Shigella sonnei utilises colicins during inter-bacterial competition.

The mammalian colon is one of the most densely populated habitats currently recognised, with 1011-1013 commensal bacteria per gram of colonic contents. Enteric pathogens must compete with the resident intestinal microbiota to cause infection. Among these enteric pathogens are Shigella species which cause approximately 125 million infections annually, of which over 90 % are caused by Shigella flexneri and Shigella sonnei. Shigella sonnei was previously reported to use a Type VI Secretion System (T6SS) to outcompete E. coli and S. flexneri in in vitro and in vivo experiments. S. sonnei strains have also been reported to harbour colicinogenic plasmids, which are an alternative anti-bacterial mechanism that could provide a competitive advantage against the intestinal microbiota. We sought to determine the contribution of both T6SS and colicins to the anti-bacterial killing activity of S. sonnei. We reveal that whilst the T6SS operon is present in S. sonnei, there is evidence of functional degradation of the system through SNPs, indels and IS within key components of the system. We created strains with synthetically inducible T6SS operons but were still unable to demonstrate anti-bacterial activity of the T6SS. We demonstrate that the anti-bacterial activity observed in our in vitro assays was due to colicin activity. We show that S. sonnei no longer displayed anti-bacterial activity against bacteria that were resistant to colicins, and removal of the colicin plasmid from S. sonnei abrogated anti-bacterial activity of S. sonnei. We propose that the anti-bacterial activity demonstrated by colicins may be sufficient for niche competition by S. sonnei within the gastrointestinal environment.

Molecular Signature of Monocytes Shaped by the Shigella sonnei 1790-Generalized Modules for Membrane Antigens Vaccine.

Shigellosis, an acute gastroenteritis infection caused by Shigella species, remains a public health burden in developing countries. Recently, many outbreaks due to Shigella sonnei multidrug-resistant strains have been reported in high-income countries, and the lack of an effective vaccine represents a major hurdle to counteract this bacterial pathogen. Vaccine candidates against Shigella sonnei are under clinical development, including a Generalized Modules for Membrane Antigens (GMMA)-based vaccine. The mechanisms by which GMMA-based vaccines interact and activate human immune cells remain elusive. Our previous study provided the first evidence that both adaptive and innate immune cells are targeted and functionally shaped by the GMMA-based vaccine. Here, flow cytometry and confocal microscopy analysis allowed us to identify monocytes as the main target population interacting with the S. sonnei 1790-GMMA vaccine on human peripheral blood. In addition, transcriptomic analysis of this cell population revealed a molecular signature induced by 1790-GMMA mostly correlated with the inflammatory response and cytokine-induced processes. This also impacts the expression of genes associated with macrophages' differentiation and T cell regulation, suggesting a dual function for this vaccine platform both as an antigen carrier and as a regulator of immune cell activation and differentiation.

WGS of a cluster of MDR Shigella sonnei utilizing Oxford Nanopore R10.4.1 long-read sequencing.

OBJECTIVES: To utilize long-read nanopore sequencing (R10.4.1 flowcells) for WGS of a cluster of MDR Shigella sonnei, specifically characterizing genetic predictors of antimicrobial resistance (AMR). METHODS: WGS was performed on S. sonnei isolates identified from stool and blood between September 2021 and October 2022. Bacterial DNA from clinical isolates was extracted on the MagNA Pure 24 and sequenced on the GridION utilizing R10.4.1 flowcells. Phenotypic antimicrobial susceptibility testing was interpreted based on CLSI breakpoints. Sequencing data were processed with BugSeq, and AMR was assessed with BugSplit and ResFinder. RESULTS: Fifty-six isolates were sequenced, including 53 related to the cluster of cases. All cluster isolates were identified as S. sonnei by sequencing, with global genotype 3.6.1.1.2 (CipR.MSM5), MLST 152 and PopPUNK cluster 3. Core genome MLST (cgMLST, examining 2513 loci) and reference-based MLST (refMLST, examining 4091 loci) both confirmed the clonality of the isolates. Cluster isolates were resistant to ampicillin (blaTEM-1), trimethoprim/sulfamethoxazole (dfA1, dfrA17; sul1, sul2), azithromycin (ermB, mphA) and ciprofloxacin (gyrA S83L, gyrA D87G, parC S80I). No genomic predictors of resistance to carbapenems were identified. CONCLUSIONS: WGS with R10.4.1 enabled rapid sequencing and identification of an MDR S. sonnei community cluster. Genetic predictors of AMR were concordant with phenotypic antimicrobial susceptibility testing.

Molecular mechanism of Hfq-dependent sRNA1039 and sRNA1600 regulating antibiotic resistance and virulence in Shigella sonnei.

Bacillary dysentery caused by Shigella spp. is a significant concern for human health. Small non-coding RNA (sRNA) plays a crucial role in regulating antibiotic resistance and virulence in Shigella spp. However, the specific mechanisms behind this phenomenon are still not fully understood. This study discovered two sRNAs (sRNA1039 and sRNA1600) that may be involved in bacterial resistance and virulence. By constructing deletion mutants (WT/ΔSR1039 and WT/ΔSR1600), this study found that the WT/ΔSR1039 mutants caused a two-fold increase in sensitivity to ampicillin, gentamicin and cefuroxime, and the WT/ΔSR1600 mutants caused a two-fold increase in sensitivity to cefuroxime. Furthermore, the WT/ΔSR1600 mutants caused a decrease in the adhesion and invasion of bacteria to HeLa cells (P<0.01), and changed the oxidative stress level of bacteria to reduce their survival rate (P<0.001). Subsequently, this study explored the molecular mechanisms by which sRNA1039 and sRNA1600 regulate antibiotic resistance and virulence. The deletion of sRNA1039 accelerated the degradation of target gene cfa mRNA and reduced its expression, thereby regulating the expression of pore protein gene ompD indirectly and negatively to increase bacterial sensitivity to ampicillin, gentamicin and cefuroxime. The inactivation of sRNA1600 reduced the formation of persister cells to reduce resistance to cefuroxime, and reduced the expression of type-III-secretion-system-related genes to reduce bacterial virulence by reducing the expression of target gene tomB. These results provide new insights into Hfq-sRNA-mRNA regulation of the resistance and virulence network of Shigella sonnei, which could potentially promote the development of more effective treatment strategies.

Vaccinomics-aided next-generation novel multi-epitope-based vaccine engineering against multidrug resistant Shigella Sonnei: Immunoinformatics and chemoinformatics approaches.

Shigella sonnei is a gram-negative bacterium and is the primary cause of shigellosis in advanced countries. An exceptional rise in the prevalence of the disease has been reported in Asia, the Middle East, and Latin America. To date, no preventive vaccine is available against S. sonnei infections. This pathogen has shown resistances towards both first- and second-line antibiotics. Therefore, an effective broad spectrum vaccine development against shigellosis is indispensable. In the present study, vaccinomics-aided immunoinformatics strategies were pursued to identify potential vaccine candidates from the S. sonnei whole proteome data. Pathogen essential proteins that are non-homologous to human and human gut microbiome proteome set, are feasible candidates for this purpose. Three antigenic outer membrane proteins were prioritized to predict lead epitopes based on reverse vaccinology approach. Multi-epitope-based chimeric vaccines was designed using lead B- and T-cell epitopes combined with suitable linker and adjuvant peptide sequences to enhance immune responses against the designed vaccine. The SS-MEVC construct was prioritized based on multiple physicochemical, immunological properties, and immune-receptors docking scores. Immune simulation analysis predicted strong immunogenic response capability of the designed vaccine construct. The Molecular dynamic simulations analysis ensured stable molecular interactions of lead vaccine construct with the host receptors. In silico restriction and cloning analysis predicted feasible cloning capability of the SS-MEVC construct within the E. coli expression system. The proposed vaccine construct is predicted to be more safe, effective and capable of inducing robust immune responses against S. sonnei infections and may be worthy of examination via in vitro/in vivo assays.

Escherichia coli killing by epidemiologically successful sublineages of Shigella sonnei is mediated by colicins.

BACKGROUND: Shigella sp. are enteric pathogens which causes >125 million cases of shigellosis annually. S. sonnei accounts for about a quarter of those cases and is increasingly prevalent in industrialising nations. Being an enteric pathogen, S. sonnei benefits from outcompeting gut commensals such as Escherichia coli to establish itself and cause disease. There are numerous mechanisms that bacterial pathogens use to outcompete its rivals including molecules called colicins. A Type 6 Secretion System (T6SS) was recently described as contributing to E. coli killing in S. sonnei. METHODS: We used Bulk Phenotyping of Epidemiological Replicates (BPER) which combined bacterial Genome Wide Association Studies (bGWAS) and high throughput phenotyping on a collection of S. sonnei surveillance isolates to identify the genetic features associated with E. coli killing and explore their relationship with epidemiological behaviour. We further explored the presence of colicins and T6SS components in the isolates using genomics, laboratory experimentation, and proteomics. FINDINGS: Our bGWAS analysis returned known and novel colicin and colicin related genes as significantly associated with E. coli killing. In silico analyses identified key colicin clusters responsible for the killing phenotype associated with epidemiologically successful sub-lineages. The killing phenotype was not associated with the presence of a T6SS. Laboratory analyses confirmed the presence of the key colicin clusters and that killing was contact-independent. INTERPRETATION: Colicins are responsible for E. coli killing by S. sonnei, not a T6SS. This phenotype contributes to shaping the observed epidemiology of S. sonnei and may contribute to its increasing prevalence globally. BPER is an epidemiologically relevant approach to phenotypic testing that enables the rapid identification of genetic drivers of phenotypic changes, and assessment of their relevance to epidemiology in natural settings. FUNDING: Biotechnology and Biological Sciences Research Council, Biotechnology and Biological Sciences Research Council Doctoral Training Partnership studentship, Wellcome Trust, Medical Research Council (UK), French National Research Agency.

Case of Extensively Drug-Resistant Shigella sonnei Infection, United States.

We report extensively drug-resistant (XDR) Shigella sonnei infection in an immunocompromised patient in Texas, USA. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry failed to identify XDR Shigella, but whole-genome sequencing accurately characterized the strain. First-line antimicrobials are not effective against emerging XDR Shigella. Fosfomycin, carbapenems, and tigecycline are potential alternatives.

Multidrug-Resistant Shigella sonnei Bacteremia among Persons Experiencing Homelessness, Vancouver, British Columbia, Canada.

Increased invasive bloodstream infections caused by multidrug resistant Shigella sonnei were noted in Vancouver, British Columbia, Canada, during 2021-2023. Whole-genome sequencing revealed clonal transmission of genotype 3.6.1.1.2 (CipR.MSM5) among persons experiencing homelessness. Improvements in identifying Shigella species, expanding treatment options for multidrug resistant infections, and developing public health partnerships are needed.

The evolution and international spread of extensively drug resistant Shigella sonnei.

Shigella sonnei causes shigellosis, a severe gastrointestinal illness that is sexually transmissible among men who have sex with men (MSM). Multidrug resistance in S. sonnei is common including against World Health Organisation recommended treatment options, azithromycin, and ciprofloxacin. Recently, an MSM-associated outbreak of extended-spectrum ß-lactamase producing, extensively drug resistant S. sonnei was reported in the United Kingdom. Here, we aimed to identify the genetic basis, evolutionary history, and international dissemination of the outbreak strain. Our genomic epidemiological analyses of 3,304 isolates from the United Kingdom, Australia, Belgium, France, and the United States of America revealed an internationally connected outbreak with a most recent common ancestor in 2018 carrying a low-fitness cost resistance plasmid, previously observed in travel associated sublineages of S. flexneri. Our results highlight the persistent threat of horizontally transmitted antimicrobial resistance and the value of continuing to work towards early and open international sharing of genomic surveillance data.

Involvement of RNA chaperone hfq in the regulation of antibiotic resistance and virulence in Shigella sonnei.

The host factor for RNA phage Qß replicase (Hfq) is a crucial post-transcriptional regulator in many bacterial pathogens, facilitating the interaction between small non-coding RNAs (sRNAs) and their target mRNAs. Studies have suggested that Hfq plays a role in antibiotic resistance and virulence in bacteria, although its functions in Shigella are not fully understood. In this study, we investigated the functional roles of Hfq in Shigella sonnei (S. sonnei) by constructing an hfq deletion mutant. Our phenotypic assays showed that the hfq deletion mutant was more sensitivity to antibiotics and had impaired virulence. Transcriptome analyses supported the results concerning the phenotype of the hfq mutant and showed that differentially expressed genes were mainly enriched in the KEGG pathways two-component system, ABC transporters, ribosome, and Escherichia coli biofilm formation. Additionally, we predicted eleven novel Hfq-dependent sRNAs, which were potentially involved in the regulation of antibiotic resistance and/or virulence in S. sonnei. Our findings suggest that Hfq plays a post-transcriptional role in regulating antibiotic resistance and virulence in S. sonnei, and could provide a basis for future studies on Hfq-sRNA-mRNA regulatory networks in this important pathogen.

Complete genome sequence and antimicrobial resistance analysis of ESBL-producing Shigella sonnei carrying small cryptic plasmids isolated in northern Italy.

OBJECTIVES: Herein, we sequenced and assembled the genome of a Shigella sonnei isolate carrying several small plasmids using a hybrid approach that combined Oxford Nanopore Technologies and Illumina platforms. METHODS: Whole-genome sequencing was conducted using the Illumina iSeq 100 and Oxford Nanopore MinION systems, and the resulting reads were used for hybrid genome assembly via Unicycler. Coding sequences were annotated using RASTtk, while genes involved in antimicrobial resistance and virulence were identified using AMRFinderPlus. Plasmid nucleotide sequences were aligned to the NCBI non-redundant database using BLAST, and replicons were identified using PlasmidFinder. RESULTS: The genome consisted of 1 chromosome (4 801 657 bp), 3 major plasmids (212 849 bp, 86 884 bp, and 83 425 bp, respectively) and 12 small cryptic plasmids (ranging from 8390 bp to 1822 bp). BLAST analysis revealed that all plasmids were highly similar to previously deposited sequences. Genome annotation predicted 5522 coding regions, including 19 antimicrobial resistance genes and 17 virulence genes. Four of the antimicrobial resistance genes were located in small plasmids, and four of the virulence genes were located in a large virulence plasmid. CONCLUSION: The presence of antimicrobial resistance genes in small cryptic plasmids may represent an overlooked mechanism for the propagation of these genes among bacterial populations. Our work provides new data on these elements that may inform the development of new strategies to control the spread of extended spectrum ß-lactamase-producing bacterial strains.

Rapid emergence of extensively drug-resistant Shigella sonnei in France.

Shigella sonnei, the main cause of bacillary dysentery in high-income countries, has become increasingly resistant to antibiotics. We monitored the antimicrobial susceptibility of 7121 S. sonnei isolates collected in France between 2005 and 2021. We detected a dramatic increase in the proportion of isolates simultaneously resistant to ciprofloxacin (CIP), third-generation cephalosporins (3GCs) and azithromycin (AZM) from 2015. Our genomic analysis of 164 such extensively drug-resistant (XDR) isolates identified 13 different clusters within CIP-resistant sublineage 3.6.1, which was selected in South Asia ∼15 years ago. AZM resistance was subsequently acquired, principally through IncFII (pKSR100-like) plasmids. The last step in the development of the XDR phenotype involved various extended-spectrum beta-lactamase genes (blaCTX-M-3, blaCTX-M-15, blaCTX-M-27, blaCTX-M-55, and blaCTX-M-134) carried by different plasmids (IncFII, IncI1, IncB/O/K/Z) or even integrated into the chromosome, and encoding resistance to 3GCs. This rapid emergence of XDR S. sonnei, including an international epidemic strain, is alarming, and good laboratory-based surveillance of shigellosis will be crucial for informed decision-making and appropriate public health action.

A Shigella sonnei clone with extensive drug resistance associated with waterborne outbreaks in China.

Antimicrobial resistance of Shigella sonnei has become a global concern. Here, we report a phylogenetic group of S. sonnei with extensive drug resistance, including a combination of multidrug resistance, coresistance to ceftriaxone and azithromycin (cefRaziR), reduced susceptibility to fluoroquinolones, and even colistin resistance (colR). This distinct clone caused six waterborne shigellosis outbreaks in China from 2015 to 2020. We collect 155 outbreak isolates and 152 sporadic isolates. The cefRaziR isolates, including outbreak strains, are mainly distributed in a distinct clade located in global Lineage III. The outbreak strains form a recently derived monophyletic group that may have emerged circa 2010. The cefRaziR and colR phenotypes are attributed to the acquisition of different plasmids, particularly the IncB/O/K/Z plasmid coharboring the blaCTX-M-14, mphA, aac(3)-IId, dfrA17, aadA5, and sul1 genes and the IncI2 plasmid with an mcr-1 gene. Genetic analyses identify 92 accessory genes and 60 single-nucleotide polymorphisms associated with the cefRaziR phenotype. Surveillance of this clone is required to determine its dissemination and threat to global public health.

Development of multiplex cross displacement amplification combined with lateral flow biosensor assay for detection of virulent shigella sonnei.

Shigella sonnei is the most common Shigella spp. in developed areas and the second most common in undeveloped regions. In this study, a multiple cross displacement amplification (MCDA) assay was used in combination with a lateral flow biosensor (LFB) assay to detect virulent S. sonnei strains containing the ipaH and wbgX genes. The multiplex MCDA-LFB assay detected wbgX at ≥1 pg/µL and ipaH at ≥10 fg/µL within 30 min in pure cultures maintained at 63°C. This assay was sensitive for ~37 CFU of virulent S. sonnei and ~3.7 CFU of Shigella spp. and enteroinvasive E. coli in stimulated fecal samples and had 100% specificity among 59 reference strains. The MCDA-LFB assay was also able to differentiate between virulent S. sonnei and other Shigella spp. and enteroinvasive E. coli among 99 clinical isolates. In summary, a multiplex MCDA-LFB assay was developed for rapid, convenient, point-of-care, and accurate identification of virulent S. sonnei within 30 min and at a constant temperature without the need for expensive lab equipment.

The Effect of Mutation in Lipopolysaccharide Biosynthesis on Bacterial Fitness.

This paper presents the genome sequence of a Shigella sonnei mutant strain (S. sonnei 4351) and the effect of mutation in lipopolysaccharide biosynthesis on bacterial fitness. Lipopolysaccharides are the major component of the outer leaflet of the Gram-negative outer membrane. We report here a frameshift mutation of the gene gmhD in the genome of S. sonnei 4351. The mutation results in a lack of epimerization of the core heptose while we also found increased thermosensitivity, abnormal cell division, and increased susceptibility to erythromycin and cefalexin compared to the S. sonnei 4303. Comparative genomic analysis supplemented with structural data helps us to understand the effect of specific mutations on the virulence of the bacteria and may provide an opportunity to study the effect of short lipopolysaccharides.

Outbreak of sexually transmitted, extensively drug-resistant Shigella sonnei in the UK, 2021-22: a descriptive epidemiological study.

BACKGROUND: Shigellosis, traditionally a foodborne and waterborne infection, causes substantial morbidity globally. It is now a leading cause of sexually transmitted gastroenteritis among gay, bisexual, and other men who have sex with men (MSM). We describe an ongoing outbreak of extensively drug-resistant (XDR) Shigella sonnei in the UK. METHODS: Routine laboratory surveillance (Second Generation Surveillance System, Gastrointestinal Data Warehouse) identified an exceedance of S sonnei clade 5 in England, first detected in September, 2021. Cases within this clade were subsequently reported from Scotland, Wales, and Northern Ireland. Confirmed cases in this outbreak were defined as individuals diagnosed with S sonnei clade 5 in the UK, with a specimen date between Sept 1, 2021, and Feb 9, 2022, who were genomically confirmed as part of a ten-single nucleotide polymorphism (SNP) linkage cluster. We used whole-genome sequencing with SNP typing to identify genomic clusters and antimicrobial-resistance determinants, analysing cases across the UK. We collected demographic, epidemiological, and clinical data from people infected with S sonnei clade 5 in England using questionnaires (standard and bespoke outbreak questionnaires). We used descriptive summary statistics to characterise cases. FINDINGS: 72 cases (70 [97%] male, median age 34 years [IQR 27-39]) belonging to the ten-SNP single linkage cluster of S sonnei clade 5 were identified between Sept 4, 2021, and Feb 9, 2022. Isolates were predominantly XDR, with 66 (92%) of 72 harbouring blaCTX-M-27, a plasmid-mediated gene for production of extended-spectrum ß-lactamases (ESBLs). Of 33 cases with clinical data, 19 (58%) received antibiotics and eight (24%) were hospitalised. 21 (78%) of 27 cases with completed bespoke outbreak questionnaires were HIV-negative MSM taking HIV pre-exposure prophylaxis (PrEP) who reported sexual contacts in the UK and Europe within the incubation period. INTERPRETATION: We highlight the rapid dissemination of XDR ESBL-producing S sonnei in sexual networks of MSM. We recommend strengthening shigella testing where clinically indicated, antimicrobial-resistance surveillance, and integrated health promotion messaging among all MSM, including PrEP users, to reduce the burden of shigellosis. FUNDING: National Institute for Health Research Health Protection Research Unit in Gastrointestinal Infections at the University of Liverpool in partnership with the UK Health Security Agency.

Characterization of Extended-Spectrum ß-Lactamase-Producing Shigella sonnei in Spain: Expanding the Geographic Distribution of Sequence Type 152/CTX-M-27 Clone.

We describe the first occurrence in Spain of community cases of CTX-M-27-producing Shigella sonnei sequence type 152 (ST152), resistant to quinolones and azithromycin. The cases included adult males and also one pediatric case. The isolates were clustered together with an Australian isolate and differed from other outbreak-causing strains in England by more than 50 alleles. They carried the blaCTX-M-27 gene on an 83-Kb F2:A-:B- plasmid, similar to that found in a British isolate.

Clinical and Genomic Investigation of an International Ceftriaxone- and Azithromycin-Resistant Shigella sonnei Cluster among Men Who Have Sex with Men, Montréal, Canada 2017-2019.

Multidrug-resistant (MDR) Shigella sonnei have become prevalent among men who have sex with men and have become a global public health concern. From June 2017 to April 2019, 32 men were infected with MDR S. sonnei acquired locally, in Montréal, which was suggestive of an outbreak. Antimicrobial susceptibility testing, whole-genome sequencing (WGS), phylogenetic analysis, antimicrobial resistance and virulence characterization, and association to international clusters were performed. The outbreak strain was ceftriaxone- and azithromycin-resistant due to the acquisition of blaCTX-M-27, and mphA and ermB genes, respectively, with reduced susceptibility to ciprofloxacin due to a single point mutation (gyrA S83L). One out of 27 patients treated with a fluoroquinolone experienced microbiological failure. Epidemiological evidence first supported by a rare unique MDR Shigella sonnei documented only in men in 2017 followed by similar pulsed-field gel electrophoresis profiles was confirmed by WGS. A core genome high-quality single-nucleotide variant (hqSNV)-based phylogeny found a median of 6 hqSNV differences among isolates. Virulence gene content was investigated, but no Shiga toxins were detected. An international cluster of highly related isolates was identified (PDS000019750.208) and belonged to the 3.7.29.1.4.1 S. sonnei genotype (Global III VN2.KH1.Aus). Genomic analysis revealed that this Montréal cluster was connected to other documented outbreaks in Australia, the United States, and the United Kingdom. This study highlights the urgent need for public health measures to focus on the prevention and the early detection of S. sonnei, since global transmission patterns of MDR strains is concerning and few antimicrobial treatment options are available. IMPORTANCE Shigella sonnei, an important foodborne pathogen, recently became a frequent sexually transmitted agent involved in large and persistent outbreaks globally among men who have sex with men. Most strains also harbor several multidrug-resistant (MDR) determinants of particular concern. This study characterizes an outbreak strain at the source of an important MDR cluster identified in Montréal in 2017. Associations were made to many high-profile international outbreaks, and the causative S. sonnei lineage of these clusters was identified, which was not evident in past reports. The worldwide occurrence of this strain is of concern since treatment with antimicrobials like ceftriaxone and azithromycin may not be effective, and rare microbiological failures have been documented in patients treated with ciprofloxacin. Our investigation highlights the threats of Shigella spp. infection and the necessity for antimicrobial susceptibility monitoring in order to mitigate S. sonnei's impact on public health and to avoid transmission to other at-risk communities.

Phylogenetic analysis and antibiotic resistance of Shigella sonnei isolates.

Shigellosis is one of the most important gastric infections caused by different species of Shigella, and has been regarded as a serious threat to public health. Lineage/sublineage profile of Shigella sonnei is strongly associated with the antibiotic resistance and population structure of this pathogen. In this study, we determined the phylogeny and antibiotic resistance profiles of S. sonnei strains, isolated from 1246 stool and 580 food samples, using multiplex PCR-HRMA genotyping and Kirby-Bauer disk diffusion methods, respectively. A total of 64 S. sonnei strains were isolated (13 food and 51 clinical isolates). Multiplex PCR-HMR assay was able to differentiate the lineages II and III, and sublineages IIIb and IIIc strains successfully considering the definite melting curves and temperatures. Lineage I and sublineage IIIa strain were not isolated in this study. We also demonstrated that most of the S. sonnei strains isolated from both food and clinical samples clustered within the lineage III and sublineage IIIc. Resistance against trimethoprim-sulfamethoxazole, tetracycline, chloramphenicol, and streptomycin antibiotics were the most prevalent phenotypes among the S. sonnei lineage III and sublineage IIIc strains.