Enterobacterial repetitive intergenic consensus (ERIC)-PCR analysis as a reliable evidence for suspected Shigella spp. outbreaks

Abstract Background Shigellosis remains a serious public health problem and an important cause of morbidity and mortality worldwide. The aim of this study was to characterize fliC and the genetic relatedness of Shigella spp. isolated during a one-year period from children in a suspected outbreak in Tehran, Iran. Methods and results Fifty Shigella spp. were isolated from 3779 stool samples of children with diarrhea (prevalence rate: 1.32%). Among the isolates, 92% were characterized as Shigella sonnei, while 6% and 2% were identified as S. flexneri and S. boydii, respectively. S. dysenteriae was not recovered from the patients. All isolates were negative for fliC except for Shigella standard strains. The enterobacterial repetitive intergenic consensus-polymerase chain reaction (ERIC-PCR) profiles allowed differentiating the 50 isolates into 5 ERIC types, which were grouped into five clusters (ET1-ET5). Computer-assisted clustering of the strains showed a high degree of similarity among the isolates. Conclusion In conclusion, given the clonal correlation of the Shigella strains isolated in this study and the lack of fliC among them, we propose that probably a single or limited fliC-defected Shigella clone spread and caused the outbreak.

Relationship among Shigella spp. and enteroinvasive Escherichia coli (EIEC) and their differentiation

Shigellosis produces inflammatory reactions and ulceration on the intestinal epithelium followed by bloody or mucoid diarrhea. It is caused by enteroinvasive E. coli (EIEC) as well as any species of the genus Shigella, namely, S. dysenteriae, S. flexneri, S. boydii, and S. sonnei. This current species designation of Shigella does not specify genetic similarity. Shigella spp. could be easily differentiated from E. coli, but difficulties observed for the EIEC-Shigella differentiation as both show similar biochemical traits and can cause dysentery using the same mode of invasion. Sequencing of multiple housekeeping genes indicates that Shigella has derived on several different occasions via acquisition of the transferable forms of ancestral virulence plasmids within commensal E. coli and form a Shigella-EIEC pathovar. EIEC showed lower expression of virulence genes compared to Shigella, hence EIEC produce less severe disease than Shigella spp. Conventional microbiological techniques often lead to confusing results concerning the discrimination between EIEC and Shigella spp. The lactose permease gene (lacY) is present in all E. coli strains but absent in Shigella spp., whereas β-glucuronidase gene (uidA) is present in both E. coli and Shigella spp. Thus uidA gene and lacY gene based duplex real-time PCR assay could be used for easy identification and differentiation of Shigella spp. from E. coli and in particular EIEC.

Shigella in Brazilian children with acute diarrhoea: prevalence, antimicrobial resistance and virulence genes

Diarrhoeal disease is still considered a major cause of morbidity and mortality among children. Among diarrhoeagenic agents, Shigella should be highlighted due to its prevalence and the severity of the associated disease. Here, we assessed Shigella prevalence, drug susceptibility and virulence factors. Faeces from 157 children with diarrhoea who sought treatment at the Children's Hospital João Paulo II, a reference children´s hospital in Belo Horizonte, state of Minas Gerais, Brazil, were cultured and drug susceptibility of the Shigella isolates was determined by the disk diffusion technique. Shigella virulence markers were identified by polymerase chain reaction. The bacterium was recovered from 10.8% of the children (88.2% Shigella sonnei). The ipaH, iuc, sen and ial genes were detected in strains isolated from all shigellosis patients; set1A was only detected in Shigella flexneri. Additionally, patients were infected by Shigella strains of different ial, sat, sen and set1A genotypes. Compared to previous studies, we observed a marked shift in the distribution of species from S. flexneri to S. sonnei and high rates of trimethoprim/sulfamethoxazole resistance.

Antimicrobial Resistance of Shigella spp. isolated in the State of Pará, Brazil / Resistência Antimicrobiana de Shigella spp. isoladas no Estado do Pará

INTRODUCTION: Shigella spp. are Gram-negative, nonsporulating, rod-shaped bacteria that belong to the family Enterobacteriaceae and are responsible for shigellosis or bacillary dysentery, an important cause of worldwide morbidity and mortality. METHODS: We studied the antibiotic resistance profiles of 122 Shigella spp. strains (81 S. flexneri, 41 S. sonnei, 1 S. boydii) isolated from patients (female and male from 0 to 80 years of age) presenting diarrhea in different districts of the State of Pará, in the North of Brazil. The antibiotic resistance of the strains, isolated from human fecal samples, was determined by the diffusion disk method and by using the VITEK-2 system. RESULTS: The highest resistance rate found was the resistance rate to tetracycline (93.8 percent), followed by the resistance rate to chloramphenicol (63.9 percent) and to trimethoprim/sulfamethoxazole (63.1 percent). Resistance to at least three drugs was more common among S. flexneri than S. sonnei (39.5 percent vs. 10 percent). Six (4.9 percent) strains were susceptible to all the antibiotics tested. All strains were susceptible to cefotaxime, ceftazidime, ciprofloxacin, nalidixic acid and nitrofurantoin. CONCLUSIONS: High rates of multidrug resistance in Shigella spp. are a serious public health concern in Brazil. It is extremely important to continuously monitor the antimicrobial resistances of Shigella spp. for effective therapy and control measures against shigellosis.

INTRODUÇÃO: Shigella spp. são bactérias Gram-negativas, não esporuladas, em forma de bastonete, pertencentes a família Enterobacteriaceae responsáveis pela shigelose ou disenteria bacilar, uma importante causa de mortalidade e morbidade mundial. MÉTODOS: Foi estudado o perfil de resistência a antimicrobianos de 122 amostras de Shigella spp. (81 S. flexneri, 41 sonnei, 1 S. boydii) isoladas de pacientes (sexo feminino e masculino com faixa etária de 0 a 80 anos) com distúrbios gastrointestinais em diferentes municípios no Estado do Pará, Brasil. A resistência antimicrobiana das amostras isoladas de coprocultura, foi determinada pelo método de difusão em disco e pelo sistema Vitek II. RESULTADOS: A maior resistência foi observada em relação à tetraciclina (93,8 por cento), seguida de cloranfenicol (63,9 por cento), e trimetoprimsulfametoxazol (63,1 por cento). Multirresistência a pelo menos três antimicrobianos foi mais comum em S. flexneri comparada a S. sonnei (39,5 por cento vs. 10 por cento). Seis (4,9 por cento) amostras foram sensíveis a todos antimicrobianos testados. Todas as amostras apresentaram sensibilidade a cefotaxima, ceftazidima, ciprofloxacina, ácido nalidixico e nitrofurantoína. CONCLUSÕES: As altas taxas de multirresistência de Shigella spp. são um sério problema de saúde pública no Brasil. Sendo assim, torna-se extremamente importante um monitoramento contínuo da resistência antimicrobiana de Shigella spp. para uma terapia efetiva e medidas de controle contra shigelose.

Frequency and antimicrobial susceptibility of Shigella species isolated in Children Medical Center Hospital, Tehran, Iran, 2001-2006

Appropriate antimicrobial treatment of shigellosis depends on identifying its changing resistance pattern over time. We evaluated 15,255 stool culture submitted from July 2001 to June 2006 to the Laboratory of Children Medical Center Hospital. Specimen culture, bacterial identification, and disk diffusion susceptibility testing were performed according to National Committee for Clinical Laboratory Standards guidelines. From 15,255 stool samples, 682 (4.5 percent) were positive for Shigella species. The most common species of Shigella were S. flexneri (48 percent) and S. sonnei (45 percent); other results were S. dysenteriae (5 percent) and S. boydii (2 percent). The rate of Sensitivity to ceftriaxone (95 percent), ceftizoxime (94 percent), and nalidixic acid (84 percent) were among our isolates. Resistance to co-trimoxazole and ampicillin was 87 percent and 86 percent, respectively. S. flexneri was more multiresistant than other species (47.9 percent). Our isolates are overall most sensitive to ceftriaxone, ceftazidime, and nalidixic acid (> 84 percent). They were most resistant to co-trimoxazole and ampicillin (> 86 percent). Because resistance varies according to specific location, continuous local monitoring of resistance patterns is necessary for the appropriate selection of empirical antimicrobial therapy.

Characterization of Shigella spp. by antimicrobial resistance and PCR detection of ipa genes in an infantile population from Porto Velho (Western Amazon region), Brazil

The incidence of Shigella spp. was assessed in 877 infants from the public hospital in Rondônia (Western Amazon region, Brazil) where Shigella represents the fourth cause of diarrhea. Twenty-five isolates were identified: 18 were Shigella flexneri, three Shigella sonnei, three Shigella boydii and one Shigella dysenteriae. With the exception of S. dysenteriae, all Shigella spp. isolated from children with diarrhea acquired multiple antibiotic resistances. PCR detection of ipa virulence genes and invasion assays of bloody diarrhea and fever (colitis) were compared among 25 patients testing positive for Shigella. The ipaH and ipaBCD genes were detected in almost all isolates and, unsurprisingly, all Shigella isolates associated with colitis were able to invade HeLa cells. This work alerts for multiple antibiotic resistant Shigella in the region and characterizes presence of ipa virulence genes and invasion phenotypesin dysenteric shigellosis.

A food borne outbreak of gastroenteritis due to shigella and possibly salmonella in a school.

On August 5, 2005, a private hospital reported a large number of students with gastrointestinal illness from the same school in Bangkok, Thailand. The Bureau of Epidemiology along with the Bangkok Metropolitan Administration investigated this outbreak, to determine risk factors, identify the source of infection and possible causative organism, and recommend prevention and control strategies. A case was defined as a person who was studying or working at School A and who developed abdominal pain, diarrhea, nausea or vomiting during the five-day period of August 4 to 8, 2005. A descriptive study was carried out for active case-finding, medical records review, and case interviews. We conducted the retrospective cohort study among third and fourth grade students. Stool samples were collected and tested at the Thai National Institute of Health and at private hospital laboratories. The overall attack rate was 37%. Main symptoms were diarrhea, fever, headache, abdominal pain, vomiting, and nausea. The highest attack rate (63%) was among fourth-grade students. Based on food-history data collected from ill and well students, a multiple logistic regression analysis showed that a mixed chicken and rice dish served for lunch on August 4 was associated with illness (OR 3.28, 95% CI 1.46-7.36). Among stools samples from 103 cases, Shigella group D was found in 18 cases, Salmonella group C in 5 cases, and Salmonella group E in 2 cases. This food borne outbreak of gastroenteritis was most likely caused by Shigella spp although the possibility of mixed contamination with Shigella and Salmonella spp cannot be ruled out. Food borne outbreaks such as this can be prevented through simple and effective hygienic measures.

Frequency of serovars and antimicrobial resistance in Shigella spp. from Brazil

A total of 296 Shigella spp. were received from State Public Health Laboratories, during the period from 1999 to 2004, by National Reference Laboratory for Cholera and Enteric Diseases (NRLCED) - IOC/Fiocruz, Rio de Janeiro, Brazil. The frequency of Shigella spp. was: S. flexneri (52.7 percent), S. sonnei (44.2 percent), S. boydii (2.3 percent), and S. dysenteriae (0.6 percent). The most frequent S. flexneri serovars were 2a and 1b. The highest incidence rates of Shigella isolation were observed in the Southeast (39 percent) and Northeast (34 percent) regions and the lowest rate in the South (3 percent) of Brazil. Strains were further analyzed for antimicrobial susceptibility by disk diffusion method as part of a surveillance program on antimicrobial resistance. The highest rates of antimicrobial resistance were to trimethoprim-sulfamethozaxole (90 percent), tetracycline (88 percent), ampicillin (56 percent), and chloramphenicol (35 percent). The patterns of antimicrobial resistance among Shigella isolates pose a major difficulty in the determination of an appropriate drug for shigellosis treatment. Continuous monitoring of antimicrobial susceptibilities of Shigella spp. through a surveillance system is thus essential for effective therapy and control measures against shigellosis.

Childhood shigellosis at King Chulalongkorn Memorial Hospital, Bangkok, Thailand: a 5-year review (1996-2000).

The objective of this study was to evaluate the demographic data and clinical presentation of childhood shigellosis, and to study the microbiological data and antimicrobial susceptibilities of Shigella spp. Nine thousand nine hundred fourteen stool culture specimens from children aged 0-15 years who were treated at King Chulalongkorn Memorial Hospital, Bangkok, Thailand, between 1996 and 2000 were retrospectively reviewed. Data were collected from microbiological records and medical charts of childhood shigellosis in terms of demographic data, symptoms, signs, and complications of the patients, and the species and antimicrobial susceptibilities of the organisms. The data were analyzed in terms of means, ranges, and percentages. Of 1,523 children whose stool cultures were positive for pathogenic bacteria, 80 (5.3%) were infected with Shigella spp; 34 females and 46 males. The age distribution ranged from 1 day to 13 years with a mean age of 3.6 years. Common clinical presentations included diarrhea (96.6%), fever (77.6%) and vomiting (44.8%); seizures were the most common complication found (27.6%). Watery and mucous were the most common characteristics of stools. The major Shigella spp found was S. sonnei (62.8%), which was susceptible to co-trimoxazole, ampicillin, cefazolin and ciprofloxacin in 2.3, 84.1, 100 and 100%, respectively. A short course of quinolones or oral cephalosporins should be recommended for the treatment of childhood shigellosis in areas with low susceptibility rates to co-trimoxazole and ampicillin.

Shigella and the fluorinated quinolones.

Between July 1996 to June 1997 a total of 531 Stool samples from cases of acute diarrhoea, dysentry, colitis admitted to Kasturba Medical College Hospital, Manipal were investigated. Shigella were isolated from 29 samples (5.46%). Out of them, Shigella flexneri were 16 (55.17%), Shigella boydii 8 (27.58%), Shigella dysentriae 3 (10.34%) and Shigella sonnei 2 (6.89%). Five strains of Shigella flexneri and one strain of Shigella dysentriae were found to show resistance to Nalidixic acid and the newer fluoroquinolones viz., Ciprofloxacin, Norfloxacin and Ofloxacin. The in vitro 'R' factor transfer study showed that the resistance was not transferable.

Susceptivilidad in vitro de azitromicina frente a cepas de Shigella sp. provenientes deniños con diarrea disentéria / In vitro susceptibility of azitromicine to shigella sp strain in clhildren with dysenteric diarrhea

Shigela sp., probablemente representa el principal agente etiológico de la diarrea enteroinvasiva en nuestro paías y se caracteriza por exhibir un elevado perfil de resistencia a diferentes antimicrobianos. la azitromicina, constituye na nueva clase de antibióticos denominada azálido, con espectro de actividad ampliada contra Gram negativos y Gram positivos. Nosotros estudiamos la sensibilidad in vitro, de la azitromicina frente a las cepas de Shigella sp. aisladas de niños con diarrea enteroinvasiva en nuestro país. 150 cepas de Shigella sp fueron estudiadas mediante la técnica de difusión con discos (Kirby-Bauer), 125 (83 porciento9 fueron S. fexneri, 20 (14) S sonnei y 5 (3 porciento) S. dysenteriae. El 100 porciento de las cpas de Shigelli sp. estudiadas demostraron ser sensibles in vitro a la azitroicina. se concluye en la necesidad de realizar estudios clínicos prospectivos a fin de determinar el rol de la azitromicina en niños con shigellosis.

A study on transferable R-plasmids among Shigella species at Lucknow.

A total of 500 Shigella strains were included in the present study out of which 233 were isolated at National Shigella Centre of Postgraduate Department of Microbiology, K.G. Medical College, Lucknow during last 5 years and 267 strains were received from different parts of the country. Predominent serotypes were Sheigella flexneri 292 (48.4%) followed by Shigella dysenteriae 153 (30.6%), Shigella boydii 32 (6.4%) and Shigella sonnei 23 (4.6%). All the strains were sensitive to Nalidix acid, Gentamycin, Kanamycin, Neomycin and Furazolidine. Varying degree of resistance was shown to Streptomycin, Chloramphenicol, Tetracycline and Ampicillin. R. plasmid extraction was done and transfer was studied on E. Coli K-12.

Estudo da resistencia antimicrobiana in vitro das coproculturas positivas para Shigella sp / Study of in vitro antimicrobial resistance of positive coprocultures to Shigella sp

Introduçäo gastroenterite causada pela Shigella sp (shigelose) representa importante causa de morbimortalidade, especialmente em crianças abaixo de 2 anos de idade. Sabe-se que o tratamento antimicrobiano adequado pode melhorar o quadro clínico e diminuir a disseminaçäo da doença. Têm sido descritos níveis crescentes de resistência aos antimicrobianos comumente usados para o tratamento, e a sensibilidade da Shigella näo foi recentemente avaliada em nosso meio. Objetivo: avaliar a resitência antimicrobiana das espécies de Shigella em nosso meio. Método: Delineou-se um estudo retrospectivo, utilizando-se 106 coproculturas de pacientes, hospitalizados ou näo, que tenham procurado atendimento no HCPA ou no HPV durante o período de 1994 a 1996 com diagnóstico de shigelose firmado por coprocultura...

Shiguelose: diagnóstico e tratamento / Shiguellosis: diagnosis and therapy

É uma doença diarréica aguda infecciosa mundialmente distribuída, causada por um bacilo gram-negativo da família das enterobactérias, transmitido por contato inter-humano, por água e alimentos contaminados, causando diarréia mucopiossanguinolenta, associada à febre, cólicas abdominais, urgência e tenesmo retal. O diagnóstico é baseado na história clínica e exame físico, sendo confirmado principalmente pela coprocultura e, mais raramente, pela hemocultura. Outros exames inespecíficos podem ser usados, como o hemograma e a análise do mucofecal. A sorologia e o PCR podem ser úteis, porém nem sempre facilmente disponíveis. O tratamento visa a correçÝo dos distúrbios hidroeletrolíticos já que a shiguelose costuma ser autolimitada, sendo porém, eventualmente necessário o uso de antimicrobianos. Quando a shiguela isolada se mostra sensível à ampicilina, esta droga tem sido utilizada como primeira escolha; entretanto, nÝo se isolando o agente causal, o sulfametoxazoltrimethopim pode ser indicado. atualmente, os melhores resultados (e com baixos ídices de resistência) têm sido obtidos com o uso das quinolonas, devendo, porém, ser usadas com critério devido ao aparecimento de cepas multirresistentes.

Serogroup prevalence of Shigellae in Bombay.

Prevalence of Shigellae serotypes in Bombay was studied from June 1988 to May 1991. A total of 2758 faecal specimens were collected from paediatric patients (< 12 yrs) with acute gastroenteritis. A total of 90 Shigella were isolated giving the isolation rate of 3.2%. Shigella flexneri was the predominant serogroup (73.3%) followed by Shigella dysenteriae (16.6%). All the isolates were sensitive to nalidixic acid. Eighty percent of the Shigellae were multidrug resistant. Present data were compared with the study carried out during the period of 1983-87 from the same institute. A change in the serogroup prevalence was noted wherein Shigella flexneri dominated over Shigella dysenteriae since 1985. Increase in resistance to ampicillin and cotrimoxazole was seen in Shigella flexneri strains as compared to previous years.

Shigella isolates in stool.

A total of 447 Shigella strains were isolated from stool samples during 1989-1991. Of these 270 (60%) were from children. Among the different species and serotypes Sh. flexneri 60 (13.4%) and Sh. sonnei Phage 139 (65%) were the most frequently isolated strains. 154 (34.4%) strains were resistant to three and 179 (40%) to more than three antibiotics. Some strains of Shigella were found to be resistant to furazolidine and neomycin.

Prevalent serogroups and antimicrobial susceptibility of Shigella strains in Metro Manila, 1982-1988.

From 1982 to 1988, Shigella was detected in 4% of stool specimens cultured at The Research Institute for Tropical Medicine (RITM), Manila. S. flexneri was the most prevalent serogroup with 2a, 1b and 1a as the predominant serotypes. Isolation was most frequent during the July to December period. Drug resistance was most often observed in S. flexneri with serotype 2a exhibiting resistance to the combination of ampicillin, chloramphenicol, tetracycline and streptomycin. More multiple-drug than single-drug resistant strains were observed for the last four years. Ampicillin is still the most frequently used drug for most forms of shigellosis in the Philippines. However, the demonstration of increasing resistance among the isolates studied in RITM underscores the need to monitor antibiotic susceptibility studies and document the emergence of resistant strains in the entire country.