and practices with regard to the prevention of vector-borne diseases.

Here, we estimated the seroprevalence of past CHIKV infection in the population of Mayotte in 2018–2019. We also identified sociodemographic factors for CHIKV seropositivity, and described the association between attitudes and practices in arboviral disease prevention, and CHIKV seroprevalence. These results provide an insight into factors that could be considered as relevant to predict, prevent, control and manage future CHIKV outbreaks in Mayotte.

Materials and methods

Setting

Mayotte is a densely populated island of 374 km2 with roughly 250,000 people; generally hot and humid, it is characterized by high rainfall, especially in the center-west and north parts of the island. Currently, roughly half of the Mahoran population holds a foreign nationality and is below age 18 [18]. Foreign population, especially coming from the Comoros islands, and high population growth rate have been particularly evident in the north-eastern part of the Island. Although the Mahoran population is in rapid growth, the number of young adults, especially between age 15 and 24, is decreasing due to the migratory outward flow of this generation towards metropolitan France [19].

The majority of the Mahoran population is poor and lives in precarious conditions, e.g. in inadequate housing; this situation has been particularly worrying for immigrants coming from neighboring islands [20, 21]. In 2017, roughly 39% of the population was reported to live in metal sheet, wood, vegetation or mud housing, and without access to running water or toilets inside their house [19, 20]. The population with these precarious living conditions is especially concentrated in Mamoudzou and surrounding communes [22], and some villages in the north [23].

Study design

The Unono wa Maore study, conducted between December 2018 and June 2019, consists of a cross-sectional household-based study exploring health status, social determinants of health, vaccination coverage in children, behaviors and attitudes toward sex, prevalence of major chronic diseases and sexually transmitted diseases, childcare and mosquito-borne disease prevention.

We used a multi-stage cluster sampling design to select study participants. First, we randomly selected addresses of buildings from the 2017 register of localized buildings (RIL), which contains all the housing addresses necessary for the population census. We stratified addresses by number of households: (i) one household, (ii) 2 to 9 households, (iii) 10 or more. When there were up to 10 households per address, all households were included. Otherwise, a maximum of 10 households were randomly selected. In each household, we randomly selected a maximum of five individuals of which 3 individuals were aged 15 years and older. Further details on the methodology can be found elsewhere [24].

Data collection

Trained interviewers conducted face-to-face questionnaire interviews in French and two local languages, Kibushi and Shimaore.

Two questionnaires were administered: one comprehensive questionnaire for the randomly selected first participant aged 15 and older included in the household, and a short one for the other participants of the same household. Both questionnaires explored participants’ socio-demographic characteristics; alcohol, tobacco, and drugs use; perceived health; hygienic practices; vaccination; perception and behavior related to vector borne diseases. Only the comprehensive questionnaire collected data on family origin, socio-economic status and education, healthcare seeking behavior; food intake, and sexual behavior. Additionally, interviewers collected information on the type of dwelling as well as access to water.

Following the interviews, participants made an appointment with a nurse for anthropometric measurements and collection of biological samples. Only participants over 15 years old were asked to provide biological samples, and only those aged 15–69 years were tested for past CHIK infection.

Laboratory analysis

Blood Samples of 5 mL collected by venipuncture were frozen at –20°C and subsequently defrosted for CHIKV total IgG antibodies testing using an in-house direct ELISA on inactivated viruses (in-house prepared antigens) [25]. The threshold for ELISA positivity was set at a ratio of measured optical density to negative antigen >3.

The National Reference Center for arboviral diseases in Marseille (Institut de Recherche biomédicale des Armées) conducted all serologic testing.

Statistical analysis

Statistical analyses were performed with STATA 16.0 (Statacorp. College Station, TX, USA) using the “svy” command to account for the study design. Age, gender, and location-adjusted weights were calculated and used to estimate IgG weighted Prevalence Ratios (wPR), and their 95% confidence intervals for each characteristic (sociodemographic, knowledge, attitude to personal protection and vector control, and health status). We opted to report the weighted seroprevalence by sectors, a group of communes based on an administrative division already in place, with socio-demographic and logistic similarities, and existing road links. In S1 File. Map of Mayotte with communes and corresponding sectors on the right table, we listed the communes grouped by sectors.

We estimated anti CHIKV IgG seroprevalence among all individuals aged 15 to 69 that provided a blood sample. For the descriptive analysis and bi- and multivariable analysis, we used the data obtained from all participants who answered to the comprehensive questionnaire.

For the univariate analysis (binomial regression), we used anti CHIKV IgG seropositivity as binary outcome and calculated weighted Prevalence Ratios (svy-weighted analysis).

We tested independent associations between serological status as binary outcome and explanatory variables using a Poisson model with robust variance, and computed weighted and adjusted prevalence ratios (w/a PR). We performed three different Poisson models exploring (i) all variables selected, (ii) sociodemographic factors only, (iii) and environmental/housing and knowledge and attitude on prevention and control of arboviral diseases. In these models, we included explanatory variables that were associated with seropositivity in the univariable analysis with a P-value ≤0.20. Collinearity was checked via variance inflation factor (VIF) and Spearman’s rank correlation test. Although the VIF test provided an average value of 1.62 (therefore not suggesting collinearity), a few variables appeared to have slightly higher VIF values (around 2–3); therefore, we opted to run the Spearman’s test. For the latter, when two variables had a pairwise correlation coefficient >0.5 [26], we combined them and reduced the number of categories to reach a final list of variables to be used in the three multivariable models (see also S6 File. Information on datasets, weights and variables used in the analysis, for further clarifications). In each of these models, a backward stepwise elimination [27] and retention procedure of covariables with a P value< 0.05 with a svy-adjusted Wald test was used to finalize each model. Interactions among variables were also explored.

Ethics statement

The UNONO WA MAORE study is interventional research involving the human being with minimal risks and constraints and with a public interest purpose (L.1121-1 2° of the public health code). Therefore, the methodology of the study complies with the provisions of the public health code relating to this type of research and in accordance with the Declaration of Helsinki.

The protocol was validated by the French ethical committee for biomedical research, the Committee for the Protection of Persons (CPP, no. 2017-A02782-51), and complied with MR001 reference methodology (agreement from the National Commission for Informatics and Freedoms of 25 September 2018, no. 918233.

Information on the study and consent forms were read to the participants and a written informed consent was obtained from them or from a legal representative when participants were ≤17 years old. Samples and data were anonymized at the time of collection; therefore, sample testing and data analysis were conducted anonymously [24].

Results

Household and population characteristics

The survey was performed in all communes of the island. 2716 households were included in the survey, and 4817 individuals between the age of 15 and 69 accepted to participate, of whom 2853 were tested for past CHIKV infection. Among those tested for past CHIKV infection, 2778 answered the questionnaires among whom 1438 answered the comprehensive questionnaire.

Participants tested for CHIKV (n = 2778) had a median age of 34 years (min 15, max 69) and were mainly between age 30 and 49 (43.5%), and female (63.8%). Most of them resided in the Mamoudzou sector (36.5%) (Table 1).

Table 1. Weighed seroprevalence of chikungunya by age, sex and residence sector.

Characteristics Total   CHIK positive N CHIK positive (%) IgGCHIK (+) Weighed prevalence (%) 95% CI
Tested for arboviral exposure   2853   1007 35.30 34.75 32.32 37.26
N tested and interviewed* 2778 N % CHIK positive n CHIK positive (%) IgGCHIK (+) Weighed prevalence (%) 95% CI
Age (Years) 15–17 353 12.71 112 31.73 31.43 25.78 37.69
18–29 716 25.77 257 35.89 33.55 29.32 38.06
30–49 1207 43.45 443 36.70 36.12 32.62 39.78
50–69 502 18.07 165 32.87 36.02 31.08 41.26
Sex Male 1007 36.25 365 36.25 36.26 32.63 40.06
Female 1771 63.75 612 34.56 33.45 30.59 36.45
Sector of residence Centre 773 27.83 252 32.60 31.59 27.72 35.73
Mamoudzou 1015 36.54 439 43.25 41.78 37.83 45.83
North 255 9.18 103 40.39 40.85 33.63 48.5
Petite-Terre 442 15.91 126 28.51 29.93 25.03 35.33
South 293 10.55 57 19.45 17.19 12.44 23.29

*Short questionnaire only

Among those who answered the comprehensive questionnaire (n = 1438), the median age was 35 years, (min 15, max 69), 38.4% (n = 552) were born in Mayotte and 52.5% (n = 754) in the Comoros Islands. 63.6% (n = 913) did not receive any type of education, 74.7% (n = 1072) were unemployed, or pensioners or students, 38.6% (n = 555) did not have any social security and 41.0% (n = 589) lived in a household with ≥4 children (Table 1). Housing was 40.4% (n = 581) traditional wooden houses (called “bangas”) or wood-sheet metal hut, 49.5% (n = 712) did not have running water and 51.7% (n = 744) did not have toilets inside the premises. Additional information is available in S2 File. Weighed chikungunya seroprevalence, weighed prevalence ratios and univariate analysis for all covariates.

Overall anti-CHIKV IgG seroprevalence and seroprevalence by age, sex and sector

IgG positive to CHIKV were found in 35.3% (n = 1007) of those tested (n = 2853). The overall weighted seroprevalence of anti-CHIKV IgG in Mayotte was 34.75% (95%IC [32.3–37.3], n = 2853) (Table 1). In those tested and having completed a questionnaire, there were no significant differences in seroprevalence between the different age groups nor by sex. Mamoudzou and North were the sectors with the highest (weighted) seroprevalence (41.8%, 95%CI [37.8–45.8], and 40.9%, 95%CI [33.6–48.5], respectively), and South the one with the lowest (17.2%, 95%CI [12.4–23.3]) (Fig 1).

Fig 1. CHIKV seroprevalence in North, Mamoudzou, Petite-Terre, Center and South sectors of Mayotte.

Fig 1

Determinants of anti-CHIKV IgG seropositivity

For the analysis of anti-CHIKV IgG seropositivity determinants (n = 1438), results from the univariable analysis can be found in S2 File. Weighed chikungunya seroprevalence, weighed prevalence ratios and univariate analysis for all covariates. Among those variables that were significantly associated with past CHIKV exposure, availability of social security or social security plus complementary health insurance were negatively associated with seropositivity. On the contrary, accessing the river for daily washing, being very annoyed by mosquitoes, covering water reservoirs, and smoking habits were all factors associated with anti-CHIKV IgG seropositivity. All these variables were initially added to the multivariable models as found statistically significant in the univariable analysis; however, some of them were subsequently eliminated during the backwards stepwise process as no longer significant in the models. More information on other variables that were tested in the univariable analysis can be found in S2 File. Weighed chikungunya seroprevalence, weighed prevalence ratios and univariate analysis for all covariates.

We performed three models, the first one in which we included all the covariates that from the Spearman test did not appear to create collinearity (M1), and two other models in which we considered only those variables related to specific domains (sociodemographic determinants–M2), and attitude and knowledge related to arboviral disease prevention and vector control and environmental/housing factors (M3) (Table 2).

Table 2. Weighed and adjusted prevalence ratios for CHIKV seropositivity (w/a PR) according to sociodemographic, environmental and attitude factors.

Model 1 includes all covariates, model 2 sociodemographic variables, and model 3 variables related to knowledge /attitudes in arboviral prevention and control and environment/housing factors.

Model 1 (all covariates) (n = 1435) Model 2 (n = 1432) Model 3 (n = 1437)
Characteristics   w/a PR (lincom) 95% CI P value w/a PR 95% CI P value w/a PR 95% CI P value
Sociodemographic factors                        
Age (Years) 15–17 ref ref ref
18–29 1.04 0.74 1.45 0.834 1.16 0.76 1.78 0.48 1.01 0.73 1.41 0.94
30–49 0.93 0.67 1.3 0.673 1.07 0.68 1.7 0.762 0.95 0.69 1.3 0.73
50–69 0.99 0.69 1.43 0.94 1.12 0.69 1.83 0.638 1.01 0.7 1.5 0.934
Sex Male ref ref ref
Female 0.94 0.79 1.11 0.448 0.92 0.76 1.1 0.362 1.01 0.84 1.2 0.939
Sector of residence Centre ref       ref  
Mamoudzou 1.49 1.21 1.83 <10 −4 1.44 1.16 1.78 0.001  
North 1.41 1.08 1.84 0.011 1.45 1.11 1.9 0.007  
Petite-Terre 1.05 0.78 1.4 0.772 1.01 0.75 1.37 0.842  
South 0.50 0.32 0.77 0.002 0.48 0.31 0.75 0.001        
Education level No education ref       ref  
Education below university degree 1.01 0.82 1.24 0.911 0.94 0.75 1.2 0.599  
University degree / higher education 0.50 0.29 0.86 0.012 0.44 0.25 0.77 0.004  
Other education 0.66 0.33 1.32 0.238 0.64 0.32 1.3 0.208        
Professional situation Employed   ref  
Unemployed   1.45 0.96 2.25 0.074  
Student, unpaid trainee / intern   1.35 1.01 1.81 0.041  
Inactive (pensioner, home maker etc,)   1.1 0.84 1.47 0.469  
Work placement, training course, apprenticeship, or paid internship   1.46 0.89 2.4 0.133        
Place of birth Mayotte ref       ref  
Comoros 1.3 1.03 1.61 0.026 1.43 1.15 1.78 0.001  
Madagascar 1.08 0.7 1.68 0.728 1.04 0.64 1.68 0.885  
Metropolitan France and other countries 0.54 0.22 1.32 0.174 0.47 0.19 1.15 0.097        
Household & environment                        
Habitation type House on cement     ref
Traditional wooden house or wood-sheet metal hut     1.30 1.02 1.67 0.035
Apartment                 1.23 0.82 1.85 0.312
Interaction term: Habitation type and access to the river for washing dishes or clothes                 no significant
Access to the river for washing clothes or dishes Never     ref
Sometimes/often for either reason     1.17 0.65 2.1 0.593
Always                 0.53 0.12 2.26 0.391
Running water and WC in the household No ref   ref
Yes 0.64 0.51 0.80 <10 −4         0.61 0.47 0.79 <10 −4
Go to the river for bathing Never           ref
Sometimes/often     1.02 0.66 1.59 0.911
Always                 1.72 1.1 2.7 0.018
Attitudes to mosquito-borne disease prevention                      
Understanding whether transmission of malaria is via mosquitoes No     ref
Yes     1.42 1.03 1.97 0.033
no answer                 1.30 0.9 1.9 0.162

Looking at the model with all the covariates (M1), residing either in Mamoudzou (w/a PR = 1.49, [95%CI: 1.21–1.83]) or North (w/a PR = 1.41 [1.08–1.84]) was associated with higher CHIKV seroprevalence compared to residing in Centre or Petite Terre. Living in the southern sector was associated with the lowest level of CHIKV seroprevalence (w/a PR = 0.49 [0.32–0.77]). Participants originating from Comoros Islands were more likely to be anti-CHIKV IgG positive than those born in Mayotte (w/a PR = 1.30 [1.03–1.61]).

Compared to individuals with no education, persons with university degrees were less likely to have anti-CHIKV IgG (w/a PR = 0.49 [0.29–0.85]). Finally, individuals having running water and a toilet in the household were less likely to have been infected with CHIKV (w/a PR = 0.64 [0.51–0.80]).

M2 exploring socio-demographic covariates/factors, showed that compared to individuals with employment, anti-CHIKV IgG seroprevalence was significantly higher in students (w/a PR = 1.35[1.01–1.81]). In M3 exploring attitudes on mosquito borne diseases prevention and environmental factors, anti-CHIKV IgG seroprevalence was higher among individuals living in traditional wooden houses or wood-sheet metal huts compared to individuals living in houses on cement or apartment (w/a PR = 1.31 [1.01–1.68]). M3 results corroborated the association of high anti-CHIKV IgG seroprevalence with not having running water or a toilet in the household (w/a PR = 0.61 [0.47–0.81]), compared to those individuals with both, as found in M1. M3 model also indicated that going to the river for bathing routinely (w/a PR = 1.72 [1.10–2.7]), and knowing that malaria is transmitted by mosquito bites (w/a PR = 1.42 [1.03–1.97]) were associated with anti-CHIKV IgG seroprevalence.

Discussion

We observed an overall weighed seroprevalence of anti-CHIKV IgG of 35% (95%CI: 32–37) amongst people aged 15 and above. This estimate is very similar to the seroprevalence of anti-CHIKV IgG as found in 2006 just after the outbreak in Mayotte, suggesting that there was no or minimal transmission of the virus in the interim [12]. CHIKV re-emerged in the Indian Ocean in 2009–2010 [28], namely in Reunion island and Madagascar, but not in Mayotte. Since 2006, no further cases of CHIKV infection have been reported in Mayotte [1517, 29], and our findings therefore suggest long-lasting seropositivity against CHIKV in the Mayotte population.

According to our results, 65% of the Mayotte population aged 15 to 69 remains susceptible to CHIKV infection. Although we did not sample children below 15 years, this age group has most likely not been substantially exposed to CHIKV in Mayotte since the last major outbreak in 2005. Pooling together the 44% of the population under 15 years [19] and the estimated number of susceptible individuals, roughly 80% of the Mahoran population remained is susceptible to CHIKV.

It appears, inevitable that new outbreaks could take place ifit is therefore, inevitable the occurrence of future epidemics in Mayotte if CHIK is reintroduced in this population. For this reason, particular attention to CHIKV transmission is warranted to anticipate and prepare for a potential CHIKV introduction and outbreaks in Mayotte.

We studied determinants of CHIKV seropositivity among which sector of residence was an important risk factor for past CHIKV infection. In agreement with a serosurvey performed after the 2006 CHIKV outbreak [30], Mamoudzou and the Northern sector of Mayotte had higher seroprevalence than the center and the mid-west areas (Centre sector). Seroprevalence was lowest in the Southern sector. This geographical distribution of cases may reflect higher past CHIKV transmission in areas with high annual rainfall (northern part of the island), densely populated (northern and north-eastern areas), or urban areas such as Mamaoudzou, Koungou, Dzouadzi and Pamandi.

Geographical area of birth was also significantly associated with anti-CHIKV IgG seroprevalence, with individuals born in the Comoros having higher seroprevalence compared to those born in other geographical areas. This result is in agreement with the higher seroprevalence reported during the CHIKV infection outbreak in these islands in 2005–2006 (over 60%), compared to the seroprevalence reported in Mayotte (around 37%) [9, 30, 31].

Knowledge of other mosquito-borne diseases such as malaria, appeared to be positively associated with the presence of CHIKV antibodies. This association could reflect a heightened awareness of mosquito-borne infectious diseases among individuals affected by malaria or chikungunya.

Other factors associated with the presence of anti-CHIKV IgG were lower education and lack of access to running water or toilets in the house. Models 2 and 3 suggest that living in a traditional wooden house or wood-sheet metal hut, being a student (and unemployed) and always using the river for bathing are all associated with the presence of anti CHIKV antibodies. These results are in agreement with the data obtained in the seroprevalence survey performed in 2006 [30], where factors indicative of socio-economic disadvantage (e.g., length of schooling, makeshift housing) were found associated with anti-CHIKV IgG seropositivity.

Altogether, these results suggest that today’s socio-economically disadvantaged groups that were not exposed to CHIKV infection in 2006, may be a high risk of infection in future CHIKV outbreaks, compared to other groups.

Improvement of housing conditions should be part of CHIK infection prevention, and more broadly, part of prevention strategies for all arboviral diseases. Makeshift and precarious housing (where generally, mosquitoes breeding sites are found more frequently) associated with high population density, increase substantially the chance of transmission of endemic arboviral diseases, making this type of environment prone to disease outbreaks. In Mayotte, several communes in the Mamoudzou sector have the highest number of people living in slums, under precarious socio-economic conditions [32]. Arboviral diseases preparedness plans could target these areas as a priority, as well as people living in similar conditions in the north part of the island, and improvement of housing and environmental conditions should be part of these plans.

Limitations

Our study has some limitations. Data on individuals younger than age 15 and older than 69 were not available, feasibility of sample collection and ethical issues being the main reasons for not including them; hence, the results from this cross sectional seroprevalence study cannot be extrapolated to these age groups. Not including individuals above age 69 (~<1% of the population), has likely not impacted our results as based on data from the last outbreak [12], CHIKV seroprevalence in the population between age 56 and 79 was very similar to other age groups (~ between 23 and 50%). Not including children below age 15, has eliminated a part of the population which was most likely not exposed to CHIKV during the outbreak in 2006. With the addition of this young (naïve) population, the estimated CHIKV seroprevalence is likely to be lower than our estimates.

Although we could estimate CHIKV seroprevalence differences among the different sectors with statistical significance, the study was not designed to determine risk factors at the sector level but only at the department level. For this reason, we could not estimate the association between seroprevalence and the selected determinants in sectors, which could have been more informative and useful from a public health response perspective.

Conclusions

This study has indicated the presence of antibodies against CHIKV in a third of the Mahoran population between the age 15 and 69. This finding implies that two thirds of the adult population is susceptible to CHIKV infection, namely those living under better socio-economic conditions, and children below the age of 15. Our study indicates also that socio-economically disadvantaged groups naïve to CHIKV infection are at higher risk than others in future epidemics.

In this context where the majority of the population is still naïve with regard to CHIKV, and the risk of another epidemic is high due to the continuous transmission of chikungunya virus in other parts of the world, a vaccine would be extremely effective in reducing the risk of infection and future outbreaks. Unfortunately, CHIKV vaccines are still in a development phase and yet to be deployed [33].

Despite the worldwide transmission of chikungunya throughout the tropics, no vaccines are available at the moment. Although planning for chikungunya vaccine trials might be challenging, the global effort to develop and distribute vaccines should be accelerated as core preparedness and response to upcoming CHIK outbreaks [34]. Serosurveys should be conducted whenever possible at least after each CHIK outbreak to monitor its spread and assess population immunity and inform efficient vaccine rollout.

In the absence of an effective vaccine, a viable strategy for preventing CHIKV infections (and other arboviral diseases) is to roll back social inequalities by improving the living conditions in rural, and densely populated urban areas, especially in the center and north part of the island.

Maintaining the screening for CHIKV and other pathogens in febrile patients as it is currently done in Mayotte, is paramount to prepare for future epidemics, as well as insuring a close monitoring of CHIKV circulation in the Indian Ocean region and neighboring African countries. These routine assessments would help in understanding whether CHIKV is circulating, and therefore, informing on whether public health measures should be initiated to prevent further spreading.

Finally, it is of utmost importance to continue to raise awareness among the whole population of Mayotte on how to prevent arboviral infections.

Supporting information

S1 File. Map of Mayotte with communes and corresponding sectors in the table on the right.

(DOCX)

S2 File. Weighed chikungunya seroprevalence and weighed prevalence ratios for all covariates (N = 1438, it may slightly differ for some of the covariates due to missing values).

(DOCX)

S3 File. Dataset CHIK analysis_PLOSOne dataset_n2853.

(DTA)

S4 File. Dataset CHIK analysis_PLOSOne dataset_n2778.

(DTA)

S5 File. Dataset CHIK analysis_PLOSOne dataset_n1438.

(DTA)

S6 File. Information on datasets, weights and variables used in the analysis.

(DOCX)

Acknowledgments

The authors are especially grateful to the investigators of the Unono Wa Maore group: Marc Ruello, Marion Fleury, Jean-Baptiste Richard, Jean-Louis Solet, Laurent Filleul, Delphine Jezewski-Serra, Julie Chesneau

Data Availability

All relevant data are within the paper and its Supporting Information files.

Funding Statement

The author(s) received no specific funding for this work.

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Decision Letter 0

Joel Mossong

3 Feb 2023

PONE-D-22-35076Chikungunya long lasting immunity in the Mayotte population will not be enough to prevent future outbreaks: A seroprevalence study, 2019.PLOS ONE

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5. Review Comments to the Author

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Reviewer #1: The study investigated the susceptibility level of CHIKV in human population in Mayotte using the serological data randomly sampled in the region. They concluded that the immunity in CHIKV is long-lasting and the current immunity from the previous outbreak is not enough to protect from future outbreaks. I found this conclusion a bit inaccurate and may need to revise some discussion points. Some of the analyses in the manuscript should also be revised.

The previous outbreak in Mayotte was in 2005-2006 and the serological data from 2019 was investigated in this study. Unfortunately, the serological data is missing age group < 15, which contains crucial information to know if there are any additional outbreaks in the region 15 years since 2005. The authors were aware of this problem and already discussed it. Besides no report of CHIKV transmission in the region since 2005, by observing that the seroprevalence in the population remains the same compared to the previous study across all age groups, we have a stronger evidence that there were no activity of the virus since 2005. This point should be added to the discussion section.

I suggest the authors should assess for multicollinearity using VIF rather than just collinearity. I also wonder why the threshold of 0.5 was chosen for the pairwise correlation to reduce the number of variables. Why not 0.7 or any other numbers? I suggest the authors attempt to reduce the dimension of the variables by other means such as stepwise regresion.

The results from the univariate analysis should be summarized or discussed somewhere in the manuscript.

I also wonder why the authors perform the analyses on the three models with different fixed set of covariates. Some kind of variables selection procedure here may make the method more robust.

“Based on an estimated basic reproduction number of 2.8 reported for areas where Aedes albopticus is the primary vector (29), roughly 65-70% of the population should be immune to CHIKV to avoid an epidemic; therefore, the possibility of future epidemics in Mayotte cannot be excluded.” I found this point to be problematic. The 65-70% figure is derived from the R0 of 2.8 assuming that the population is homogenous. This in reality is not true for aborviruses, where their transmission dynamic are highly heterogenous. Given the heteogenousity, the herd immunity threshold can be much lower than 65-70%, hence current immunity level in the population maybe enough to prevent epidemic to happen in the near future. Furthermore, the authors seems to ignore the uncertainty of the R0 estimation, which lead to the uncertainty in the herd immunity threshold as well. I also found the conclusion for this point is weak, since in the future, as the susceptible pool is increasing over time from the newborns, it is inevitable that “the possibility of future epidemics in Mayotte cannot be excluded.”

“Knowledge of other mosquito-borne diseases such as malaria, appeared to be positively associated with the presence of CHIKV antibodies. This association could reflect a heightened awareness of mosquito-borne infectious diseases among individuals affected by malaria or chikungunya.” I failed to see the authors’ logic of this point. I believe there is a potential confounder here, knowledge of malaria and presence of CHIKV antibodies are both correlated with high risk of arboviruses infection.

The lines in the manuscript should be numbered.

Reviewer #2: Dear authors,

Excellent work conducting this work and conveying key information about a neglected disease. Please see some specific comments below:

Is it possible to add a map illustrating the different regions of Mayotte and their respective seroprevalence? A figure would be helpful for readers unfamiliar with the island when discussing the geographic findings of the study.

Introduction, 2nd paragraph, line 4, remove “has” in “where presumably CHIKV has originated”

Introduction, final paragraph, line 4, change “insight of” to “insight into”

Setting, paragraph 1, line 3, change “is holding” to “holds”

Study design, paragraph 1, line 1, change “consists in” to “consists of”

Data collection, paragraph 2, line 7, change “Besides,” to “Additionally,”

Determinants of anti-CHIKV IgG seropositivity, paragraph 3, line 1, change “being resident either in Mamoudzou” to “residing in either Mamoudzou”

Determinants of anti-CHIKV IgG seropositivity, paragraph 3, line 2, remove “the” in “associated with the higher”

Determinants of anti-CHIKV IgG seropositivity, paragraph 3, line 3, change “being resident” to “residing”

Determinants of anti-CHIKV IgG seropositivity, paragraph 4, line 3, change “toilette” to “toilet”

Discussion, paragraph 2 - “therefore we should add this population to the group of susceptible individuals.” Is it possible to provide a quantitative estimate of this value? Ie. if you assume all members of the population below 15 are seronegative and calculate a new percentage including these individuals, by how much does the 65% susceptibility estimate increase?

Reviewer #3: Ortu et al. performed a serosurvey and questionnaire in Mayotte, where a CHIKV outbreak took place in 2006. The authors performed IgG ELISA to determine previous CHIKV exposure and applied questionnaires exploring socio-demographic factors and knowledge of mosquito-borne diseases. Although the part of the study looking at CHIKV IgG could be substantially expanded upon, by doing quantitative ELISA, neutralization assays, and T-cell assays to assess the population immunity, the igG data presented here will be a valuable addition to the literature for understanding CHIKV present and future outbreaks.

Major comments:

1.The title of the manuscript claims that the current immunity will not be enough to prevent future outbreaks. This claim is based on a report that estimates that at least 65% of the population would need to be immune to prevent future epidemics. However, in this paper the authors only perform a positive/negative IgG, and do not assess other parts of the immune system that could give partial immunity to CHIKV such as T-cells, and proportion of neutralizing antibodies, cross-neutralizing antibodies from similar alphaviruses. Therefore, the authors should consider changing the title to reflect their data accurately.

2. The authors stored the sera at -20C. Please add how long the sera was stored for. How long are IgGs stable for at -20C?

3. The references need to be improved on several occasions:

•The authors cite a manuscript in preparation; this is highly unusual, and I strongly suggest removing that citation. The details on the methodology should be included in this manuscript, in the methods or supplemental files, or a published source could be cited.

•References 15-17 look like they need more information, is that citing a book chapter?

•While citing reputable websites is acceptable, the authors cite many websites. I would suggest reducing the number of website citations wherever possible and adding the date accessed for any that will be kept. Links to websites tend to break with time and no longer be accessible.

•Reference 31 is all in caps, while the others are not.

4. More details are needed regarding the ELISA. Please clarify if the ELISA is for total IgG, IgG-1 or other. Please specify how the ELISA threshold was chosen. If it was recommended by a manufacturer or based on previous experience or statistical method.

5. Since the authors are working with human blood from a region that may have CHIKV present, the authors should specify how the samples were handled, ie/ level of containment, any heat inactivation done, if anything was done in case blood was collected from a patient with CHIKV symptoms. Although the samples were not known to contain CHIKV, this virus is typically BSL-3 in Europe and the US.

Minor comments:

1. Would it be possible for the authors to use a known sample as standard curve to quantify the IgGs?

2. The "attitudes to mosquito-borne disease prevention" data is not showing on table 2, I think it was cut off.

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Reviewer #1: No

Reviewer #2: No

Reviewer #3: No

**********

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PLoS One. 2023 May 18;18(5):e0285879. doi: 10.1371/journal.pone.0285879.r002

Author response to Decision Letter 0


2 Apr 2023

Manuscript

PONE-D-22-35076

Chikungunya long lasting immunity in the Mayotte population will not be enough to prevent future outbreaks: A seroprevalence study, 2019.

PLOS ONE

REPLY TO REVIEWERS

Reviewer #1:

The study investigated the susceptibility level of CHIKV in human population in Mayotte using the serological data randomly sampled in the region. They concluded that the immunity in CHIKV is long-lasting and the current immunity from the previous outbreak is not enough to protect from future outbreaks. I found this conclusion a bit inaccurate and may need to revise some discussion points. Some of the analyses in the manuscript should also be revised.

The previous outbreak in Mayotte was in 2005-2006 and the serological data from 2019 was investigated in this study. Unfortunately, the serological data is missing age group < 15, which contains crucial information to know if there are any additional outbreaks in the region 15 years since 2005. The authors were aware of this problem and already discussed it. Besides no report of CHIKV transmission in the region since 2005, by observing that the seroprevalence in the population remains the same compared to the previous study across all age groups, we have stronger evidence that there were no activity of the virus since 2005. This point should be added to the discussion section.

Answer:

The authors acknowledge your general feedback – thank you. We mentioned the evidence of not circulation of CHIKV in Mayotte in the discussion - please see lines 288-292.

I suggest the authors should assess for multicollinearity using VIF rather than just collinearity.

Answer:

By using Spearman test we did consider multicollinearity rather than collinearity, as we checked the correlation byrunning the Spearman test with all the variables at once (although the output shows all possible combinations of correlation among each 2 variables).

We did not opt for using the VIF command (https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9901837/) as it is linked to the use of regression. We were using the poisson function, therefore we could not use the command VIF. The command collin could have been used instead as it does not require the regression function beforehand, but we opted for using the Spearman test as it seems to be more appropriate when variables are ordinal, not linear or not following a normal distribution, a situation that pretty much reflected our group of variables.

I also wonder why the threshold of 0.5 was chosen for the pairwise correlation to reduce the number of variables. Why not 0.7 or any other numbers?

Answer:

Based on some reading on Spearman test, the use of it, and the interpretation of thresholds - see for instance these:

https://geographyfieldwork.com/SpearmansRankCalculator.html,

https://statistics.laerd.com/stata-tutorials/spearmans-correlation-using-stata.php

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3576830/

we opted for choosing 0.5 as the threshold. The range between 0.5 and 0.7 is generally associated to moderate correlation. As we did not want to have correlation as a factor that would somehow impact the analysis, 0.5 seemed to be an appropriate and conservative threshold to use (alongside a significant P value).

I suggest the authors attempt to reduce the dimension of the variables by other means such as stepwise regression.

Answer:

We could not conduct an automated stepwise logistic regression using the stepwise command in STATA as the svy prefix command is not compatible with it. However, we did stepwise regression “manually”, specifically we used a backward stepwise process with the commands svy,:poisson and test.

On this note, the svy command was necessary as we could not avoid performing a weighed analysis in this case. https://www.stata.com/support/faqs/statistics/stepwise-regression-with-svy-commands/

The results from the univariate analysis should be summarized or discussed somewhere in the manuscript.

Answer:

We have taken on board your comment and briefly discussed the results from the univariate analysis. Please find the results in lines 241-250.

We opted for not adding any further point related to the univariable analysis in the discussion as not relevant to the most essential results of the study.

I also wonder why the authors perform the analyses on the three models with different fixed set of covariates. Some kind of variables selection procedure here may make the method more robust.

Answer:

After performing the Spearman test and eliminating (or simply recategorizing or combining) variables that were creating collinearity, we added all the remaining variables to one model. By doing the backward stepwise method and reaching a list of variables that were significative in the final model, we realized that we were possibly losing some relevant information that could have been useful to report.

We then run several models with different set of variables, and eventually we reached the conclusion that based on the concept that a model should be parsimonious, it would have been preferable to use and report several models with a smaller number of variables than try to fit all the variables in one single model in which some of the information would have been lost.

Finally, we were not performing prediction models where it would be more appropriate to fit all the variables, but testing potential risk factors that may have been impacting the outcome we were measuring; therefore, (and considering the concept of model parsimony) it would have been preferable not to add all variables at once in the model.

“Based on an estimated basic reproduction number of 2.8 reported for areas where Aedes albopticus is the primary vector (29), roughly 65-70% of the population should be immune to CHIKV to avoid an epidemic; therefore, the possibility of future epidemics in Mayotte cannot be excluded.” I found this point to be problematic. The 65-70% figure is derived from the R0 of 2.8 assuming that the population is homogenous. This in reality is not true for aboviruses, where their transmission dynamic are highly heterogenous. Given the heterogeneity, the herd immunity threshold can be much lower than 65-70%, hence current immunity level in the population maybe enough to prevent epidemic to happen in the near future. Furthermore, the authors seems to ignore the uncertainty of the R0 estimation, which lead to the uncertainty in the herd immunity threshold as well. I also found the conclusion for this point is weak, since in the future, as the susceptible pool is increasing over time from the new-borns, it is inevitable that “the possibility of future epidemics in Mayotte cannot be excluded.”

Answer:

We do agree with your comment and we took off the reference to the Ro. We also added the % of the population under the age 15 that is susceptible and rephrased the paragraph. We also highlighted the resulting expected proportion of susceptible in the population of Mayotte. Please see lines 291-299.

“Knowledge of other mosquito-borne diseases such as malaria, appeared to be positively associated with the presence of CHIKV antibodies. This association could reflect a heightened awareness of mosquito-borne infectious diseases among individuals affected by malaria or chikungunya.” I failed to see the authors’ logic of this point. I believe there is a potential confounder here, knowledge of malaria and presence of CHIKV antibodies are both correlated with high risk of arboviruses infection.

Answer:

Our exposure of interest was “knowledge of other mosquito- borne diseases such as malaria” and the outcome is “CHIKV seropositivity”. This is the relationship we were looking at. A third variable that may have impacted this relationship is the strategies used to fight malaria between 2007 and 2014. During this period, people may have become more aware of malaria transmission and cases as prevention and surveillance for this disease were enhanced. (see this: https://malariajournal.biomedcentral.com/articles/10.1186/s12936-015-0837-6 )

If people had been already impacted by CHIKV infection during the 2005 outbreak, perhaps they may have been keener in understanding more about malaria as well, considering the similarities in transmission. We may infer that the exposure to CHIKV has possibly improved the knowledge of vector borne diseases in the population including malaria. We decided not to report this as it was as looking at the CHIKV as exposure and “knowledge of other mosquito- borne diseases such as malaria” as outcome.

The lines in the manuscript should be numbered.

Answer:

We did add the line number as suggested

Reviewer #2:

Dear authors,

Excellent work conducting this work and conveying key information about a neglected disease. Please see some specific comments below:

Is it possible to add a map illustrating the different regions of Mayotte and their respective seroprevalence? A figure would be helpful for readers unfamiliar with the island when discussing the geographic findings of the study.

Answer:

Thank you for your suggestion, we have added a map with the requested information – see Fig 1.

Introduction, 2nd paragraph, line 4, remove “has” in “where presumably CHIKV has originated”

Introduction, final paragraph, line 4, change “insight of” to “insight into”

Setting, paragraph 1, line 3, change “is holding” to “holds”

Study design, paragraph 1, line 1, change “consists in” to “consists of”

Data collection, paragraph 2, line 7, change “Besides,” to “Additionally,”

Determinants of anti-CHIKV IgG seropositivity, paragraph 3, line 1, change “being resident either in Mamoudzou” to “residing in either Mamoudzou”

Determinants of anti-CHIKV IgG seropositivity, paragraph 3, line 2, remove “the” in “associated with the higher”

Determinants of anti-CHIKV IgG seropositivity, paragraph 3, line 3, change “being resident” to “residing”

Determinants of anti-CHIKV IgG seropositivity, paragraph 4, line 3, change “toilette” to “toilet”

Answer:

Thank you for highlighting all the lines where language changes were needed. We addressed them all.

Discussion, paragraph 2 - “therefore we should add this population to the group of susceptible individuals.” Is it possible to provide a quantitative estimate of this value? Ie. if you assume all members of the population below 15 are seronegative and calculate a new percentage including these individuals, by how much does the 65% susceptibility estimate increase?

Add % of the population 0-15 on top of 65%

Answer:

Thank you for this very useful suggestion. We calculated the % and added to the 65% - Please see line 291-296.

Reviewer #3:

Ortu et al. performed a serosurvey and questionnaire in Mayotte, where a CHIKV outbreak took place in 2006. The authors performed IgG ELISA to determine previous CHIKV exposure and applied questionnaires exploring socio-demographic factors and knowledge of mosquito-borne diseases. Although the part of the study looking at CHIKV IgG could be substantially expanded upon, by doing quantitative ELISA, neutralization assays, and T-cell assays to assess the population immunity, the IgG data presented here will be a valuable addition to the literature for understanding CHIKV present and future outbreaks.

Major comments:

1.The title of the manuscript claims that the current immunity will not be enough to prevent future outbreaks. This claim is based on a report that estimates that at least 65% of the population would need to be immune to prevent future epidemics. However, in this paper the authors only perform a positive/negative IgG, and do not assess other parts of the immune system that could give partial immunity to CHIKV such as T-cells, and proportion of neutralizing antibodies, cross-neutralizing antibodies from similar alphaviruses. Therefore, the authors should consider changing the title to reflect their data accurately.

Answer:

Thank you for your comment – we do agree and we have modified the word in the title– we used “IgG seropositivity” rather than immunity

2. The authors stored the sera at -20C. Please add how long the sera was stored for. How long are IgGs stable for at -20C?

Answer:

First, as appropriate for long-term stability (Hendriks et al. DOI: 10.4155/bio.14.96, Ma et al. https://doi.org/10.1016/j.biochi.2020.08.019), we stored samples at -20°C as we collected them between December 2018 and May 2019. Accounting for the collection period, storage, transportation delays due to COVID-19, a maximum of 2 years elapsed between collection and defrosting upon the analysis in the National Reference Centre for arboviral diseases in mainland France in Dec 2020. We added this information in the methods.

3. The references need to be improved on several occasions:

•The authors cite a manuscript in preparation; this is highly unusual, and I strongly suggest removing that citation. The details on the methodology should be included in this manuscript, in the methods or supplemental files, or a published source could be cited.

Answer:

Unfortunately, the publication of the cited source is still pending for lack of relevant reviewers; even after submission to PloS One, it had to be resubmitted elsewhere. To address this comment of Reviewer #3, we included a reference to the detailed methods as we did in a recently published study using the same samples (Bastard et al. doi: 10.1038/s43856-022-00230-4): Ruello, M. & Richard, J. Enquête de santé à Mayotte 2019—Unono Wa Maore. Méthode (Santé publique France, 2022, : https://www.santepubliquefrance.fr/docs/enquete-de-sante-a-mayotte-2019-unono-wa-maore.-methode ).

•References 15-17 look like they need more information, is that citing a book chapter?

Answer:

These are series that are published by Sante Publique France or regional public health agencies. In these cases, they are surveillance reports done by the agency in the Indian Ocean. We have amended the references and attached the reports.

•While citing reputable websites is acceptable, the authors cite many websites. I would suggest reducing the number of website citations wherever possible and adding the date accessed for any that will be kept. Links to websites tend to break with time and no longer be accessible.

Answer:

We have eliminated one web sites but left the others, which we thought were necessary. We added the access date and for a few we also included the pdf of the document.

•Reference 31 is all in caps, while the others are not.

Answer:

We have amended it.

4. More details are needed regarding the ELISA. Please clarify if the ELISA is for total IgG, IgG-1 or other. Please specify how the ELISA threshold was chosen. If it was recommended by a manufacturer or based on previous experience or statistical method.

Answer:

For this serosurvey, we performed a homemade indirect ELISA, certified according to ISO 1589 standards. This assay using whole inactivated virus detected total IgG. More details and the following reference were added accordingly in manuscript (Denis J, et al. High specificity and sensitivity of Zika EDIII-based ELISA diagnosis highlighted by a large human reference panel. PLoS Negl. Trop. Dis. 2019;13:e0007747. doi: 10.1371/journal.pntd.0007747.)

5. Since the authors are working with human blood from a region that may have CHIKV present, the authors should specify how the samples were handled, ie/ level of containment, any heat inactivation done, if anything was done in case blood was collected from a patient with CHIKV symptoms. Although the samples were not known to contain CHIKV, this virus is typically BSL-3 in Europe and the US.

Answer:

Samples were opened and handled under a microbiological safety cabinet. Although participants were asymptomatic and their blood samples did not qualify as diagnostic specimens, they underwent a heat-inactivation step (56°C for 30 minutes).

Minor comments:

1. Would it be possible for the authors to use a known sample as standard curve to quantify the IgGs?

Answer:

Unfortunately, this does not apply because our ELISA method was qualitative.

2. The "attitudes to mosquito-borne disease prevention" data is not showing on table 2, I think it was cut off.

Answer:

There was actually one extra variable that was not supposed to be there – we have removed it. Thank you.

Attachment

Submitted filename: Reply to Reviewers_PLOSOne_31March2023.docx

Decision Letter 1

Joel Mossong

11 Apr 2023

PONE-D-22-35076R1Long lasting anti-IgG chikungunya seropositivity in the Mayotte population will not be enough to prevent future outbreaks: A seroprevalence study, 2019.PLOS ONE

Dear Dr. Ortu,

Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process.

==============================

ACADEMIC EDITOR:Please address the remaining queries and suggestions by one of the reviewers. 

==============================

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PLOS ONE

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Please review your reference list to ensure that it is complete and correct. If you have cited papers that have been retracted, please include the rationale for doing so in the manuscript text, or remove these references and replace them with relevant current references. Any changes to the reference list should be mentioned in the rebuttal letter that accompanies your revised manuscript. If you need to cite a retracted article, indicate the article’s retracted status in the References list and also include a citation and full reference for the retraction notice.

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Reviewer's Responses to Questions

Comments to the Author

1. If the authors have adequately addressed your comments raised in a previous round of review and you feel that this manuscript is now acceptable for publication, you may indicate that here to bypass the “Comments to the Author” section, enter your conflict of interest statement in the “Confidential to Editor” section, and submit your "Accept" recommendation.

Reviewer #1: (No Response)

Reviewer #2: All comments have been addressed

**********

2. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #1: Partly

Reviewer #2: Yes

**********

3. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: Yes

Reviewer #2: Yes

**********

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The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: Yes

Reviewer #2: Yes

**********

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PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #1: Yes

Reviewer #2: Yes

**********

6. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: "The authors acknowledge your general feedback – thank you. We mentioned the evidence of not circulation of CHIKV in Mayotte in the discussion - please see lines 288-292."

No report does not mean no circulation of the virus. Please add the point I have made in my previous comment for the discussion. The positive proportion remains the same compared to the previous study in 2006 means that there are no or minimal transmission of the virus since then. If there was an outbreak recently, the positive proportion must be way higher than the previous one. This is a stronger argument for the no circulation in the region point and should be added to the discussion.

"By using Spearman test we did consider multicollinearity rather than collinearity, as we checked the correlation byrunning the Spearman test with all the variables at once (although the output shows all possible combinations of correlation among each 2 variables). We did not opt for using the VIF command"

No pair-wise co-linearity does not exclude the possibility of multicolinearity.

I do not get the difficulty to calculate VIF since the authors only need to calculate it based on the variables

"Based on some reading on Spearman test, the use of it, and the interpretation of thresholds - see for instance these:

https://geographyfieldwork.com/SpearmansRankCalculator.html,

https://statistics.laerd.com/stata-tutorials/spearmans-correlation-using-stata.php

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3576830/ we opted for choosing 0.5 as the threshold. The range between 0.5 and 0.7 is generally associated to moderate correlation. As we did not want to have correlation as a factor that would somehow impact the analysis, 0.5 seemed to be an appropriate and conservative threshold to use (alongside a significant P value)."

Please add these reference to the manuscript to explain why you choose the number.

"We could not conduct an automated stepwise logistic regression using the stepwise command in STATA as the svy prefix command is not compatible with it. However, we did stepwise regression “manually”, specifically we used a backward stepwise process with the commands svy,:poisson and test."

I am not familiar with STATA syntax so I cannot get what the authors trying to say here. Please use statistical terms to explain your reasoning.

Reviewer #2: Thank you for addressing the comments submitted by the reviewers. All of my comments and questions are now resolved.

**********

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Reviewer #1: No

Reviewer #2: No

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PLoS One. 2023 May 18;18(5):e0285879. doi: 10.1371/journal.pone.0285879.r004

Author response to Decision Letter 1


27 Apr 2023

Reviewer #1

"The authors acknowledge your general feedback – thank you. We mentioned the evidence of not circulation of CHIKV in Mayotte in the discussion - please see lines 288-292."

No report does not mean no circulation of the virus. Please add the point I have made in my previous comment for the discussion. The positive proportion remains the same compared to the previous study in 2006 means that there are no or minimal transmission of the virus since then. If there was an outbreak recently, the positive proportion must be way higher than the previous one. This is a stronger argument for the no circulation in the region point and should be added to the discussion.

Answer:

We entirely concur. Therefore, we made our agreement clearer by adding the following (lines 287-288): “This estimate is very similar to the seroprevalence of anti-CHIKV IgG as found in 2006 just after the outbreak in Mayotte, suggesting that there was no or minimal transmission of the virus in the interim.”

"By using Spearman test we did consider multicollinearity rather than collinearity, as we checked the correlation by running the Spearman test with all the variables at once (although the output shows all possible combinations of correlation among each 2 variables). We did not opt for using the VIF command"

No pair-wise co-linearity does not exclude the possibility of multicollinearity.

I do not get the difficulty to calculate VIF since the authors only need to calculate it based on the variables

Answer:

As the reviewer requested, we calculated the VIF with all the variables that we wanted to use for the multivariable analysis (see page 3 of S6), and we obtained an average VIF of 1.62, with a variable showing a maximum of 3.2 VIF. Based on this average VIF value we would have used all the variables for the models, but we felt that this value was not enough to highlight the collinearity issues that we felt very likely present among some of these variables. Therefore, we run the Spearman test, that actually showed us which couples of variables were showing collinearity. Based on this test, we then decided to combine those variables that appear to be collinear, and/or eliminate one of the two that showed collinearity. The final list of variables that we then used for the running our models is the one showed in page 4 of the S6 information sheet. We run again the VIF test with these and the final average value was 1.52, a bit lower than before as we did address the suspected collinearity issues.

We hope that this explanation clarifies why eventually we decided initially, to report the Spearman test on our manuscript rather than the VIF, which we felt, did not provide a satisfactory answer to whether there was collinearity and for which variables. We have added now a few sentences to reflect the request of the reviewer and this issue. We hope that this amendment is acceptable for the reviewer (see lines : 188-190).

"Based on some reading on Spearman test, the use of it, and the interpretation of thresholds - see for instance these:

https://geographyfieldwork.com/SpearmansRankCalculator.html,

https://statistics.laerd.com/stata-tutorials/spearmans-correlation-using-stata.php

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3576830/ we opted for choosing 0.5 as the threshold. The range between 0.5 and 0.7 is generally associated to moderate correlation. As we did not want to have correlation as a factor that would somehow impact the analysis, 0.5 seemed to be an appropriate and conservative threshold to use (alongside a significant P value)."

Please add these references to the manuscript to explain why you choose the number.

Answer:

As initially we understood from the reviewers that it was preferred not to use many web sites among the references, we opted to add only the manuscript among the above references (see No: 26)

"We could not conduct an automated stepwise logistic regression using the stepwise command in STATA as the svy prefix command is not compatible with it. However, we did stepwise regression “manually”, specifically we used a backward stepwise process with the commands svy,:poisson and test."

I am not familiar with STATA syntax so I cannot get what the authors trying to say here. Please use statistical terms to explain your reasoning.

Answer:

By "manual stepwise regression ", we meant that we followed the alternate procedure presented on the Stata website: William Sribney. 2022. Is there a way in Stata to do stepwise regression with svy: logit or any of the svy commands? Stata FAQ. StataCorp Available at: https://www.stata.com/support/faqs/statistics/stepwise-regression-with-svy-commands/ Last Update Date: 16Nov2022. This website is referenced (see ref number: 27)

In brief, we arranged the selected covariates for each model into groupings ordered by decreasing presumed importance. Then we ran each full model before testing the last group of covariates. If not significant, the entire group was discarded. Otherwise, we kept the whole group. Then we repeated the procedure. Hopefully, adding it to our manuscript reference list will not pose a problem since we understood that the reviewers preferred that there were as few references to online resources as possible.

Attachment

Submitted filename: Rebuttal letter _final_25April2023.docx

Decision Letter 2

Joel Mossong

4 May 2023

Long lasting anti-IgG chikungunya seropositivity in the Mayotte population will not be enough to prevent future outbreaks: A seroprevalence study, 2019.

PONE-D-22-35076R2

Dear Dr. Ortu,

We’re pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it meets all outstanding technical requirements.

Within one week, you’ll receive an e-mail detailing the required amendments. When these have been addressed, you’ll receive a formal acceptance letter and your manuscript will be scheduled for publication.

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Kind regards,

Joel Mossong, PhD

Academic Editor

PLOS ONE

Additional Editor Comments (optional):

Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. If the authors have adequately addressed your comments raised in a previous round of review and you feel that this manuscript is now acceptable for publication, you may indicate that here to bypass the “Comments to the Author” section, enter your conflict of interest statement in the “Confidential to Editor” section, and submit your "Accept" recommendation.

Reviewer #1: All comments have been addressed

**********

2. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #1: Yes

**********

3. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: Yes

**********

4. Have the authors made all data underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: Yes

**********

5. Is the manuscript presented in an intelligible fashion and written in standard English?

PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #1: Yes

**********

6. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: (No Response)

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Reviewer #1: No

**********

Acceptance letter

Joel Mossong

9 May 2023

PONE-D-22-35076R2

Long lasting anti-IgG chikungunya seropositivity in the Mayotte population will not be enough to prevent future outbreaks: A seroprevalence study, 2019.

Dear Dr. Ortu:

I'm pleased to inform you that your manuscript has been deemed suitable for publication in PLOS ONE. Congratulations! Your manuscript is now with our production department.

If your institution or institutions have a press office, please let them know about your upcoming paper now to help maximize its impact. If they'll be preparing press materials, please inform our press team within the next 48 hours. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information please contact onepress@plos.org.

If we can help with anything else, please email us at plosone@plos.org.

Thank you for submitting your work to PLOS ONE and supporting open access.

Kind regards,

PLOS ONE Editorial Office Staff

on behalf of

Dr. Joel Mossong

Academic Editor

PLOS ONE

Associated Data

    This section collects any data citations, data availability statements, or supplementary materials included in this article.

    Supplementary Materials

    S1 File. Map of Mayotte with communes and corresponding sectors in the table on the right.

    (DOCX)

    S2 File. Weighed chikungunya seroprevalence and weighed prevalence ratios for all covariates (N = 1438, it may slightly differ for some of the covariates due to missing values).

    (DOCX)

    S3 File. Dataset CHIK analysis_PLOSOne dataset_n2853.

    (DTA)

    S4 File. Dataset CHIK analysis_PLOSOne dataset_n2778.

    (DTA)

    S5 File. Dataset CHIK analysis_PLOSOne dataset_n1438.

    (DTA)

    S6 File. Information on datasets, weights and variables used in the analysis.

    (DOCX)

    Attachment

    Submitted filename: Reply to Reviewers_PLOSOne_31March2023.docx

    Attachment

    Submitted filename: Rebuttal letter _final_25April2023.docx

    Data Availability Statement

    All relevant data are within the paper and its Supporting Information files.


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