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PLOS Neglected Tropical Diseases logoLink to PLOS Neglected Tropical Diseases
. 2025 Nov 6;19(11):e0013644. doi: 10.1371/journal.pntd.0013644

Prevalence of Shigellosis among household contacts of index cases in the EFGH catchment area, Dhaka, Bangladesh

Md Taufiqul Islam 1,☯, Farhana Khanam 1,☯, Md Nazmul Hasan Rajib 1, Md Ismail Hossen 1, Syed Qudrat-E-Khuda 1, Mahzabeen Ireen 1, Md Golam Firoj 1, Faisal Ahmmed 1, Prasanta Kumar Biswas 1, Amirul Islam Bhuiyan 1, S M Azadul Alam Raz 1, Md Parvej Mosharraf 1, Md Taufiqur Rahman Bhuiyan 1, Firdausi Qadri 1,*
Editor: Nicholas J Mantis2
PMCID: PMC12614799  PMID: 41196885

Abstract

Background

This study aimed to estimate the prevalence of shigellosis among household contacts (HHCs) using data from the ‘Enterics for Global Health (EFGH)’ study, conducted at seven Asian and African countries.

Methods

In Bangladesh, the EFGH study was conducted in Maniknagar, Dhaka, to determine the burden of shigellosis among children aged 6–35 months. HHCs of Shigella-positive patients (index cases) were enrolled in this study. Stool specimens form contacts were collected within 7 days of enrollment of index cases and culture and qPCR were performed. Sociodemographic and behavioral information were obtained to identify risk factors.

Results

A total of 400 HHCs of 118 index cases were enrolled, of which 36 (9%) were positive for Shigella spp. by culture, while qPCR revealed 21% (42/200) of contacts had Shigella infections. Individuals who failed to reheat meals before consumption had a two-fold higher risk of shigellosis compared to those who reheated meals, although other sociodemographic and behavioral factors did not show any significant association.

Conclusions

The study revealed a high burden of asymptomatic Shigella-infected individuals, emphasizing the need for systematic surveillance to assess the burden and develop effective preventive strategies to prevent the spread of the disease.

Author summary

Asymptomatic individuals can play a key role in transmitting Shigella within communities in endemic settings. We assessed the prevalence and risk factors of Shigella infection among household contacts of culture-confirmed index cases in an urban slum of Dhaka, Bangladesh. Our study revealed a high proportion of household contacts were infected, with 97% of them excreting the organism asymptomatically. Index cases were children aged 6–35 months, and household members represented a range of age groups, with 75% being adults. Failure to reheat meals before consumption and the use of non-flush toilets were significant risk factors. These findings highlight the urgent need for interventions that extend beyond symptomatic case management, emphasizing safe food handling practices and improved sanitation to interrupt transmission chains effectively.

Background

Shigellosis is a severe intestinal infection caused by the gram-negative bacterium Shigella spp., primarily transmitted through the fecal-oral route [1]. Shigellosis is one of the leading causes of diarrheal illness worldwide, contributing to substantial morbidity and mortality, particularly among children under five years of age [2]. This disease represents a major public health challenge, particularly in low- and middle-income countries like Bangladesh, where inadequate sanitation and limited access to clean water exacerbate its spread. In a hospital-based study, the incidence of shigellosis among children under five in a densely populated urban area of Dhaka, Bangladesh was 4.6 cases per 100 person-years [3,4]. A previous study in rural Bangladesh highlighted the increased risk of Shigella transmission among infected individuals’ household contacts, underscoring this disease’s widespread nature [2]. Another retrospective cohort study conducted in Amsterdam between 2002 and 2009 showed that the laboratory-confirmed Shigella infection in high-risk household contacts of primary cases was 7.4% [5].

The Enterics for Global Health (EFGH) Shigella burden study, conducted in urban Dhaka from 2022 to 2024, aimed to estimate the incidence rates of Shigella-attributed diarrhea in children aged 6–35 months, documenting health outcomes and informing public health strategies and interventions. A sub-study was planned using the EFGH platform. The objective of the study was to estimate the proportion of Shigella infection among household contacts and also to identify the risk factors associated with the infection among household contacts (HHCs).

Methods

Ethical clearance

This study (PR 23034) was approved by the Research Review Committee (RRC) and Ethical Review Committee (ERC) of International Centre for Diarrhoeal Disease Research, Bangladesh, icddr,b. We obtained informed written consent or assent from all eligible study participants or their legal guardian prior to the enrollment.

Enrollment of study participants

The EFGH study was conducted in wards 6, 7, 48, 49, 50, 71, and 72 of Dhaka South City Corporation, Bangladesh, from the EFGH Shigella surveillance sites. Patients aged 6–35 months presented with diarrhea to the icddr,b Dhaka hospital, the Mugda Medical College Hospital, and the EFGH field office in the Maniknagar area were enrolled in the EFGH study. A rectal swab was collected from them for microbiological culturing. A culture-confirmed Shigella patient was considered as an index case and their household contacts were selected for this sub-study. The HHCs were individuals of all ages who permanently resided at the time of the study in the same household as the index cases, shared food from the same kitchen as the index cases, and were present for at least 7 of the last 14 days.

All HHCs of Shigella-positive index cases were approached and enrolled in this study from April 2023 to June 2024 after completing the proper consenting procedure. Enrollment occurred within 7 days of receiving the culture reports of EFGH index cases. Study staff visited the households on the 6th day and provided stool containers to the parents or guardians of index cases to collect stool specimens from all household members on day 7. The participants were reminded by phone calls or household visits to ensure the collection of day 7 stool specimens. In addition, if any household members developed acute watery diarrhea (≥3 abnormally loose or watery stools in 24 hours) within seven days, a stool specimen was obtained from them. No further stool collection was carried out.

Specimen collection, preparation and transportation

Stool specimens were collected in stool containers from the contacts of the index cases. A stool swab was prepared from a stool specimen collected from HHCs and placed dry in a transport tube (2 ml screw top vial compatible with a bead beater) for TaqMan Array Card (TAC) testing. All specimens were then immediately transported to the icddr,b laboratory, maintaining a temperature of 2–8˚C [6].

Laboratory methods

Stool specimens were streaked directly onto MacConkey agar and Xylose Lysine Deoxycholate agar media and incubated at 37°C for 18–24 hours. Morphologically suspicious Shigella colonies were inoculated onto Triple Sugar Iron (TSI), Motility Indole Urea (MIU) and Tryptic Soy Agar (TSA). Shigella isolates were then confirmed by serotyping test using commercially available antisera (Denka Seiken Co., Ltd., Tokyo, Japan) for agglutination. The antibiotic susceptibility testing (AST) of confirmed Shigella isolates was done by the Kirby-Bauer disk diffusion method and zone diameters were recorded and classified as susceptible, resistant, or intermediate based on the Clinical and Laboratory Standards Institute guideline [7]. AST was performed for various antibiotics, including ampicillin, azithromycin, ceftriaxone, ciprofloxacin, nalidixic acid, pivmecillinam, and trimethoprim/sulfamethoxazole.

The QIAamp Fast DNA Stool Mini kit (Qiagen, Valencia, CA) was used to extract nucleic acid from stool specimens for TAC testing. The quantitative analysis was performed to determine the likely attribution of Shigella and other particular etiologies of diarrhea using the QuantStudio 7 Flex. A customized TaqMan Array Card (TAC) was used, which had a total of 82 targets. Among them, 37 targets were for bacteria, 8 for viruses, 1 for fungi, 5 for protozoa, and 4 targets for extraction and plate control. After completion of the plate run, the dataset was uploaded to the Multi-Schema Information Capture database platform for analysis. Shigella was targeted using ipaH/Shigella EIEC, and the cut-off for positivity was set at a quantification cycle (Cq) ≤29.8. When the Cq value exceeded 29.8, Shigella serotypes were not pursued further. For other pathogens, the Cq cut-offs were the same as that was used in previous studies were applied [8].

Data capturing and management

Comprehensive documentation of each participant was maintained throughout the study. Data on demographic, socioeconomic, clinical characteristics, and water sanitation and hygiene (WASH) behaviour and practices were collected using structured case report forms and entered into the electronic data capturing system and then transferred to the SQL-Server of icddr,b. Errors in the database were detected and corrected following a standard operating procedure of maintaining an audit trail.

Statistical analysis

The prevalence of Shigella infection among household contacts was determined using numbers and percentages and the prevalence of households with Shigella infection was calculated based on various household-level demographic, socio-economic, and WASH characteristics. The numerator was the number of households with Shigella infection, and the denominator was the total number of households at risk. A 95% confidence interval (CI) for the prevalence was calculated using binomial proportions of the Wilson method. Bar diagrams and pie charts were used to present the summarized TAC results for various organisms, serotypes of Shigella and the antimicrobial susceptibility patterns. Association between Shigella infection among HHCs and individual- and household-level demographic, socio-economic, and WASH characteristics were assessed using the Chi-square and the Fisher’s exact test.

Logistic regression was carried out to measure the relationship between predisposing factors in household contacts and Shigella infection. Exponential coefficients of the binomial Logistic regression models with log link function have been used and presented as risk ratios (RR) with a corresponding 95% CI. Variables with a significance level of ≤0.2 in univariate analysis were included in the multiple regression model as independent variables. All tests were two-tailed, where p-value <0.05 was considered statistically significant. The statistical analysis was conducted using R (version 4.3.3). For plots and figures, “ggplot2” package and Microsoft Excel were used.

Results

Assembly of the participants

We identified 118 culture-confirmed Shigella index cases in the EFGH study. A total of 443 household members of the EFGH index cases were approached for enrollment in this study. Of these, 24 refused to participate, and 19 were not enrolled in this study as they developed acute watery diarrhea (AWD) within one day of the index case’s enrollment (Fig 1). Among 400 household members enrolled in the study, 205 (51%) were female, and 299 (75%) were adults.

Fig 1. CONSORT diagram of household contact study.

Fig 1

Shigella infection among household contacts by using microbiological culture of stool specimens

Microbiological culture of stool samples collected from 400 enrolled household contacts revealed that 36 individuals (9%) were Shigella-positive. Of these, only one had diarrhea before enrollment, while the remaining 35 cases were asymptomatic (Fig 1). The Shigella isolates consisted of four species: Shigella flexneri (n = 15), Shigella sonnei (n = 10), Shigella boydii (n = 7), and Shigella dysenteriae (n = 4). Among the S. flexneri isolates, subtyping revealed a range of serotypes, including S. flexneri 2a (n = 4), S. flexneri 4a (n = 1), S. flexneri 1a (n = 1), and 9 non-typable S. flexneri isolates (Table 1). Not all household members with stool cultures positive for Shigella had the same serogroup as the index case: 11 shared the same serogroups, while the others had different ones. 29 households out of 118 had an infection. 2 members from 5 households and 3 members from one HH were infected.

Table 1. Proportion of Shigella species detected by culture and pathogens detected by TAC among HH contacts of index cases.

Culture results No. of positive cases Percentage
Shigella flexneri 15 3.8
 Shigella flexneri 2a 4 26.7
 Shigella flexneri 4a 1 6.7
 Shigella flexneri 1a 1 6.7
 Shigella flexneri non-typable 9 60
Shigella sonnei 10 2.5
Shigella boydii 7 1.8
Shigella dysenteriae 4 1
TaqMan Array Card (TAC) results No. of positive cases Percentage
EAEC 139 69.5
EPEC 125 62.5
ETEC_LT 100 50
Giardia 96 48
ETEC_ST 93 46.5
Shigella_EIEC 78 39
C infans 51 25.5
C jejuni coli 23 11.5
Adenovirus 17 8.5
Plesiomons 16 8
E.bieneusi 14 7
Sapovirus 124 12 6
STEC 9 4.5
Cryptosporidium 8 4
Aeromonas 7 3.5
Vibrio choler ae 6 3
C_Upsalensis 6 3
H pylori 4 2
Norovirus G-II 4 2
Rotavirus 3 1.5
Norovirus G-I 2 1
Salmonella 1 0.5
SARS-CoV-2 1 0.5
Astrovirus 1 0.5
Cyclospora 1 0.5

Risk factors of Shigella infection among HH contacts

For individual-level characteristics, there were no significant differences in Shigella infection by age group, gender, or religion. Education level showed a trend, though the difference was not statistically significant (p = 0.063): there were more Shigella-positive cases among individuals with secondary education (19%) compared to those with higher education (6%). Occupational status, food consumption outside the home, and use of soap for personal hygiene did not significantly differ between Shigella-positive cases and Shigella-negative individuals. Notably, individuals who did not heat their meals before consumption had a significantly higher proportion of Shigella positivity (19%) compared to those who did (8%) (p = 0.041).

At the household level, no significant association between family size and Shigella positivity was observed. Duration of residence in the current household (more than one year or less than one year) did not show significant differences between Shigella-positive and Shigella-negative participants. Additionally, construction materials of the roof, walls, and floors had no significant association with Shigella positivity. Sanitation facilities, including toilet type, toilet sharing, kitchen sharing, and water sources, also showed no significant differences. Water adequacy and treatment practices, along with waste disposal methods, did not reveal notable associations with Shigella infection among household members (Table 2).

Table 2. Household and individual level demographic, socio-economic, and WASH characteristics of the Shigella positive and negative HH contacts.

Household level
Factors Labels Shigella positive n = 36 Shigella negative n = 364 p-value
Number of family member <4 20 (12%) 153 (88%) 0.158
≥4 16 (7%) 211 (93%)
Number of persons live per room ≤2 14 (9%) 149 (91%) 0.861
>2 22 (9%) 215 (91%)
Living in this household till ≤1 year 18 (10%) 159 (90%) 0.486
>1 year 18 (8%) 205 (92%)
Construction materials of the roof Brick/Cement 22 (8%) 249 (92%) 0.455
Others 14 (11%) 115 (89%)
Construction materials of the wall Brick/Cement 31 (9%) 323 (91%) 0.588
Others 5 (11%) 41 (89%)
Construction materials of the floor Brick/Cement 36 (9%) 362 (91%) >0.999
Others 0 (0%) 2 (100%)
Toilet type Flush or pour flush 35 (9%) 363 (91%) 0.172
Non-flush 1 (50%) 1 (50%)
Shared toilet Yes 17 (9%) 166 (91%) 0.863
No 19 (9%) 198 (91%)
Shared kitchen Yes 18 (8%) 195 (92%) 0.728
No 18 (10%) 169 (90%)
Source of water Tube-well/Bottled/ ATM booth 9 (8%) 111 (92%) 0.571
Supply/Tanker/Public tap/Rain/River etc. 27 (10%) 253 (90%)
Amount of water adequate Yes 36 (9%) 364 (91%) –
No – –
Treatment of water Boiled/filtered/ chemical 18 (9%) 190 (91%) 0.862
Not treated 18 (9%) 174 (91%)
Water available 24 hours and 7 days Yes 36 (9%) 364 (91%) –
No – –
Distance of water source from toilet/sewerage system in feet Within 3 feet 19 (8%) 231 (92%) 0.212
More than 3 feet 17 (11%) 133 (89%)
Place of waste disposal Fixed 36 (9%) 359 (91%) >0.999
Indiscriminate 0 (0%) 5 (100%)
Individual level
Age <5 years 2 (9%) 21 (91%) 0.856
5 to 17 years 8 (10%) 70 (90%)
≥18 years 26 (9%) 273 (91%)
Gender Male 17 (9%) 178 (91%) 0.863
Female 19 (9%) 186 (91%)
Religion Muslim 36 (9%) 364 (91%) –
Educational status Higher educated 4 (6%) 62 (94%) 0.063
Secondary* 5 (19%) 21 (81%)
Primary 7 (5%) 124 (95%)
Child/Illiterate 20 (11%) 157 (89%)
Occupational status Work Outside 14 (8%) 161 (92%) 0.834
House maker 12 (10%) 111 (90%)
Unemployed/ Child/Student 10 (10%) 92 (90%)
Taken food outside home in last 7 days Yes 20 (9%) 207 (91%) >0.999
No 16 (9%) 157 (91%)
Heat meal before intake Yes 28 (8%) 329 (92%) 0.041
No 8 (19%) 35 (81%)
Use soap after defecation Yes 35 (9%) 356 (91%) 0.576
No 1 (11%) 8 (89%)
Use soap before taking food Yes 36 (9%) 359 (91%) >0.999
No 0 (0%) 5 (100%)

*Secondary education includes schooling for 5 years, classes from VI to X.

The multivariate analysis revealed that secondary education posed a higher risk for Shigella infection among household members, with an adjusted relative risk (RR) of 3.19 (95% CI: 0.95, 10.68; p = 0.060), indicating a threefold higher risk compared to higher educational levels. Additionally, failing to reheat meals before consumption showed a significant risk (RR = 2.27, 95% CI: 1.07, 4.83; p = 0.034) for being infected. Also, the type of toilet used indicated that non-flush users had a higher risk for Shigella infection, with an adjusted relative risk of 6.16 (95% CI: 1.31, 29.03; p = 0.021). Additionally, the distance of >3 feet between the water source and toilet/sewerage system was associated with a risk of infection (RR = 2.35, 95% CI: 0.70, 2.59), but this association was not statistically significant. However, the analysis did not show any significant effect of using soap before eating in reducing the risk of infection (Table 3).

Table 3. Predisposing factors in household contacts for developing Shigella infection.

Factors Labels Crude RR (95%CI) p value Adj. RR (95%CI) p value
Age ≥18 years Ref. – – –
5 to 17 years 1.18 (0.56, 2.5) 0.667 – –
<5 years 1 (0.25, 3.95) >0.999 – –
Gender Male Ref. – – –
Female 1.06 (0.57, 1.98) 0.848 – –
Educational status Higher educated Ref. – Ref. –
Secondary 3.17 (0.92, 10.9) 0.067 3.19 (0.95, 10.68) 0.060
Primary 0.88 (0.27, 2.9) 0.836 0.69 (0.21, 2.28) 0.542
Child/Illiterate 1.86 (0.66, 5.25) 0.238 1.32 (0.46, 3.78) 0.608
Occupational status Work Outside Ref. – – –
House maker 1.22 (0.58, 2.55) 0.597 – –
Unemployed/ Child/Student 1.23 (0.57, 2.66) 0.607 – –
Taken food outside home in last 7 days No Ref. – – –
Yes 0.95 (0.51, 1.78) 0.879 – –
Heat meal before intake Yes Ref. – Ref. –
No 2.37 (1.16, 4.87) 0.019 2.27 (1.07, 4.83) 0.034
Use soap after defecation Yes Ref. – – –
No 1.24 (0.19, 8.09) 0.821 – –
Use soap before taking food Yes Ref. – – –
No 0 (0, Inf) 0.989 – –
Number of family member <4 Ref. – Ref. –
≥4 0.61 (0.33, 1.14) 0.122 0.62 (0.33, 1.17) 0.142
Number of persons live per room ≤2 Ref. – – –
>2 1.08 (0.57, 2.05) 0.812 – –
Living in this household till ≤1 year Ref. – – –
>1 year 0.79 (0.43, 1.48) 0.467 – –
Construction materials of the roof Brick/Cement Ref. – – –
Others 1.34 (0.71, 2.53) 0.371 – –
Construction materials of the wall Brick/Cement Ref. – – –
Others 1.24 (0.51, 3.03) 0.635 – –
Construction materials of the floor Brick/Cement Ref. – – –
Others 0 (0, Inf) 0.989 – –
Toilet type Flush or pour flush Ref. – Ref. –
Non-flush 5.69 (1.37, 23.56) 0.017 6.16 (1.31, 29.03) 0.021
Shared toilet No Ref. – – –
Yes 1.06 (0.57, 1.98) 0.853 – –
Shared kitchen No Ref. – – –
Yes 0.88 (0.47, 1.64) 0.682 – –
Source of water Tube-well/Bottled/ ATM booth Ref. – – –
Supply/Tanker/Public tap/Rain/River etc. 1.29 (0.62, 2.65) 0.496 – –
Treatment of water Boiled/filtered/ chemical Ref. – – –
Not treated 1.08 (0.58, 2.02) 0.801 – –
Distance of water source from toilet/sewerage system in feet Within 3 feet Ref. – Ref. –
More than 3 feet 1.49 (0.8, 2.78) 0.208 2.35 (0.70, 2.59) 0.369
Place of waste disposal Fixed Ref. – – –
Indiscriminate 0 (0, Inf) 0.989 – –

Antimicrobial resistance pattern of isolated Shigella

Antibiotic susceptibility profiles revealed distinct resistance patterns among the four Shigella species. All isolates were uniformly susceptible to pivmecillinam. S. flexneri exhibited high resistance to ampicillin (60.0%) and notable resistance to nalidixic acid (46.7%). S. sonnei showed complete resistance to ciprofloxacin and nalidixic acid (100.0%), and high resistance to azithromycin (70.0%). S. boydii was highly susceptible to most antibiotics but showed resistance to nalidixic acid (85.7%). S. dysenteriae was susceptible to all antibiotics except nalidixic acid, in which 50.0% of isolates were resistant. (Fig 2).

Fig 2. Antimicrobial susceptibility pattern of isolated Shigella strains among the household contacts.

Fig 2

Pathogens identified from TaqMan Array Card (TAC) assay

Among the 400 participants, the first 200 stool specimens collected were tested using the TAC assay. Shigella was identified in 42 (21%) of the samples, with the majority (69%, 29 samples) classified as Shigella flexneri, while 13 (31%) samples were identified as Shigella sonnei. Among the Shigella flexneri isolates, the most prevalent serotypes were Shigella flexneri 1a, 2a, 2b, 4a, 5a, 6, 7a and X in addition to Shigella, the analysis detected 26 other organisms, including enterotoxigenic Escherichia coli, adenovirus, rotavirus and cryptosporidium spp. A total of 5 (38.4%) samples had co-infection with Shigella (Table 1).

Discussion

There is a substantial burden of (9%) Shigella infection among asymptomatic household members of culture-confirmed Shigella index cases, identified in the EFGH Bangladesh site conducted between June 2022 and October 2024 [7]. The high prevalence of asymptomatic cases underscores the potential for asymptomatic carriage, which may play a crucial role in transmission of Shigella within families and communities, contributing to a high burden of Shigella infection in this setting. Our findings also indicate that households with identified index cases are at particularly high risk of shigellosis. However, the differences in Shigella serogroups observed between some index cases and their household members suggest that these infections were not directly related to the index cases, or that secondary transmission did not occur.

Similarly, studies conducted in Amsterdam from 1992 to 1998 reported 8% rate of Shigella infection among household members and asymptomatic individuals [9]. Between April and August 2008, an outbreak of Shigella sonnei infection was documented in the Orthodox Jewish community of Antwerp, Belgium, with a household attack rate of 8.5% [10]. A study conducted in Bangladesh on the intra-family transmission of enteric diseases reported that the intra-familial Shigella transmission rate was 8% [11]. Furthermore, a study in rural Bangladesh indicated that household members were likely to transmit Shigella infection within the week following the initial diagnosis of Shigella-infected patients [2].

The antibiotic susceptibility testing (AST) of Shigella isolates revealed higher sensitivity to pivmecillinam, ceftriaxone, and trimethoprim/sulfamethoxazole, with a significant proportion of isolates demonstrating resistance to nalidixic acid and ciprofloxacin. These findings will be helpful in selecting appropriate antibiotics for treating Shigella-infected patients, particularly in settings where AST facilities are inadequate. A study conducted in Bangladesh showed similar patterns, with 93% sensitivity to pivmicillinam, 96.8% sensitivity to ceftriaxone, and 93% resistance to nalidixic acid. Additionally, a meta-analysis based on studies from Bangladesh revealed resistance patterns against Shigella, with 38.8% resistance to azithromycin, 34.5% resistance to ampicillin, 31.1% resistance to ciprofloxacin, and 10.8% resistance to ceftriaxone, which was similar to our findings [12,13].

Our analysis revealed several significant risk factors for Shigella infection among HHCs. Education up to secondary education level only was associated with a higher risk, with an adjusted relative risk (RR) of 3.19, indicating a threefold higher risk compared to individuals with higher educational levels. This finding aligns with a previous study identifying lower educational levels as a risk factor for various infectious diseases [14]. Additionally, failing to reheat meals before consumption was associated with a significant risk (RR = 2.27), consistent with studies highlighting the importance of proper food handling and reheating practices in preventing foodborne illnesses [15]. The type of toilet used also appeared as a risk factor, with non-flush toilets posing a significant risk. This finding is supported by a study showing that inadequate sanitation facilities contribute to the spread of enteric diseases [16].

Our study had several limitations. Firstly, it was conducted in the densely populated urban area of Dhaka city, which may not accurately reflect transmission dynamics in rural areas of Bangladesh or less inhabited regions in LMICs globally. Secondly, the use of self-reported data for certain variables in the risk factor analysis, such as hygiene practices, could introduce reporting bias. Lastly, although the sample size was calculated based on the primary objective (the proportion of infection among household members), this may have limited power to identify additional risk factors. Despite these limitations, our findings have great implications, highlighting a high proportion of asymptomatic Shigella infections. This contributes to a sustained high burden of infection, with morbidity and subsequent consequences such as growth failure, hospitalization, and death among young children.

In conclusion, the evidence generated from this study shows the spread of infection within households and may be leading to carriage. This information will support policymakers in the development of comprehensive and targeted preventive strategies, ultimately leading to better health outcomes and a reduced disease burden by effectively interrupting the transmission chains in the community.

Acknowledgments

The icddr,b is grateful to the governments of Bangladesh and Canada for providing core/unrestricted support. We would like to thank Patricia B. Pavlinac, Kirk Tickell, Stephanie Edlund Cho, Sonia Rao, Hannah Atlas from the University of Washington and Beth Tippett Barr from Nyanja Health Research Institute, Eric Houpt and Queen Saidi from the University of Virginia.

Data Availability

A de-identified analytical data set will be made available upon requests directed to the Institutional Review Board (IRB) coordinator of the icddr,b at info@icddrb.org. Only after approval of a proposal data can be shared through a secure online platform. Approval of the proposal will be subject to scientific review by the institutional review board at icddr,b. Sharing of data will also be subject to the published data access rules of the icddr,b. The requestor will need to sign a standard data access agreement required by the icddr,b.

Funding Statement

This study was supported by the Bill & Melinda Gates Foundation [INV-045988]. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. No authors received any salary from the funder.

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PLoS Negl Trop Dis. doi: 10.1371/journal.pntd.0013644.r002

Decision Letter 0

Stuart Blacksell

29 Jul 2025

Prevalence of Shigellosis among household contacts of index cases in the EFGH catchment area, Dhaka, Bangladesh

PLOS Neglected Tropical Diseases

Dear Dr. Qadri,

Thank you for submitting your manuscript to PLOS Neglected Tropical Diseases. After careful consideration, we feel that it has merit but does not fully meet PLOS Neglected Tropical Diseases'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.

Please submit your revised manuscript within 60 days Sep 27 2025 11:59PM. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at plosntds@plos.org. When you're ready to submit your revision, log on to https://www.editorialmanager.com/pntd/ and select the 'Submissions Needing Revision' folder to locate your manuscript file.

Please include the following items when submitting your revised manuscript:

* A rebuttal letter that responds to each point raised by the editor and reviewer(s). You should upload this letter as a separate file labeled 'Response to Reviewers'. This file does not need to include responses to any formatting updates and technical items listed in the 'Journal Requirements' section below.

* A marked-up copy of your manuscript that highlights changes made to the original version. You should upload this as a separate file labeled 'Revised Manuscript with Track Changes'.

* An unmarked version of your revised paper without tracked changes. You should upload this as a separate file labeled 'Manuscript'.

If you would like to make changes to your financial disclosure, competing interests statement, or data availability statement, please make these updates within the submission form at the time of resubmission. Guidelines for resubmitting your figure files are available below the reviewer comments at the end of this letter.

We look forward to receiving your revised manuscript.

Kind regards,

Nicholas J Mantis

Academic Editor

PLOS Neglected Tropical Diseases

Stuart Blacksell

Section Editor

PLOS Neglected Tropical Diseases

Shaden Kamhawi

co-Editor-in-Chief

PLOS Neglected Tropical Diseases

orcid.org/0000-0003-4304-636XX

Paul Brindley

co-Editor-in-Chief

PLOS Neglected Tropical Diseases

orcid.org/0000-0003-1765-0002

Additional Editor Comments:

There was a split decision between the two reviewers about the merit of the manuscript. While both Reviewers recognize the significance of the study, Reviewer 1 raised concerns about the analysis conducted to define relationships between the index cases and the household cases. The reviewer suggested you re-analyze and re-write the paper from the data set, but with more consideration of the types of analysis needed. This comment in particular influenced the decision for a major revision.

Journal Requirements:

1) Please ensure that the CRediT author contributions listed for every co-author are completed accurately and in full.

At this stage, the following Authors/Authors require contributions: Md Taufiqul Islam, Farhana Khanam, Md Nazmul Hasan Rajib, Md Ismail Hossen, Syed Qudrat-E-Khuda, Mahzabeen Ireen, Md Golam Firoj, Faisal Ahmmed, Prasanta Kumar Biswas, Amirul Islam Bhuiyan, S.M. Azadul Alam Raz, Md. Parvej Mosharraf, Md Taufiqur Rahman Bhuiyan, and Firdausi Qadri. Please ensure that the full contributions of each author are acknowledged in the "Add/Edit/Remove Authors" section of our submission form.

The list of CRediT author contributions may be found here: https://journals.plos.org/plosntds/s/authorship#loc-author-contributions

2) Please provide an Author Summary. This should appear in your manuscript between the Abstract (if applicable) and the Introduction, and should be 150-200 words long. The aim should be to make your findings accessible to a wide audience that includes both scientists and non-scientists. Sample summaries can be found on our website under Submission Guidelines:

https://journals.plos.org/plosntds/s/submission-guidelines#loc-parts-of-a-submission

3) Tables should not be uploaded as individual files. Please remove these files and include the Tables in your manuscript file as editable, cell-based objects. For more information about how to format tables, see our guidelines:

https://journals.plos.org/plosntds/s/tables 

4) We note that your Data Availability Statement is currently as follows: "All relevant data are within the paper and its supporting information files.". Please confirm at this time whether or not your submission contains all raw data required to replicate the results of your study. Authors must share the “minimal data set” for their submission. PLOS defines the minimal data set to consist of the data required to replicate all study findings reported in the article, as well as related metadata and methods (https://journals.plos.org/plosone/s/data-availability#loc-minimal-data-set-definition).

For example, authors should submit the following data: 

1) The values behind the means, standard deviations and other measures reported;

2) The values used to build graphs;

3) The points extracted from images for analysis..

Authors do not need to submit their entire data set if only a portion of the data was used in the reported study.

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5) Please provide a detailed Financial Disclosure statement. This is published with the article. It must therefore be completed in full sentences and contain the exact wording you wish to be published.

1) Please clarify all sources of financial support for your study. List the grants, grant numbers, and organizations that funded your study, including funding received from your institution. Please note that suppliers of material support, including research materials, should be recognized in the Acknowledgements section rather than in the Financial Disclosure

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3) State what role the funders took in the study. If the funders had no role in your study, please state: "The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript."

4) If any authors received a salary from any of your funders, please state which authors and which funders..

If you did not receive any funding for this study, please simply state: u201cThe authors received no specific funding for this work.u201d

6) Your current Financial Disclosure states, "The author(s) received no specific funding for this work.".

However, your funding information on the submission form indicates  receiving funds.. 

Please indicate by return email the full and correct funding information for your study and confirm the order in which funding contributions should appear. Please be sure to indicate whether the funders played any role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript.

Reviewers' Comments:

Reviewer's Responses to Questions

Key Review Criteria Required for Acceptance?

As you describe the new analyses required for acceptance, please consider the following:

Methods

-Are the objectives of the study clearly articulated with a clear testable hypothesis stated?

-Is the study design appropriate to address the stated objectives?

-Is the population clearly described and appropriate for the hypothesis being tested?

-Is the sample size sufficient to ensure adequate power to address the hypothesis being tested?

-Were correct statistical analysis used to support conclusions?

-Are there concerns about ethical or regulatory requirements being met?

Reviewer #1: The objectives are clear - that is to carry out a family study of shigellosis. However, there is no testable hypothesis. The population could be better described by age, sex, season.

Reviewer #2: The methods are clearly described and are appropriate for addressing the objectives of this study.

**********

Results

-Does the analysis presented match the analysis plan?

-Are the results clearly and completely presented?

-Are the figures (Tables, Images) of sufficient quality for clarity?

Reviewer #1: The analysis presented match the plan, however, the analysis could have gone into more depth and detail.

i felt the figures 2 and 4 are not needed.

Reviewer #2: The results match the analysis plan and are supported by the data presented. The tables and figures are clear and helpfully illustrate the key findings.

**********

Conclusions

-Are the conclusions supported by the data presented?

-Are the limitations of analysis clearly described?

-Do the authors discuss how these data can be helpful to advance our understanding of the topic under study?

-Is public health relevance addressed?

Reviewer #1: The authors found many shigella infections in household members of index patients, but it is difficult to know what this means.

Reviewer #2: The conclusions are clearly presented and are well-supported by the data provided.

**********

Editorial and Data Presentation Modifications?

Use this section for editorial suggestions as well as relatively minor modifications of existing data that would enhance clarity. If the only modifications needed are minor and/or editorial, you may wish to recommend “Minor Revision” or “Accept”.

Reviewer #1: Editorial comments (the numbers refer to the line numbers).

37, Why mention vaccinations in the abstract when there are no licensed vaccines

83. Is there validation that a cold temp is needed. Could it be even harmful to maintain survival of shigellae?

85 stool specimens (plural) were…

92 reference(s) needed for the AST assays

98 is this the first time this customized card was used ? is this the same assay as used in the GEMs studies

114 “All HH stools positive for Shigella were not of the same species or serotype.” This sentence needs to ber explained. Not the same as what?

163 Explain “secondary education.” How many years of schooling is this?

189 I believe the ERC number should be included.

Reviewer #2: Minor comments/typographical:

Ln 37- I would favor replacing "vaccination" with " prevention" as the paper does not deal at all with vaccination and other prevention strategies are alluded to in the results (i.e reheating meals)

Ln 136- fill in the % value

Ln 186- Ethical clearance should go under methods

Fig 1- "CONSORT" should be capitalized

**********

Summary and General Comments

Use this section to provide overall comments, discuss strengths/weaknesses of the study, novelty, significance, general execution and scholarship. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. If requesting major revision, please articulate the new experiments that are needed.

Reviewer #1: This is an ambitious study to study shigella infections in households of index patients with shigellosis. It is an interesting study, but it seems the analysis could have gone into more depth in their analysis. For example:

1. When the study on household infections was being considered, did the authors have a hypothesis about the household infections? For example, did they want to determine if the infections were being spread between family members and if so, how? To answer this question, they would need to track the specific strains between household members. Perhaps alternatively, they just wanted to see if index cases identified households that have a high risk for shigellosis in general because of poor water/sanitation. If there was a hypothesis, what was the conclusion after doing the study?

2. What were the characteristics of the index cases? Did they all have dysentery or did some have watery diarrhea and were classified as an index case only after a positive culture was detected. Did the clinical symptoms change the rate of infections in household members?

3. What were the species and serotypes of the index cases, and how did these match the species / serotype of the infections in the household members. It would seem, there were many different strains of shigella in the household members, but they may not be related to the infection in the index cases.

4. What is the rate of shigella infection in the community? In this slum area of the city, it may be that shigella infections are common in the general population, especially infections that can be detected by PCR. Without a comparison of rates in a control group, it is difficult to know if the households had a higher rate than would be expected.

5. The analysis of households should be by household as well as individual. How many of the households had an infection. What was the distribution of the numbers of infections in the infected households. Did some households have many people infected and others had 0 or 1? It was not clear if the risk factors looked at households or only at individuals with infections.

6. I believe Table 1 can better describe the characteristics of the household members by age and sex.

7. Was there any seasonality among the index cases.

8. I believe the discussion section could be improved by first describing what the authors found in their study. I prefer to understand all the findings from this study first. Then, in later paragraphs compare their findings to other studies.

9. In terms of limitations, I would highlight the lack of control group. Ideally this could have been a case control study in which households of index patients with a non-diarrheal illness could be compared. Obviously, it is too late to do this now, but is there another way to estimate the rates in the community?

10. The antibiotic sensitivity patterns must have differed by species and serotype. By lumping together, I believe we do not have an appreciation of the true antibiotic sensitivity patterns.

Figure 2. I do not think a figure is needed. A table would be better.

Figure 3. Include the number of isolates tested

Figure 4. The figure does not have a label for the y axis. With so few pathogens identified with the Taq card, it would seem this does not deserve a figure. The data could be included in the text.

Reviewer #2: This manuscript describes a study assessing factors associated with Shigella carriage among household contacts of index children aged 6-35 months presenting with Shigella infection in Dhaka, Bangladesh. The cohort was relatively large, but still of limited size to detect differences in some factors. The methods were rigorously defined and support the conclusions drawn, which showed frequent asymptomatic carriage in household contacts and risk factors that included failure to reheat meals. I admit to being a little confused about how toilet type is shown to be significant in even the univariate analysis despite having only 1 infection in a total 2 individuals without flush toilets.

I think this paper does add to the general knowledge of household transmission of shigella and could be useful in developing strategies for screening or prevention.

**********

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If you choose “no”, your identity will remain anonymous but your review may still be made public.

Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy .

Reviewer #1: No

Reviewer #2: No

Figure resubmission:

While revising your submission, please upload your figure files to the Preflight Analysis and Conversion Engine (PACE) digital diagnostic tool, https://pacev2.apexcovantage.com/. PACE helps ensure that figures meet PLOS requirements. To use PACE, you must first register as a user. Registration is free. Then, login and navigate to the UPLOAD tab, where you will find detailed instructions on how to use the tool. If you encounter any issues or have any questions when using PACE, please email PLOS at figures@plos.org. Please note that Supporting Information files do not need this step. If there are other versions of figure files still present in your submission file inventory at resubmission, please replace them with the PACE-processed versions.

Reproducibility:

?>

PLoS Negl Trop Dis. doi: 10.1371/journal.pntd.0013644.r004

Decision Letter 1

Ana LTO Nascimento

14 Oct 2025

Dear Dr. Qadri,

We are pleased to inform you that your manuscript 'Prevalence of Shigellosis among household contacts of index cases in the EFGH catchment area, Dhaka, Bangladesh' has been provisionally accepted for publication in PLOS Neglected Tropical Diseases.

Before your manuscript can be formally accepted you will need to complete some formatting changes, which you will receive in a follow up email. A member of our team will be in touch with a set of requests.

Please note that your manuscript will not be scheduled for publication until you have made the required changes, so a swift response is appreciated.

IMPORTANT: The editorial review process is now complete. PLOS will only permit corrections to spelling, formatting or significant scientific errors from this point onwards. Requests for major changes, or any which affect the scientific understanding of your work, will cause delays to the publication date of your manuscript.

Should you, your institution's press office or the journal office choose to press release your paper, you will automatically be opted out of early publication. We ask that you notify us now if you or your institution is planning to press release the article. All press must be co-ordinated with PLOS.

Thank you again for supporting Open Access publishing; we are looking forward to publishing your work in PLOS Neglected Tropical Diseases.

Best regards,

Nicholas J Mantis

Academic Editor

PLOS Neglected Tropical Diseases

Stuart Blacksell

Section Editor

PLOS Neglected Tropical Diseases

Shaden Kamhawi

co-Editor-in-Chief

PLOS Neglected Tropical Diseases

orcid.org/0000-0003-4304-636XX

Paul Brindley

co-Editor-in-Chief

PLOS Neglected Tropical Diseases

orcid.org/0000-0003-1765-0002

***********************************************************

p.p1 {margin: 0.0px 0.0px 0.0px 0.0px; line-height: 16.0px; font: 14.0px Arial; color: #323333; -webkit-text-stroke: #323333}span.s1 {font-kerning: none

Reviewer's Responses to Questions

Key Review Criteria Required for Acceptance?

As you describe the new analyses required for acceptance, please consider the following:

Methods

-Are the objectives of the study clearly articulated with a clear testable hypothesis stated?

-Is the study design appropriate to address the stated objectives?

-Is the population clearly described and appropriate for the hypothesis being tested?

-Is the sample size sufficient to ensure adequate power to address the hypothesis being tested?

-Were correct statistical analysis used to support conclusions?

-Are there concerns about ethical or regulatory requirements being met?

Reviewer #1: yes, the objectives are clear

**********

Results

-Does the analysis presented match the analysis plan?

-Are the results clearly and completely presented?

-Are the figures (Tables, Images) of sufficient quality for clarity?

Reviewer #1: Yes

**********

Conclusions

-Are the conclusions supported by the data presented?

-Are the limitations of analysis clearly described?

-Do the authors discuss how these data can be helpful to advance our understanding of the topic under study?

-Is public health relevance addressed?

Reviewer #1: Yes

**********

Editorial and Data Presentation Modifications?

Use this section for editorial suggestions as well as relatively minor modifications of existing data that would enhance clarity. If the only modifications needed are minor and/or editorial, you may wish to recommend “Minor Revision” or “Accept”.

Reviewer #1: Figure legend is run-on with the Table 1

Table 2 has a wording problem in one field: "number of persons live per room"

I thought one weakness would be mentioned: no community control group to know the rate of shigella positives in the general population.

**********

Summary and General Comments

Use this section to provide overall comments, discuss strengths/weaknesses of the study, novelty, significance, general execution and scholarship. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. If requesting major revision, please articulate the new experiments that are needed.

Reviewer #1: This is much improved and the authors have addressed the concerns from my earlier review.

**********

PLOS authors have the option to publish the peer review history of their article (what does this mean? ). If published, this will include your full peer review and any attached files.

If you choose “no”, your identity will remain anonymous but your review may still be made public.

Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy .

Reviewer #1: No

PLoS Negl Trop Dis. doi: 10.1371/journal.pntd.0013644.r005

Acceptance letter

Ana LTO Nascimento

Dear Dr. Qadri,

We are delighted to inform you that your manuscript, "Prevalence of Shigellosis among household contacts of index cases in the EFGH catchment area, Dhaka, Bangladesh," has been formally accepted for publication in PLOS Neglected Tropical Diseases.

We have now passed your article onto the PLOS Production Department who will complete the rest of the publication process. All authors will receive a confirmation email upon publication.

The corresponding author will soon be receiving a typeset proof for review, to ensure errors have not been introduced during production. Please review the PDF proof of your manuscript carefully, as this is the last chance to correct any scientific or type-setting errors. Please note that major changes, or those which affect the scientific understanding of the work, will likely cause delays to the publication date of your manuscript. Note: Proofs for Front Matter articles (Editorial, Viewpoint, Symposium, Review, etc...) are generated on a different schedule and may not be made available as quickly.

Soon after your final files are uploaded, the early version of your manuscript will be published online unless you opted out of this process. The date of the early version will be your article's publication date. The final article will be published to the same URL, and all versions of the paper will be accessible to readers.

For Research Articles, you will receive an invoice from PLOS for your publication fee after your manuscript has reached the completed accept phase. If you receive an email requesting payment before acceptance or for any other service, this may be a phishing scheme. Learn how to identify phishing emails and protect your accounts at https://explore.plos.org/phishing.

Thank you again for supporting open-access publishing; we are looking forward to publishing your work in PLOS Neglected Tropical Diseases.

Best regards,

Shaden Kamhawi

co-Editor-in-Chief

PLOS Neglected Tropical Diseases

Paul Brindley

co-Editor-in-Chief

PLOS Neglected Tropical Diseases

Associated Data

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

    Supplementary Materials

    Attachment

    Submitted filename: Response to Reviewer comments.pdf

    pntd.0013644.s001.pdf (153.6KB, pdf)

    Data Availability Statement

    A de-identified analytical data set will be made available upon requests directed to the Institutional Review Board (IRB) coordinator of the icddr,b at info@icddrb.org. Only after approval of a proposal data can be shared through a secure online platform. Approval of the proposal will be subject to scientific review by the institutional review board at icddr,b. Sharing of data will also be subject to the published data access rules of the icddr,b. The requestor will need to sign a standard data access agreement required by the icddr,b.


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