SERS-based immunocapture and detection of pathogenic bacteria using a boronic acid-functionalized polydopamine-coated Au@Ag nanoprobe.

A surface-enhanced Raman scattering (SERS)-based immunocapture nanoprobe is described for the detection of pathogenic bacteria. The probe uses boronic acid-functionalized polydopamine-coated Au@Ag nanoparticles as an advanced SERS nanotag. Modified magnetic IgG@Fe3O4 nanoparticles are used for magnetic separation. Au@Ag@PDA nanoparticles, where PDA stands for polydopamine, were functionalized with boronic acid to bind to pathogenic bacteria and induce signal amplification. The Raman signal is amplified 108 times when the SERS tag binds the surface of bacteria. The SERS spectra exhibit fingerprint-like patterns that enable bacterial classification. The results of principal component analysis (PCA) and hierarchical cluster analysis (HCA) of the spectral regions were compared. The bacterial surface protein and glycan signals (1300-1450 cm-1) were the best regions for bacterial classification. Staphylococcus aureus, Escherichia coli, Shigella dysenteriae, Pseudomonas aeruginosa, and Klebsiella pneumonia were successfully classified by this method. The lowest detection limit was 10 colonies/mL (CFU·mL). The assay can be completed within 30 min. Conceivably, this method may be extended to the quantitative detection or classification of bacteria under various other conditions. Graphical abstract Schematic representation of immunocapture and detection of pathogenic bacteria using boronic acid-functionalized polydopamine-coated Au@Ag nanoprobe through the bacterial surface protein and glycan signals. Green arrow: laser; black arrow: SERS; red ball: bacteria; grey ball: IgG@Fe3O4; golden ball: boronic acid-functionalized Au@Ag@PDA.

Development of quinolone resistance and prevalence of different virulence genes among Shigella flexneri and Shigella dysenteriae in environmental water samples.

The purpose of this study was to find out the mechanism of quinolone resistance in Shigella sp. isolated from environmental water samples from various parts of Kolkata, India. Out of 196 Shigella sp. isolated from 2014 to 2017, we selected 32 Shigella isolates for antimicrobial susceptibility tests. The minimum inhibitory concentrations (MIC) for quinolones ranged from 30 to 50 µg ml-1 for ofloxacin, 5-20 µg ml-1 for ciprofloxacin and 20-30 µg ml-1 for norfloxacin. A few amino acid changes were found in quinolone resistance determining region (QRDR) of gyrA. Mutations in gyrA lead to a higher increment of MIC of quinolones. Among the plasmid-mediated (PMQR) quinolone resistance genes investigated, qnrB and aac(6')-lb-cr genes were found in all isolates. qnrA and qnrS were found in 25% and 62% of the isolates, respectively. ipaH gene was found in all of the isolates followed by the presence of other virulence genes ial, sen and stx1. Almost all the isolates having high MICs showed efflux pump activity in drug accumulation assay. All the mechanisms may or may not be present in a single strain. Several types of efflux pumps, presence of PMQR genes and mutations in drug target site of QRDR region may play the crucial role for resistance in our isolates.

An impedimetric aptasensor for Shigella dysenteriae using a gold nanoparticle-modified glassy carbon electrode.

This work describes an aptasensor for the foodborne pathogen Shigella dysenteriae (S. dysenteriae). A glassy carbon electrode (GCE) was modified with gold nanoparticles (AuNPs) by electrodeposition. Then, thiolated aptamer for S. dysenteriae detection was self-assembled on the surface of the modified GCE, and any free residual AuNPs were blocked with 6-mercapto-1-hexanol. The size, morphology, and distribution of the AuNPs were characterized by field emission scanning electron microscopy. Detection of S. dysenteriae was performed measurement of the charge transfer resistance (Rct) before and after addition of S. dysenteriae using hexacyanoferrate as an electrochemical probe. The interaction between the aptamer and outer-membrane proteins of S. dysenteriae lead to an increase in the Rct of the sensor. The assay has a linear dynamic range that extends from 101 to 106 CFU.mL-1 and a limit of detection of 100 CFU.mL-1. It can differentiate between alive S. dysenteriae and other pathogens. Dead S. dysenteriae cells do not have any effect on selectivity. Unpasteurized and pasteurized skim milk and some water samples were spiked with S. dysenteriae and then successfully examined by this method. The results were validated by real-time PCR. The method is fast, low-cost, highly sensitive, and specific. Hence, it represents a valuable tool in food quality control. Graphical abstract Schematic presentation of a label free impedimetric aptasensor for Shigella dysenteriae using a glassy carbon electrode modified with gold nanoparticles (AuNPs) and 6-mercapto-1-hexanol (MCH). The limit of detection of this aptasensor is as low as 1 CFU.mL-1 for target bacteria.

Antimicrobial resistance profiles of Shigella dysenteriae isolated from travellers returning to the UK, 2004-2017.

PURPOSE: Antimicrobial resistance (AMR) profiles of 754 strains of Shigella dysenteriae isolated between 2004 and 2017 from UK travellers reporting symptoms of gastrointestinal (GI) disease were reviewed to look for evidence of emerging AMR associated with travellers' diarrhoea. METHODOLOGY: A travel history was provided for 72.7 % (548/754) of cases, of which 90.9 % (498/548) reported travel outside the UK within 7 days of onset of symptoms, and 9.1 % (50/498) reported no travel in that time frame. During the course of this study, whole genome sequencing (WGS) was implemented for GI disease surveillance, and we compared phenotypic AMR profiles with those derived from WGS data (n=133).Results/Key findings. The phenotypic and genotypic AMR results correlated well, with 90.1 % (121/133) isolates having concordant results to 10 classes of antimicrobials. Resistance to the first-line drugs commonly used in the treatment of shigellosis was observed throughout the study (ampicillin, 54.1%; chloramphenicol, 33.7 %; sulphonamides, 76.0 %; trimethoprim, 80.0%). Between 2004 and 2017, resistance to all classes of antimicrobials (except the phenicols) increased. The proportion of isolates exhibiting reduced susceptibility to ciprofloxacin increased from 3.8 % in 2004 to 75.7 % in 2017, and this was significantly associated with cases reporting travel to Asia compared to Africa (P<0.001). Of the 201 sequenced isolates, 3.0 % (20/201) had either blaCTX-M-15 or blaCMY-4. CONCLUSIONS: Increasing MDR, along with resistance to the fluroquinolones and the third generation cephalosporins, in Shigella dysenteriae causing travellers' diarrhoea provides further evidence for the need to regulatethe use of antimicrobial agents and continuous monitoring of emerging AMR.

Risk factors and outcome of Shigella encephalopathy in Bangladeshi children.

BACKGROUND: Although, Shigella encephalopathy, a serious extra-intestinal complication of shigellosis, significantly increases the risks of death, data are very limited on predicting factors particularly related to electrolyte profiles in children below five years of age with Shigella encephalopathy. Our objective was to determine the clinical as well as laboratory predicting factors and outcome of children with Shigella encephalopathy. METHODOLOGY/PRINCIPAL FINDINGS: In this unmatched case-control design, children aged 2-59 months having a positive stool culture for Shigella and who had their serum electrolytes been done from July 2012 to June 2015 were studied. Children with Shigella encephalopathy, defined as having abnormal mentation, constituted the cases, and those without encephalopathy constituted the controls. During the study period, we identified a total of 541 children less than five years of age, who had Shigella in their stool culture. Only 139 children fulfilled the study criteria and among them 69 were cases and 70 were controls. The cases more often had fatal outcome compared to the controls (7% vs. 0%, P = 0.02). In logistic regression analysis, the cases were independently associated with shorter duration (1.2 ± 0.4 days) of diarrhea prior to admission, dehydrating diarrhea, sepsis and hyponatremia (p<0.05 for all). Among 139 Shigella isolates, S. flexneri (88/139, 63%) and S. sonnei(34/139, 24%) were the dominant species. S. dysenteriae was not isolated throughout the study period. S.sonnei was more frequently isolated from the cases (24/69, 35%) than the controls (10/70, 14%), whereas the isolation of S. flexneri was comparable between the groups (40/69, 58% vs 48/70, 69%). A total of 94 (67.6%) isolates were resistant to trimethoprim-sulphamethoxazole, 84 (60.4%) to ciprofloxacin, 66/138 (48%) to ampicillin, 5 (3.5%) to ceftriaxone, 17 (12.2%) to mecillinum and 35 (25%) to azithromycin. CONCLUSIONS/SIGNIFICANCE: The case-fatality-rate was significantly higher among the children with Shigella encephalopathy compared to those without encephalopathy. Early identification and aggressive management of simple risk factors for Shigella encephalopathy may help to reduce morbidity and deaths in such children especially in resource-limited settings.

Shigella dysenteriae infection activates proinflammatory response through ß-catenin/NF-κB signaling pathway.

Shigella dysenteriae (S.dysenteriae) the causative agent of bacillary dysentery invades the human colonic epithelium resulting in severe intestinal inflammatory response and epithelial destruction. However, the mechanism by which S.dysenteriae infection regulates proinflammatory cytokines during intestinal inflammation is still obscure. In this study, we evaluated whether the interaction of ß-catenin and NF-κB regulates proinflammatory cytokines TNF-α and IL-8 by modulating GSK-3ß activity during S.dysenteriae infection in rat ileal loop model. Here we demonstrated that S.dysenteriae infection stimulate ß-catenin degradation which in turn decreased the association between NF-κB and ß-catenin. Also, we showed that S.dysenteriae infection increased GSK-3ß kinase activity which in turn phosphorylates ß-catenin for its degradation by ubiquitination and upregulates IL-8 through NF-κB activation thereby leading to inflammation. Thus these findings revealed the role of ß-catenin/ NF-κB and GSK-3ß in modulating the inflammatory response during bacterial infection and also showed that ß-catenin acts as a critical regulator of inflammation.

Shigellosis in Nepal: 13 years review of nationwide surveillance.

BACKGROUND: Shigella is a major cause of gastroenteritis especially in children. In developing countries, the incidence is frequent and results are often life threatening. Changing epidemiology and emerging antibiotic resistance warrants continuous monitoring of susceptibility. The present study highlights the changing epidemiology and drug resistance patterns of Shigella isolated at different hospitals of Nepal over a period of 13 years (Jan. 2003-Dec. 2015). METHODS: This study was carried out in 12 participating laboratories. Stool specimens received at respective laboratories were processed for isolation and identification of Shigella species and confirmed by serotyping at National Public Health Laboratory. Antimicrobial resistance patterns were determined by Kirby Baeur disc diffusion test. RESULTS: A total of 332 isolates were identified as Shigella species of which Shigella flexneri (50 %) was the predominant serotype. Shigella dysenteriae, Shigella sonnei, Shigella boydii, and untypable Shigella spp. respectively, accounted for 28.6, 27.54, 10.2, 4.5, and 6.6 % of the total number. Change in prevalent serotype is noted over the years. S. dysenteriae was the prevalent species in Nepal in 2003 and 2004, but since 2005, S. flexneri remained prevalent. Majority of the isolates were recovered from children aged 1-10 years and was statistically significant (p = 0.023) compared to the other age groups. High resistance among all Shigella species to the first-line drugs like ampicillin (88 %), cotrimoxazole (76 %), ciprofloxacin (39 %,) and nalidixic acid (80 %) was observed; 46.1 % of total isolates were multidrug resistant (MDR), and the most common MDR profile was ampicillin, nalidixic acid, and co-trimoxazole. Prevalence of MDR increased significantly in 2010 as compared to 2003. Only few Shigella isolates were resistant to ceftriaxone. CONCLUSIONS: The study revealed S. flexneri as the predominant serogroup in Nepal. Children below 10 years were more prone to the disease. Nalidixic acid, ampicillin, co-trimoxazole, and ciprofloxacin should not be used empirically as the first-line drugs in treatment of shigellosis. Since the distribution of different species of Shigella and their antibiotic susceptibility profile may vary from one geographical location to another and may also change with time, continuous local monitoring of resistance patterns is necessary for appropriate antimicrobial therapy.

Global phylogeography and evolutionary history of Shigella dysenteriae type 1.

Together with plague, smallpox and typhus, epidemics of dysentery have been a major scourge of human populations for centuries(1). A previous genomic study concluded that Shigella dysenteriae type 1 (Sd1), the epidemic dysentery bacillus, emerged and spread worldwide after the First World War, with no clear pattern of transmission(2). This is not consistent with the massive cyclic dysentery epidemics reported in Europe during the eighteenth and nineteenth centuries(1,3,4) and the first isolation of Sd1 in Japan in 1897(5). Here, we report a whole-genome analysis of 331 Sd1 isolates from around the world, collected between 1915 and 2011, providing us with unprecedented insight into the historical spread of this pathogen. We show here that Sd1 has existed since at least the eighteenth century and that it swept the globe at the end of the nineteenth century, diversifying into distinct lineages associated with the First World War, Second World War and various conflicts or natural disasters across Africa, Asia and Central America. We also provide a unique historical perspective on the evolution of antibiotic resistance over a 100-year period, beginning decades before the antibiotic era, and identify a prevalent multiple antibiotic-resistant lineage in South Asia that was transmitted in several waves to Africa, where it caused severe outbreaks of disease.

Competitive Survival of Escherichia coli, Vibrio cholerae, Salmonella typhimurium and Shigella dysenteriae in Riverbed Sediments.

Studies on the survival of bacterial enteric pathogens in riverbed sediments have mostly focused on individual organisms. Reports on the competitive survival of these pathogens in riverbed sediments under the same experimental setup are limited. We investigated the survival of Escherichia coli, Salmonella enterica ser. Typhimurium, Vibrio cholerae and Shigella dysenteriae in riverbed sediments of the Apies River. Experiments were performed in flow chambers containing three sediment types and connected to aquarium pumps immersed in river water to maintain continuous water circulation. Each chamber was inoculated with ~107 CFU/mL (final concentration) of each microorganism and kept at 4, 20 and 30 °C. Chambers were sampled on days 0, 1, 2, 7, 14 and 28. At 4 °C, only E. coli and S. typhimurium survived throughout the 28 experimental days. V. cholerae had the shortest survival time at this temperature and was not detected in any of the sediment chambers 24 h after inoculation. S. dysenteriae only survived until day 7. At an increased temperature of 20 °C, only S. dysenteriae was not detected on day 28 of the experiment. At 30 °C, V. cholerae and Salmonella survived longer (28 days) than E. coli (14 days) and S. dysenteriae (4 days). Vibrio cholerae was shown to have the highest T 90 values (32 days) in all sediment types at 20 and 30 °C. We conclude that the sediments of the Apies River present a favourable environment for the survival of indicator and pathogenic bacteria depending on the prevailing temperature.

[Microbiological approach to a possible infective endocarditis case caused by Aggregatibacter actinomycetemcomitans]. / Aggregatibacter actinomycetemcomitans kaynakli olasi enfektif endokardit olgusuna yaklasim.

Aggregatibacter (Actinobacillus) actinomycetemcomitans, a small, gram-negative coccobacillus that grows slow and fastidious, is generally colonized in the oral cavity. It is a rarely seen bacterium because of the difficulty of isolation but it can be a causative agent for dental infections and infective endocarditis (IE) particularly in the persons having prosthetic heart valves. In this report, a possible IE case caused by A.actinomycetemcomitans in a patient with aortic valve replacement has been presented. A 36-year-old man has admitted to Trakya University Hospital, Health Center for Medical Research and Practice, with the complaints of chills, malaise, intermittent fever, severe arthralgia and weight loss (20 kg). During his follow-up period, the blood cultures that were obtained three week intervals yielded the identical gram-negative coccobacilli morphology. The patient was then diagnosed as possible IE on the basis of having one major (growth of the typical microorganisms that may cause IE in two different blood cultures) and two minor (presence of prosthetic valve and high fever) criterias. The isolate could not be identified with conventional methods, while it was identified as Francisella tularensis with VITEK 2 (bioMerieux, France) system. Hence this identification was not confirmed by real-time Taqman polymerase chain reaction, so MALDI-TOF mass spectrometry was used to identify this bacteria. In the first run of the study, the isolate was named as Shigella dysenteriae initially, however when it was retested the next day it was identified as A.actinomycetemcomitans. In order to enlighten these conflicting results, 16S and 23S ribosomal DNA sequence analysis was performed, and consequently the bacterium was identified as A.actinomycetemcomitans. Doxycycline (2 x 100 mg po, 20 days) and streptomycin (2 x 10 mg/kg im, 10 days) therapy were initiated, considering the initial suspicious identification (F.tularensis), and on the fifth day of therapy the blood culture was negative with the regression of patient's complaints. Therapy continued with the addition of rifampicin to doxycycline from the 21(st) day and the patient discharged with cure. As a result, the identification of an exceptional bacterium like A.actinomycetemcomitans may be difficult and time-consuming in certain laboratory facilities. So, the use of different identification methods in addition to classical methods are needed to overcome such a problem, especially for uncommon isolates and clinically discordant cases. This case was presented because A.actinomycetemcomitans is a rare etiological agent for IE patients and it could be a good example to draw attention to the problem when identifying the organism using automatized identification systems.

Prevalence of Shiga toxin-producing Shigella species isolated from French travellers returning from the Caribbean: an emerging pathogen with international implications.

Shiga toxins (Stxs) are potent cytotoxins that inhibit host cell protein synthesis, leading to cell death. Classically, these toxins are associated with intestinal infections due to Stx-producing Escherichia coli or Shigella dysenteriae serotype 1, and infections with these strains can lead to haemolytic-uraemic syndrome. Over the past decade, there has been increasing recognition that Stx is produced by additional Shigella species. We recently reported the presence and expression of stx genes in Shigella flexneri 2a clinical isolates. The toxin genes were carried by a new stx-encoding bacteriophage, and infection with these strains correlated with recent travel to Haiti or the Dominican Republic. In this study, we further explored the epidemiological link to this region by utilizing the French National Reference Centre for Escherichia coli, Shigella and Salmonella collection to survey the frequency of Stx-producing Shigella species isolated from French travellers returning from the Caribbean. Approximately 21% of the isolates tested were found to encode and produce Stx. These isolates included strains of S. flexneri 2a, S. flexneri Y, and S. dysenteriae 4. All of the travellers who were infected with Stx-producing Shigella had recently travelled to Haiti, the Dominican Republic, or French Guiana. Furthermore, whole genome sequencing showed that the toxin genes were encoded by a prophage that was highly identical to the phage that we identified in our previous study. These findings demonstrate that this new stx-encoding prophage is circulating within that geographical area, has spread to other continents, and is capable of spreading to multiple Shigella serogroups.

Isolation and development of bioluminescent reporter phages for bacterial dysentery.

Shigellosis is a significant cause of morbidity and mortality worldwide, most notably amongst children. Moreover, there is a global increase in the occurrence of multidrug-resistant isolates, including the epidemic and pandemic Shigella dysenteriae type 1 strain. We developed a bioluminescent reporter phage assay to facilitate detection and simultaneously determine antibiotic susceptibility. A Shigella flexneri phage (Shfl25875) was isolated from environmental wastewater and characterized by DNA sequencing. Shfl25875 is T4-like, harbors a 169,062-bp genome, and grows on most (28/29) S. flexneri strains and all 12 S. dysenteriae type 1 strains tested. The genes encoding bacterial luciferase were integrated into the Shfl25875 genome to create a "light-tagged" phage capable of transducing a bioluminescent phenotype to infected cells. Shfl25875::luxAB rapidly detects cultured isolates with high sensitivity. Specificity experiments indicate that the reporter does not respond to Shigella boydii, non-type 1 S. dysenteriae strains, and most non-Shigella Enterobacteriaceae. Shfl25875::luxAB generates ampicillin and ciprofloxacin susceptibility profiles that are similar to the standard Clinical and Laboratory Standards Institute (CLSI) growth microdilution method, but in a significantly shorter time. In addition, the reporter phage detects Shigella in mock-infected stool. This new reporter phage shows promise as a tool for the detection of cultured isolates or complex clinical samples.

Molecular characterization of serologically atypical provisional serovars of Shigella isolates from Kolkata, India.

During 2000-2004, 13 Shigella strains that were untypable by commercially available antisera were isolated from children <5 years of age with acute diarrhoea in Kolkata. These strains were subsequently identified as Shigella dysenteriae provisional serovar 204/96 (n = 3), Shigella dysenteriae provisional serovar E23507 (n = 1), Shigella dysenteriae provisional serovar I9809-73 (n = 1), Shigella dysenteriae provisional serovar 93-119 (n = 1), Shigella flexneri provisional serovar 88-893 (n = 6) and Shigella boydii provisional serovar E16553 (n = 1). In this study, characterization of those provisional serovars of Shigella was performed with respect to their antimicrobial resistance, plasmids, virulence genes and PFGE profiles. The drug resistant strains (n = 10) of Shigella identified in this study possessed various antibiotic resistance genetic markers like catA (for chloramphenicol resistance); tetA and tetB (for tetracycline resistance); dfrA1 and sul2 (for co-trimoxazole resistance); aadA1, strA and strB (for streptomycin resistance) and blaOXA-1 (for ampicillin resistance). Class 1 and/or class 2 integrons were present in eight resistant strains. Three study strains were pan-susceptible. A single mutation in the gyrA gene (serine to leucine at codon 83) was present in four quinolone resistant strains. The virulence gene ipaH (invasion plasmid antigen H) was uniformly present in all strains in this study, but the stx (Shiga toxin) and set1 (Shigella enterotoxin 1) genes were absent. Other virulence genes like ial (invasion associated locus) and sen (Shigella enterotoxin 2) were occasionally present. A large plasmid of 212 kb and of incompatibility type IncFIIA was present in the majority of the strains (n = 10) and diversity was noticed in the smaller plasmid profiles of these strains even within the same provisional serovars. PFGE profile analysis showed the presence of multiple unrelated clones among the isolates of provisional Shigella serovars. To the best of our knowledge, this is the first report on the phenotypic and molecular characterization of provisional serovars of Shigella isolates from Kolkata, India.

Genomic analysis of the emergence of 20th century epidemic dysentery.

BACKGROUND: Shigella dysenteriae type 1 (Sd1) causes recurrent epidemics of dysentery associated with high mortality in many regions of the world. Sd1 infects humans at very low infectious doses (10 CFU), and treatment is complicated by the rapid emergence of antibiotic resistant Sd1 strains. Sd1 is only detected in the context of human infections, and the circumstances under which epidemics emerge and regress remain unknown. RESULTS: Phylogenomic analyses of 56 isolates collected worldwide over the past 60 years indicate that the Sd1 clone responsible for the recent pandemics emerged at the turn of the 20th century, and that the two world wars likely played a pivotal role for its dissemination. Several lineages remain ubiquitous and their phylogeny indicates several recent intercontinental transfers. Our comparative genomics analysis reveals that isolates responsible for separate outbreaks, though closely related to one another, have independently accumulated antibiotic resistance genes, suggesting that there is little or no selection to retain these genes in-between outbreaks. The genomes appear to be subjected to genetic drift that affects a number of functions currently used by diagnostic tools to identify Sd1, which could lead to the potential failure of such tools. CONCLUSIONS: Taken together, the Sd1 population structure and pattern of evolution suggest a recent emergence and a possible human carrier state that could play an important role in the epidemic pattern of infections of this human-specific pathogen. This analysis highlights the important role of whole-genome sequencing in studying pathogens for which epidemiological or laboratory investigations are particularly challenging.

In vitro selection of a DNA aptamer targeted against Shigella dysenteriae.

To identify DNA aptamers demonstrating binding specificity for Shigella dysenteriae, a whole-bacterium Systemic Evolution of Ligands by Exponential enrichment (SELEX) method was applied to a combinatorial library of single-stranded DNA (ssDNA) molecules. After several rounds of selection using S. dysenteriae as the target, the highly enriched oligonucleotide pool was sequenced and then grouped into different families based on primary sequence homologies and similarities in the secondary structures. Aptamer S 1, which showed particularly high binding affinity in preliminary studies, was chosen for further characterisation. This aptamer displayed a dissociation constant (Kd value) of 23.47 ± 2.48 nM. Binding assays to assess the specificity of aptamer S 1 showed high binding affinity for S. dysenteriae and low apparent binding affinity for other bacteria. The ssDNA aptamers generated may serve as a new type of molecular probe for microbial pathogens, as it has the potential to overcome the tedious isolation and purification requirements for complex targets.

Haemolytic-uraemic syndrome as a sequela of diarrhoeal disease.

Haemolytic-uraemic syndrome (HUS) is a serious sequela of diarrhoea and results in a high mortality rate. This systematic review aimed at estimating the proportion of HUS cases that are linked to prior infection due to Shiga toxin-producing Escherichia coli (STEC) or Shigella dysenteriae type 1. A systematic review of the existing literature was done to identify cohort and case-control studies that examined the relationship between STEC and S. dysenteriae type 1 and HUS. After screening 2,516 articles, 11 studies were found that met the inclusion/exclusion criteria. Findings of case-control studies suggest that 60.8% of the HUS cases may be attributable to a previous infection with STEC. In cohort studies, 7.8% of participants with STEC and 8% of participants with S. dysenteriae type 1 developed HUS during follow-up. HUS is linked to diarrhoea due to both STEC and S. dysenteriae type 1. Thus, preventing infections caused by both pathogens is critical for the prevention and control of HUS, especially in areas where timely and effective treatment is not available.