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

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.(AU)

Isolamento e determinação de sensibilidade e resistência a antimicrobianos de cepas bacterianas presentes na cloaca de Trachemys scripta elegans (Wied, 1839) criadas em cativeiro em Petrolina, PE / Isolation and determination of antimicrobials sensitivity and resistance of bacterial strains present in the cloaca Trachemys scripta elegans (Wied, 1839) raised in captivity in Petrolina/PE, Brazil

Este estudo isolou e determinou o perfil de sensibilidade e de resistência a antimicrobianos de cepas bacterianas isoladas da cloaca de Trachemys scripta elegans (T. s. elegans) criadas em cativeiro. Após 120 dias de adaptação, amostras de swab cloacal obtidas de 20 animais adultos foram cultivadas e, após a identificação dos patógenos através de testes bioquímicos, submetidas ao teste de suscetibilidade a nove antimicrobianos. Enterobacter aerogenes (85%); Shigella spp. (10%) e Edwadsiella spp. (5%) foram isolados e identificados. Os isolados de E. aerogenes foram sensíveis à gentamicina (86%), enrofloxacina (79%), estreptomicina (50%), sulfazotrim (36%) e ampicilina (29%) e resistentes a penicilina (100%), eritromicina (93%), cefalexina (86%) ampicilina (71%) e sulfazotrim (64%). Isolados de Shigella spp. apresentaram sensibilidade à gentamicina (100%), enrofloxacina (50%), doxicilina (50%), estreptomicina (50%), ampicilina (50%), penicilina (50%) e sulfazotrim (50%) e resistência a doxicilina (50%), estreptomicina (50%), ampicilina (50%), penicilina (100%), cefalexina (50%) e sulfazotrim (50%), enquanto que os de Edwardsiella spp. foram sensíveis apenas à gentamicina (100%) e altamente resistentes (100%) aos demais antimicrobianos. Os resultados sugerem a participação de T. s. elegans na cadeia epidemiológica, como reservatório de patógenos importantes, como E. aerogenes, Shigella spp. e Edwardisiella spp., tornando importante a adoção de medidas preventivas pelo risco zoonótico que apresentam e corretas de tratamento e de controle em cativeiros e domicílios, assim como de estudos que enfoquem as características de sensibilidade e de resistência antimicrobiana dos isolados cloacais, pois a multirresistência a drogas pode ser transmitida aos humanos e comprometer o tratamento de indivíduos com doenças graves.(AU)

This study isolated and determined the profile of susceptibility and antimicrobials resistance of bacterial strains isolated from the cloaca Trachemys scripta elegans (T. s. elegans) raised in captivity. After 120 days of adaptation, cloacal swab samples obtained from 20 adults animals were grown and, after the pathogens identification through biochemical tests, submitted to the test of susceptibility to nine antimicrobials. Enterobacter aerogenes (85%); Shigella spp. (10%) and Edwadsiella spp. (5%) were isolated and identified. Isolates from E. aerogenes were sensitive to gentamicin (86%), enrofloxacin (79%), streptomycin (50%), sulfazotrim (36%) and ampicillin (29%) and resistant to penicillin (100%), erythromycin (93%), cephalexin (86%), ampicillin (71%) and sulfazotrim (64%). Isolates from Shigella spp. showed sensitivity to gentamicin (100%), enrofloxacin (50%), doxycycline (50%), streptomycin (50%), ampicillin (50%), penicillin (50%) and sulfazotrim (50%) and resistance to doxycycline (50 %), streptomycin (50%), ampicillin (50%), penicillin (100%), cephalexin (50%) and sulfazotrim (50%), while the Edwardsiella spp. were sensitive only to gentamicin (100%) and were highly resistant (100%) to other antibiotics. The results suggest the participation of T. s. elegans in the epidemiological chain, as reservoir of important pathogens, such as E. aerogenes, Shigella spp. and Edwardisiella spp., making it important to adopt preventive measures for zoonotic risk that present and correct treatment and control in captivity and households, as well as studies that address the sensitivity characteristics and antimicrobial resistance of isolates from cloaca, as it multidrug resistance to drugs can be transmitted to humans and compromise the treatment of patients with serious diseases.(AU)

Isolamento e determinação de sensibilidade e resistência a antimicrobianos de cepas bacterianas presentes na cloaca de Trachemys scripta elegans (Wied, 1839) criadas em cativeiro em Petrolina, PE / Isolation and determination of antimicrobials sensitivity and resistance of bacterial strains present in the cloaca Trachemys scripta elegans (Wied, 1839) raised in captivity in Petrolina/PE, Brazil

Este estudo isolou e determinou o perfil de sensibilidade e de resistência a antimicrobianos de cepas bacterianas isoladas da cloaca de Trachemys scripta elegans (T. s. elegans) criadas em cativeiro. Após 120 dias de adaptação, amostras de swab cloacal obtidas de 20 animais adultos foram cultivadas e, após a identificação dos patógenos através de testes bioquímicos, submetidas ao teste de suscetibilidade a nove antimicrobianos. Enterobacter aerogenes (85%); Shigella spp. (10%) e Edwadsiella spp. (5%) foram isolados e identificados. Os isolados de E. aerogenes foram sensíveis à gentamicina (86%), enrofloxacina (79%), estreptomicina (50%), sulfazotrim (36%) e ampicilina (29%) e resistentes a penicilina (100%), eritromicina (93%), cefalexina (86%) ampicilina (71%) e sulfazotrim (64%). Isolados de Shigella spp. apresentaram sensibilidade à gentamicina (100%), enrofloxacina (50%), doxicilina (50%), estreptomicina (50%), ampicilina (50%), penicilina (50%) e sulfazotrim (50%) e resistência a doxicilina (50%), estreptomicina (50%), ampicilina (50%), penicilina (100%), cefalexina (50%) e sulfazotrim (50%), enquanto que os de Edwardsiella spp. foram sensíveis apenas à gentamicina (100%) e altamente resistentes (100%) aos demais antimicrobianos. Os resultados sugerem a participação de T. s. elegans na cadeia epidemiológica, como reservatório de patógenos importantes, como E. aerogenes, Shigella spp. e Edwardisiella spp., tornando importante a adoção de medidas preventivas pelo risco zoonótico que apresentam e corretas de tratamento e de controle em cativeiros e domicílios, assim como de estudos que enfoquem as características de sensibilidade e de resistência antimicrobiana dos isolados cloacais, pois a multirresistência a drogas pode ser transmitida aos humanos e comprometer o tratamento de indivíduos com doenças graves.(AU)

This study isolated and determined the profile of susceptibility and antimicrobials resistance of bacterial strains isolated from the cloaca Trachemys scripta elegans (T. s. elegans) raised in captivity. After 120 days of adaptation, cloacal swab samples obtained from 20 adults animals were grown and, after the pathogens identification through biochemical tests, submitted to the test of susceptibility to nine antimicrobials. Enterobacter aerogenes (85%); Shigella spp. (10%) and Edwadsiella spp. (5%) were isolated and identified. Isolates from E. aerogenes were sensitive to gentamicin (86%), enrofloxacin (79%), streptomycin (50%), sulfazotrim (36%) and ampicillin (29%) and resistant to penicillin (100%), erythromycin (93%), cephalexin (86%), ampicillin (71%) and sulfazotrim (64%). Isolates from Shigella spp. showed sensitivity to gentamicin (100%), enrofloxacin (50%), doxycycline (50%), streptomycin (50%), ampicillin (50%), penicillin (50%) and sulfazotrim (50%) and resistance to doxycycline (50 %), streptomycin (50%), ampicillin (50%), penicillin (100%), cephalexin (50%) and sulfazotrim (50%), while the Edwardsiella spp. were sensitive only to gentamicin (100%) and were highly resistant (100%) to other antibiotics. The results suggest the participation of T. s. elegans in the epidemiological chain, as reservoir of important pathogens, such as E. aerogenes, Shigella spp. and Edwardisiella spp., making it important to adopt preventive measures for zoonotic risk that present and correct treatment and control in captivity and households, as well as studies that address the sensitivity characteristics and antimicrobial resistance of isolates from cloaca, as it multidrug resistance to drugs can be transmitted to humans and compromise the treatment of patients with serious diseases.(AU)

CTX-M extended-spectrum -lactamase-producing Klebsiella spp, Salmonella spp, Shigella spp and Escherichia coli isolates in Iranian hospitals

This study was conducted in Iran in order to assess the distribution of CTX-M type ESBLs producing Enterobacteriaceae. From January 2012 to December 2013, totally 198 E. coli, 139 Klebsiella spp, 54 Salmonella spp and 52 Shigella spp from seven hospitals of six provinces in Iran were screened for resistance to extended-spectrum cephalosporins. After identification and susceptibility testing, isolates presenting multiple-drug resistance (MDR) were evaluated for ESBL production by the disk combination method and by Etest using (cefotaxime and cefotaxime plus clavulanic acid). All isolates were also screened for bla CTX-M using conventional PCR. A total of 42.92%, 33.81%, 14.81% and 7.69% of the E. coli, Klebsiella spp, Salmonella spp and Shigella spp isolates were MDR, respectively. The presence of CTX-M enzyme among ESBL-producing isolates was 85.18%, 77.7%, 50%, and 66.7%, in E. coli, Klebsiella spp, Salmonella spp and Shigella spp respectively. The overall presence of CTX-M genes in Enterobacteriaceae was 15.4% and among the resistant isolates was 47.6%. This study indicated that resistance to -lactams mediated by CTX-M enzymes in Iran had similar pattern as in other parts of the world. In order to control the spread of resistance, comprehensive studies and programs are needed.(AU)

The importance of integrons for development and propagation of resistance in Shigella: the case of Latin America

In Latin America, the disease burden of shigellosis is found to coexist with the rapid and rampant spread of resistance to commonly used antibiotics. The molecular basis of antibiotic resistance lies within genetic elements such as plasmids, transposons, integrons, genomic islands, etc., which are found in the bacterial genome. Integrons are known to acquire, exchange, and express genes within gene cassettes and it is hypothesized that they play a significant role in the transmission of multidrug resistance genes in several Gram-negative bacteria including Shigella. A few studies have described antibiotic resistance genes and integrons among multidrug resistant Shigella isolates found in Latin America. For example, in Brazil, Bolivia, Chile, Costa Rica and Peru, class 1 and class 2 integrons have been detected among multidrug resistant strains of Shigella; this phenomenon is more frequently observed in S. flexneri isolates that are resistant to trimethoprim, sulfamethoxazole, streptomycin, ampicillin, chloramphenicol, and tetracycline. The gene cassette sul2, which is frequently detected in Shigella strains resistant to the sulfonamides, suggests that the sulfonamide-resistant phenotype can be explained by the presence of the sul2 genes independent of the integron class detected. It is to be noted that sul3 was negative in all isolates analyzed in these studies.The high frequency of sulfonamide (as encoded by sul2) and trimethoprim resistance is likely to be a result of the recurrent use of trimethoprim sulfamethoxazole as a popular regimen for the treatment of shigellosis. The observed resistance profiles of Shigella strains confirm that ampicillin and trimethoprim-sulfamethoxazole are ineffective as therapeutic options. In-depth information regarding antibiotic resistance mechanism in this pathogen is needed in order to develop suitable intervention strategies. There is a pressing need for regional and local antimicrobial resistance profiling of Shigella to be included as a part of the public health strategy.(AU)

Caracterização filogenética molecular e resistência antimicrobiana de escherichia coli isoladas de caprinos neonatos com diarreia / Molecular-phylogenetic characterization and antimicrobial resistance of Escherichia coli isolated from goats with diarrhea

Neonatal diarrhea determines significant changes in feed conversion, causing productivity loss in caprine herds. The antimicrobial resistance in bacteria is characterized as an important public healthissue; therefore, Escherichia coli may be characterized as an important pathogen due to expressingvirulence mechanisms responsible for significant clinical conditions in humans and animals. Thepresent study evaluated the presence of E. coli among 117 caprine fecal samples and analyzed theisolates for antimicrobial resistance. Suggestive colonies were submitted to biochemical screeningfollowed by genotypic group determination and phylogenetic analysis; further, the sampleswere submitted to antimicrobials susceptibility test. E. coli, Salmonella spp, Shigella sonnei andEnterobacter aerogenes were identified. E. coli isolates were phylogenetically classified as B2 (9/39),D (19/39), B1 (7/39) e A (4/29) groups. The analysis of the isolates also revealed the presence of K99(04/39) and Stx (02/39) virulence factors. Antimicrobial susceptibility test revealed sensitive isolatesto Chloramphenicol, Streptomycin, Amoxicillin and Ciprofloxacin, being all resistant to Lincomycin,Vancomycin and Penicillin. The results support the need of establishing restricted protocols forantimicrobial use, a fundamental procedure for health improvement in Brazilian caprine herds.

A diarreia neonatal determina alterações significativas na conversão alimentar, resultando na queda daprodutividade de caprinos. A resistência antimicrobiana em bactérias caracteriza-se como importanteproblema de saúde pública, assim, a Escherichia coli pode ser caracterizada como importantepatógeno por expressar mecanismos de virulência responsáveis por significativas afecções clínicasem seres humanos e animais. O presente estudo avaliou a presença de E. coli em 117 amostrasfecais de caprinos e analisou a resistência antimicrobiana dos isolados. Colônias sugestivas foramsubmetidas a testes bioquímicos, seguido de determinação do grupo genotípico e análise filogenética;posteriormente foram submetidas à prova de suscetibilidade a antimicrobianos. Foram identificadosisolados de E. coli, Salmonella spp, Shigella sonnei e Enterobacter aerogenes. Isolados de E. coliforam classificados filogeneticamente em B2 (9/39), D (19/39), B1 (7/39) e A (4/29). Os fatores devirulência identificados foram K99 (04/39) e Stx (02/39). À prova de suscetibilidade antimicrobiana,os isolados foram sensíveis a Cloranfenicol, Estreptomicina, Amoxicilina e Ciprofloxacina, sendotodos resistentes à Lincomicina, Vancomicina e Penicilina. Os resultados reforçam a necessidade deestabelecimento de protocolos criteriosos para o uso de antimicrobianos, medida fundamental para oaprimoramento da sanidade dos rebanhos caprinos brasileiros.

Caracterização filogenética molecular e resistência antimicrobiana de escherichia coli isoladas de caprinos neonatos com diarreia / Molecular-phylogenetic characterization and antimicrobial resistance of Escherichia coli isolated from goats with diarrhea

Neonatal diarrhea determines significant changes in feed conversion, causing productivity loss in caprine herds. The antimicrobial resistance in bacteria is characterized as an important public healthissue; therefore, Escherichia coli may be characterized as an important pathogen due to expressingvirulence mechanisms responsible for significant clinical conditions in humans and animals. Thepresent study evaluated the presence of E. coli among 117 caprine fecal samples and analyzed theisolates for antimicrobial resistance. Suggestive colonies were submitted to biochemical screeningfollowed by genotypic group determination and phylogenetic analysis; further, the sampleswere submitted to antimicrobials susceptibility test. E. coli, Salmonella spp, Shigella sonnei andEnterobacter aerogenes were identified. E. coli isolates were phylogenetically classified as B2 (9/39),D (19/39), B1 (7/39) e A (4/29) groups. The analysis of the isolates also revealed the presence of K99(04/39) and Stx (02/39) virulence factors. Antimicrobial susceptibility test revealed sensitive isolatesto Chloramphenicol, Streptomycin, Amoxicillin and Ciprofloxacin, being all resistant to Lincomycin,Vancomycin and Penicillin. The results support the need of establishing restricted protocols forantimicrobial use, a fundamental procedure for health improvement in Brazilian caprine herds.(AU)

A diarreia neonatal determina alterações significativas na conversão alimentar, resultando na queda daprodutividade de caprinos. A resistência antimicrobiana em bactérias caracteriza-se como importanteproblema de saúde pública, assim, a Escherichia coli pode ser caracterizada como importantepatógeno por expressar mecanismos de virulência responsáveis por significativas afecções clínicasem seres humanos e animais. O presente estudo avaliou a presença de E. coli em 117 amostrasfecais de caprinos e analisou a resistência antimicrobiana dos isolados. Colônias sugestivas foramsubmetidas a testes bioquímicos, seguido de determinação do grupo genotípico e análise filogenética;posteriormente foram submetidas à prova de suscetibilidade a antimicrobianos. Foram identificadosisolados de E. coli, Salmonella spp, Shigella sonnei e Enterobacter aerogenes. Isolados de E. coliforam classificados filogeneticamente em B2 (9/39), D (19/39), B1 (7/39) e A (4/29). Os fatores devirulência identificados foram K99 (04/39) e Stx (02/39). À prova de suscetibilidade antimicrobiana,os isolados foram sensíveis a Cloranfenicol, Estreptomicina, Amoxicilina e Ciprofloxacina, sendotodos resistentes à Lincomicina, Vancomicina e Penicilina. Os resultados reforçam a necessidade deestabelecimento de protocolos criteriosos para o uso de antimicrobianos, medida fundamental para oaprimoramento da sanidade dos rebanhos caprinos brasileiros.(AU)

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.

A novel multiplex PCR for the simultaneous detection of Salmonella enterica and Shigella species

Salmonella enterica and Shigella species are commonly associated with food and water borne infections leading to gastrointestinal diseases. The present work was undertaken to develop a sensitive and reliable PCR based detection system for simultaneous detection of Salmonella enterica and Shigella at species level. For this the conserved regions of specific genes namely ipaH1, ipaH, wbgZ, wzy and invA were targeted for detection of Shigella genus, S. flexneri, S. sonnei, S. boydii and Salmonella enterica respectively along with an internal amplification control (IAC). The results showed that twenty Salmonella and eleven Shigella spp., were accurately identified by the assay without showing non-specificity against closely related other Enterobacteriaceae organisms and also against other pathogens. Further evaluation of multiplex PCR was undertaken on 50 natural samples of chicken, eggs and poultry litter and results compared with conventional culture isolation and identification procedure. The multiplex PCR identified the presence of Salmonella and Shigella strains with a short pre-enrichment step of 5 h in peptone water and the same samples were processed by conventional procedures for comparison. Therefore, this reported multiplex PCR can serve as an alternative to the tedious time-consuming procedure of culture and identification in food safety laboratories.

Shigella sp: um problema de saúde pública

O aumento do consumo de refeições fora de casa está associado ao aumento das Doenças Veiculadas por Alimentos (DVA), que são provenientes do consumo de alimentos e água contaminados, principalmente, devido à falta de higiene no preparo dos alimentos e à ausência, em muitos casos, do tratamento da água usada no preparo destes alimentos. A shigelose é uma DVA que se caracteriza como uma doença inflamatória do trato gastrointestinal causada pela Shigella cujos sintomas e características se assemelham à salmonelose. O gênero Shigella possui quatro espécies: S. dysenteriae, S. flexneri, S. boydii e S. sonnei. Para a realização deste trabalho foi realizado levantamento de artigos publicados em revistas científicas sobre o tema, com o objetivo de verificar a incidência de Shigella em alimentos abordando suas características e danos provocados à saúde.(AU)

The increase of the consumption of meals out of house is associated to the increase of the foodborne ilnesses, which are originating from the consumption of foods and water contaminated mainly due to the lack of hygiene in the preparation of the foods and the absence in many cases of the treatment of water used in the preparation of these foods. The shigelose is a foodborne disease that is characterized as an inflammatory disease of the gastrointestinal treatment, caused by Shigella, and whose symptoms and characteristics resembles salmonelose. The type Shigella has four sorts: S. dysenteriae, S.flexneri, S. boydii and S. sonnei. To carry out this work was carried out survey of articles published in scientific journals on the subject, in order to verify the incidence of Shigella in food approaching its characteristics and damage to health.(AU)

Nota sobre infecção por Salmonella sp. e Shigella sp. em bugios-ruivos, Alouatta clamitans Cabrera, 1940 (Primates: atelidae) mantidos em cativeiro / Note about Salmonella sp. and Shigella sp. infections in southern brown howler monkeys, Alouatta clamitans Cabrera, 1940 (Primates: atelidae) on captivity

Amostras de fezes de bugios-ruivos cativos foram investigadas quanto a presença de Salmonella sp. e Shigella sp. O padrão de sensibilidade a antibióticos foi testado. Este é o primeiro relato de infecção assintomática por Salmonella sp. nesta espécie. Não foi detectada infecção por Shigella sp. Destaca-se a importância de investigação em animais submetidos a translocação ou a reintrodução e a necessidade de inquéritos periódicos em populações cativas a fim de se evitar agravos ocupacionais.(AU)

Fecal samples of captive souther brown howler monkeys were screened for the presence of Salmonella sp. and Shigella sp. The sensibility for antibiotics was also measured. This is the first report of asymptomatic infection by Salmonella sp. in this specie. Shigella sp. infection was not detected. Screening for this bacteria should be performed during translocation and reintroduction programs. We recommend periodic investigations to prevent occupational hazard on captivity.(AU)