Detecção de Aeromonas spp. e do gene de virulência aerolisina em tilápias do Nilo (Oreochromis niloticus) com a técnica de mPCR / Detection of Aeromonas spp. and virulence gene aerolysin in Nile tilapia (Oreochromis niloticus) using PCR technique

As infecções causadas por bactérias do gênero Aeromonas estão entre as doenças mais comuns em peixes cultivados em todo o mundo, com ocorrência de aeromoniose em todos os países que possuem cultivo de tilápia do Nilo (Oreochromis niloticus). O presente trabalho descreve o desenvolvimento de uma nova multiplex PCR (mPCR) para diagnóstico de Aeromonas spp. e identificação do gene aerolisina (aerA). Para padronização da mPCR foram utilizadas cepas de referência de várias espécies do gênero Aeromonas e de outros gêneros. Também foram usadas cepas de campo de A. hydrophila oriundas de cultivos de peixes pacamãs (Lophiosilurus alexandri) e Aeromonas spp. de tilápias do Nilo. Os primers foram desenhados com base na região 16S rRNA e aerA. Para verificar a melhor temperatura de anelamento foram utilizados gradientes entre 59°C a 61°C com 40ng de DNA molde. Os produtos da amplificação da região 16S rRNA e do gene aerA apresentaram 786 e 550pb, respectivamente. A mPCR apresentou melhor temperatura de anelamento a 57,6°C com limite de detecção das concentrações de DNA em ambos genes (16S rRNA and aerA) de 10-10g/μL. A mPCR padronizada é rápida, sensível e específica no diagnóstico de Aeromonas spp. e identificação do gene aerolisina. Esta metodologia apresenta vantagens quando comparada aos métodos de diagnóstico convencionais, podendo ser utilizada em cultivos comerciais de tilápias do Nilo ou outros peixes. A identificação do gene aerolisina é uma importante ferramenta na determinação do potencial patogênico dos isolados de Aeromonas spp. estudados.(AU)

Infections caused by bacteria of the genus Aeromonas are among the most common diseases in fish farming systems worldwide, and this disease occurs in all countries which have Nile tilapia (Oreochromis niloticus) farmed. The present work describes the development of a new multiplex PCR (mPCR) technique that diagnosis the genus Aeromonas and detects aerolysin gene (aerA). Reference strains of several Aeromonas species and other genera were used for standardization of mPCR. Strains of A. hydrophila from "pacaman" fish (Lophiosilurus alexandri) and Aeromonas spp. from Nile tilapia from farming systems were used too. Primers were designed based on the 16S rRNA region and aerA (aerolysin toxin). To verify a better annealing temperature were used gradients between 59°C and 61°C with 40ng of the DNA template. The 16S rRNA gene and the aerA gene amplification products showed 786 and 550 bp, respectively. The mPCR showed better annealing temperature at 57.6°C, and the detection limit for both genes (16S rRNA and aerA) was 10-10g/μL of the DNA. The standardized mPCR is quick, sensitive, and specific for Aeromonas spp. diagnosis and to detect aerolysin gene. This method showed advantages when compared to the conventional diagnostic methods and can be used in Nile tilapia or other fish farming systems. The detection of aerolysin gene is an important tool to determine the potential pathogenicity of Aeromonas spp. isolates.(AU)

Comparison of MALDI-TOF MS, Housekeeping Gene Sequencing, and 16S rRNA Gene Sequencing for Identification of Aeromonas Clinical Isolates

PURPOSE: The genus Aeromonas is a pathogen that is well known to cause severe clinical illnesses, ranging from gastroenteritis to sepsis. Accurate identification of A. hydrophila, A. caviae, and A. veronii is important for the care of patients. However, species identification remains difficult using conventional methods. The aim of this study was to compare the accuracy of different methods of identifying Aeromonas at the species level: a biochemical method, matrix-assisted laser desorption ionization mass spectrometry-time of flight (MALDI-TOF MS), 16S rRNA sequencing, and housekeeping gene sequencing (gyrB, rpoB). MATERIALS AND METHODS: We analyzed 65 Aeromonas isolates recovered from patients at a university hospital in Korea between 1996 and 2012. The isolates were recovered from frozen states and tested using the following four methods: a conventional biochemical method, 16S rRNA sequencing, housekeeping gene sequencing with phylogenetic analysis, and MALDI-TOF MS. RESULTS: The conventional biochemical method and 16S rRNA sequencing identified Aeromonas at the genus level very accurately, although species level identification was unsatisfactory. MALDI-TOF MS system correctly identified 60 (92.3%) isolates at the species level and an additional four (6.2%) at the genus level. Overall, housekeeping gene sequencing with phylogenetic analysis was found to be the most accurate in identifying Aeromonas at the species level. CONCLUSION: The most accurate method of identification of Aeromonas to species level is by housekeeping gene sequencing, although high cost and technical difficulty hinder its usage in clinical settings. An easy-to-use identification method is needed for clinical laboratories, for which MALDI-TOF MS could be a strong candidate.

Markers of pathogenicity islands in strains of Aeromonas species of clinical and environmental origin.

The aim of this study was to investigate the presence of markers of pathogenicity islands that may be informative to detect the virulent PAI carriers of clinical and environmental strains of Aeromonas spp. isolated in Mexico. virB2, virB9 and virB11 genes were found in Aeromonas strains isolated from environmental and clinical sources while cagE and tfc16 genes were only in strains of environmental origin. Having performed the wide screening presented in this study, we now have a set of strains to map and confirm the presence of a pathogenicity island in Aeromonas strains isolated in Mexico.

Aeromonas spp asociada a enfermedad diarreica aguda en Cuba: estudios de casos y controles / Aeromonas spp asociated to acute diarrheic disease in Cuba: case-control study

The members of the genus Aeromonas are currently considered important gastrointestinal pathogens in different geographical areas. From February 1985 to January 2005 several case-control studies were coordinated by the National Reference Laboratory for Diarrheal Diseases from the Pedro Kouri Institute. The study purpose was to analyze a possible pathogenic role for Aeromonas spp in Cuban children with acute diarrhea. In that period 2,322 children less than 5 years old with acute diarrhea were studied for diarhoeal pathogens and another group of 2,072 non hospitalized children without diarrhea during the similar time from the same geographical areas and matched by ages were recruited. In the group of children with diarrheas (cases), Aeromonas spp. was isolated in 166 (7.15%) and in the control group the microorganism was found in only 35 (1.76%). When Aeromonas isolation rates were compared between both groups, we found that probability to isolate this specie was significantly higher in cases than in controls (OR = 4.48, 95% IC: 3.05-6.60; P < 0.001). The Aeromonas species more frequently isolated were A. caviae, A. hydrophila, and A. veronii bv sobria. Other enteric pathogens detected in children with diarrhea were: Shigella spp in 418 (18%) (P < 0.0001), Salmonella spp in 53 (2.3%) (P < 0.01), and enteropathogenic E. coli in 58 (2.49%) (P < 0.05).

Los miembros del género Aeromonas son considerados patógenos importantes del tracto gastrointestinal en diferentes áreas geográficas. De febrero de 1985 a enero de 2005 se realizaron estudios de casos y controles en el Laboratorio Nacional de Referencia de Enfermedades Diarreicas Agudas del Instituto Pedro Kourí con el objetivo de conocer el comportamiento de los microorganismos pertenecientes al género Aeromonas en niños con diarreas en Cuba. La muestra estuvo constituida por 2.322 niños bajo 5 años de edad, ingresados por enfermedad diarreica aguda y como grupo control se estudió un total de 2.072 niños con edades y áreas geográficas similares que acudieron a los hospitales correspondientes en ese mismo período de tiempo. En el grupo de niños que presentaron diarreas, Aeromonas spp fue aislada en 166 (7,15%), y en los controles fue encontrada en 35 (1,76%). Al comparar la positividad para Aeromonas entre ambos grupos, la probabilidad de diagnosticar este microorganismo fue 4,28 veces mayor en los casos que en los controles (OR = 4,28; IC al 95%: 2,96-6,20; P < 0,001). Las especies más frecuentemente aisladas fueron A. caviae, A. hydrophila, y A. veronii bv sobria. Otros enteropatógenos aislados en niños que presentaban diarrea fueron: Shigella spp. en 418 (18%) (P < 0,0001, Salmonella spp en 53 (2,3%) (P < 0,01),) y E. coli enteropatógena en 58 (2,49%) (P < 0,05).

Caracterización fenotípica de cepas de Aeromonas aisladas de pacientes con enfermedad diarreica aguda / Phenotypic characterization of Aeromonas isolated from patients with acute diarrheal diseases

INTRODUCCIÓN: las enfermedades diarreicas agudas constituyen una de las principales causas de morbilidad y mortalidad en los niños menores de 5 años y en la población general, esto ocasiona una gran demanda de atención en los servicios de salud. La situación se agrava por el abuso de los antimicrobianos y el desarrollo de resistencia bacteriana, suceso que hoy día constituye un problema emergente de salud en diversas regiones del mundo. Entre los microorganismos causantes de enfermedades diarreicas agudas se encuentran los pertenecientes al género Aeromonas, los cuales han sido reconocidos como patógenos emergentes de riesgo II. OBJETIVOS: determinar las especies de Aeromonas más frecuentemente aisladas a partir de muestras de heces de pacientes con enfermedades diarreicas agudas y su susceptibilidad antimicrobiana. MÉTODOS: se determinó la susceptibilidad mediante el método de Bauer-Kirby frente a diferentes antimicrobianos a 100 aislamientos remitidos desde los centros provinciales de higiene y epidemiología del país durante 2007-2008. RESULTADOS: En 67 por ciento de los aislamientos se logró la identificación hasta especie, se observó el predominio de A. caviae (33 por ciento) y A. hydrophila (29 por ciento). Se demostró que 100 por ciento de los aislamientos resultaron resistentes al menos a un antimicrobiano de los investigados. El porcentaje más elevado de resistencia se observó frente a la cefalotina, las sulfonamidas y el ácido nalidíxico. CONCLUSIONES: se propone a la tetraciclina y el cloranfenicol como fármacos de elección para el tratamiento de las infecciones intestinales producidas por estos microorganismos en Cuba.

INTRODUCTION: the acute diarrheal diseases are one of the main causes of morbidity and mortality in children aged under 5 years and in the general population; this demands a great deal of care in the healthcare services. The situation worsens due to the overuse of antimicrobials and the development of bacterial resistance, being the latter an emerging health problem in different areas of the world. Among the causative microorganisms of acute diarrheal diseases are those of Aeromonas genus, recognized as second risk emerging pathogens. OBJECTIVES: to determine the most frequently isolated Aeromonas species in fecal samples from acute diarrheal patients and their antimicrobial susceptibility. METHODS: the Bauer-Kirby´s method allowed identifying the susceptibility to several antimicrobials of 100 isolated samples coming from the provincial hygiene and epidemiology centers during 2007 and 2008. RESULTS: identification of the species was accomplished in 67 percent of isolates, being A. caviae (33 percent) y A. hydrophila (29 percent) the predominant species. It was demonstrated that 100 percent of isolates got resistant to at least one of the studied antimicrobials. Drug resistance to cefalotine, sulfonamides and nalidixic acid showed the highest percentages. CONCLUSIONS: tetracycline and chloramphenicol are recommended as the drugs of choice for treating intestinal infections caused by these microorganisms in Cuba.

Caracterización fenotípica y factores de virulencia en cepas de Aeromonas aisladas de pacientes con enfermedad diarreica aguda en Cuba / Phenotypic characteristics and virulence factors in Aeromonas strains isolated from patients with diarrheic disease in Cuba

Fifty four strains of Aeromonas spp were isolated from patients with acute diarrheic episodes by using Aerokey II and Aeroesquema methods. In vitro antimicrobial susceptibility and virulence factors were analyzed. The most frequently isolated specie was Aeromonas caviae. Over 75 percent of strains exhibited resistance to penicillins and ce-phalosporins; for the other antibiotic groups resistance was under 20 percent. Twenty six strains (48.1 percent) were multiresist-ant. At least one virulence factor among those evaluated in the study was present in 53 (98.1 percent) of the 54 strains.

Se identificaron 54 cepas de Aeromonas aisladas de pacientes con enfermedad diarreica aguda mediante los métodos Aerokey II y Aeroesquema. Se determinó la susceptibilidad antimicrobiana y algunos factores de virulencia. La especie encontrada en mayor frecuencia fue Aeromonas caviae. Se observaron valores de resistencia por sobre 75 por ciento para penicilinas y cefalotina; para el resto de los antimicrobianos estos valores se encontraron bajo 20 por ciento>; 26 cepas (48,1 por ciento) presentaron multi-resistencia. Al menos un factor de virulencia de los investigados estuvo presente en 53 (98,1 por ciento) de las 54 cepas analizadas.

Aeromonas caviae septicemia in immunocompetent gastrointestinal carriers

Aeromonas caviae strains have been isolated from blood and stool cultures of three immunocompetent patients, residents of Northern India, who presented with community acquired septicemia without any recent history of diarrhea. Cell culture infectivity test performed on Hep-2 cells have shown substantial degree of invasiveness in the isolated strains. This case unleashes a possibility of asymptomatic gastrointestinal carriage of such strains of A. caviae in a very large population of India, as several areas of India have very high rates of Aeromonas induced acute diarrhea/gastroenteritis (up to 13 percent). It needs to be appraised further in India as well as other countries having high rates of Aeromonas induced acute diarrhea/gastroenteritis.

Enzymatic characterization of Vibrio alginolyticus strains isolated from bivalves harvested at Venice Lagoon (Italy) and Guanabara Bay (Brazil) / Caracterização enzimática de cepas de Vibrio alginolyticus isoladas de bivalves coletados na Lagoa de Veneza (Itália) e Baía de Guanabara (Brasil)

The aquatic ecosystem is the natural habitat of microorganisms including Vibrio and Aeromonas genus which are pathogenic to human and animals. In the present investigation the frequency of these bacteria and the enzymatic characteristics of 34 Vibrio alginolyticus strains isolated from bivalves harvested in Venice Lagoon (Italy) and Guanabara Bay (Brazil) were carried out from November 2003 to February 2004. The mussels' samples were submitted to enrichment in Alkaline Peptone Water (APW) added with 1 percent of sodium chloride (NaCl) and APW plus 3 percent NaCl incubated at 37 ºC for 18-24h. Following the samples were streaked onto TCBS Agar (Thiossulfate Citrate Bile Sucrose Agar) and the suspected colonies were submitted to biochemical characterization. Also, the Vibrio alginolyticus strains were evaluated to collagenase, elastase and chondroitinase production. The results showed the isolation of 127 microorganisms distributed as follows: 105 Vibrio strains such as V. alginolyticus (32.4 percent), V. harveyi (19 percent) and V. parahaemolyticus (7.6 percent), 20 Aeromonas strains and two Plesiomonas shigelloides were the main pathogens isolated. We observed the production of the three enzymes from V. alginolyticus strains considered as the main virulence factors of the bacteria, especially in cases of human dermatological infection.

O ecossistema aquático é o habitat natural de microrganismos incluindo aqueles dos gêneros Vibrio e Aeromonas os quais são patogênicos para o homem e animais. Na presente investigação foi avaliada a freqüência destas bactérias e a característica enzimática de 34 cepas de Vibrio alginolyticus isoladas de bivalves coletados na Lagoa de Venice (Itália) e Baía de Guanabara (Brasil) durante o período de Novembro-2003 a Fevereiro-2004. As amostras de mexilhões foram submetidas a enriquecimento em Água Peptonada Alcalina (APA) adicionada de 1 por cento de Cloreto de Sódio (NaCl) e APA com 3 por cento de NaCl (37 ºC/18-24h). Em seguida as amostras foram semeadas em Agar TCBS (Agar Tiossulfato Citrato Bile Sacarose) e as colônias suspeitas foram submetidas à caracterização bioquímica. As cepas de Vibrio alginolyticus foram avaliadas quanto à produção das enzimas colagenase, elastase e condroitinase. Os resultados demonstraram o isolamento de 127 microrganismos assim distribuídos: 105 cepas de Vibrio das quais V. alginolyticus (32,4 por cento), V. harveyi (19 por cento) e V. parahaemolyticus (7,6 por cento), 20 cepas de Aeromonas e 2 Plesiomonas shigelloides foram os principais patógenos isolados. Observou-se a produção das três enzimas a partir de V. alginolyticus, consideradas principais fatores de virulência da bactéria, em especial em casos de infecção dermatológica humana.

Caracterização de Aeromonas spp isoladas de neonatos hospitalizados / Characterization of Aeromonas spp isolates from newborns hospitalized

Aeromonas spp é reconhecida como patogênica para o homem após o consumo de água e alimentos contaminados. Na presente investigação, foram avaliadas 2.323 amostras de swabs retais de neonatos hospitalizados no Rio de Janeiro objetivando o isolamento de Aeromonas. As amostras foram coletadas e enviadas ao Laboratório de Referência Nacional de Cólera e outras enteroinfecções bacterianas, Instituto Oswaldo Cruz, Fundação Oswaldo Cruz. Os swabs foram submetidos ao enriquecimento em água peptonada alcalina adicionada de 1 por cento de cloreto de sódio (NaCl) e água peptonada alcalina adicionada de 3 por cento de NaCl (37ºC/18-24h) e semeadas em agar seletivo para Pseudomonas aeromonas (Agar GSP). Foram isoladas 56 cepas de Aeromonas assim distribuídas: Aeromonas caviae (42,8 por cento), Aeromonas media (25 por cento), Aeromonas veronii biogrupo sobria (10,7 por cento), Aeromonas hydrophila (9 por cento), Aeromonas veronii biogrupo veronii (5,3 por cento), Aeromonas sobria (1,8 por cento), Aeromonas jandaei (1,8 por cento), Aeromonas schubertii (1,8 por cento) e Aeromonas sp (1,8 por cento). Foi observada resistência a uma ou mais drogas antimicrobianas em 26,8 por cento das cepas. Considerando a relevância de Aeromonas torna-se urgente alertar sobre sua importância para o controle de infecções hospitalares.

Aeromonas spp is recognized as pathogenic to humans after consumption of contaminated water and food. In the present investigation, 2,323 rectal swab samples from newborns hospitalized in Rio de Janeiro were evaluated with a view to isolating Aeromonas. The samples were collected and sent to the national reference laboratory for cholera and other bacterial intestinal infections, at the Oswaldo Cruz Institute of the Oswaldo Cruz Foundation. The swabs were subjected to enrichment in alkaline peptonated water with the addition of 1 percent sodium chloride (NaCl) and alkaline peptonated water plus 3 percent NaCl (37°C/18-24h) and were streaked onto agar that was selective for Pseudomonas-Aeromonas (GSP Agar). Fifty-six Aeromonas strains were isolated, distributed as follows: Aeromonas caviae (42.8 percent), Aeromonas media (25 percent), Aeromonas veronii biogroup sobria (10.7 percent), Aeromonas hydrophila (9 percent), Aeromonas veronii biogroup veronii (5.3 percent), Aeromonas sobria (1.8 percent), Aeromonas jandaei (1.8 percent), Aeromonas schubertii (1.8 percent) and Aeromonas sp (1.8 percent). Resistance to one or more antimicrobial drugs was observed in 26.8 percent of the strains. Considering the importance of Aeromonas, there is an urgent need to warn about this in relation to nosocomial infection control.

Concordância entre perfil de restrição do fragmento 16S rDNA e testes fenotípicos para determinação do posicionamento taxonômico de cepas de Aeromonas / Consistency between profile of restriction fragment of the 16S rDNA and phenotypic tests to determine the taxonomic position of strains of Aeromonas

A taxonomia do gênero Aeromonas é controversa e sofre mudanças constantemente, isso devido às novas espécies que vêm sendo descritas, reclassificadas ou de estudo mais aprofundados em espécies já conhecidas. Devido à heterogeneidade fenotípica apresentada entre as espécies do gênero, consideráveis esforços estão direcionados para o desenvolvimento de métodos que permitam identificar e reclassificar corretamente as espécies Aeromonas. Com base no 16S rDNA das espécies de Aeromonas, o presente estudo propõe um par de inibidores gênero-específico para Aeromonas e um esquema utilizando enzimas de restrição para identificação das espécies do gênero. Foram analisadas 40 cepas de amostras ambientais, previamente identificadas fenotipicamente como membros do gênero Aeromonas, todas as cepas foram confirmadas com o par de inibidores gênero-específico. Os resultados da identificação das espécies utilizando o esquema de RFLP foi seguinte A. jandaei 35 por cento, A. hydrophila 30 por cento, A. trota 5 por cento, A. aquariorum 12,5 por cento. A. veronii 10 por cento, e A. media 7,5 por cento. Os métodos apresentados no estudo podem ser empregados na rotina laboratorial podendo ser útil no estudo epidemiológico do gênero Aeromonas bem como na determinação da real distribuição dos organismos deste gênero nos diferentes ambientes, em alimentos e em casos clínicos.

Aislamiento e identificación de especies pertenecientes a los géneros Aeromonas, Vibrio y Plesiomonas procedentes de muestras extra-intestinales en Cuba / Isolation and identification of species from the genera Aeromonas, Vibrio, and Plesiomonas from extraintestinal samples in Cuba

Las infecciones extra-intestinales producidas por los géneros Aeromonas, Vibrio y Plesiomonas presentan una elevada tasa de morbimortalidad en diferentes áreas geográficas. De enero 2002 a diciembre 2003 se recibieron en el Laboratorio Nacional de Referencia de Enfermedades Diarreicas Agudas del Instituto de Medicina Tropical Pedro Kourí, 95 cepas de bacilos gramnegativos, anaerobios facultativos, oxi-dasa positiva, procedentes de muestras extra-intestinales (hemocultivos, exudados óticos, pus de heridas, exudados conjuntivales, urocultivos, catéter, entre otras) remitidas de diferentes provincias del país. Se demostró la presencia de Aeromonas caviae, Aero-monas veronii bv sobria, Aeromonas jandaei, Vibrio cholerae no -O1, Vibrio vulnificus, Vibrio fluvialis y Plesiomonas shigelloides en las muestras estudiadas.

Extraintestinal infections caused by the genera Aeromonas, Vibrio and Plesiomonas have high morbidity and mortality rates in different areas of world. From January 2002 to December 2003, the National Reference Laboratory for Acute Diarrhoeal Diseases of the Pedro Kourí Tropical Medicine Institute received 95 gramnegative, facultative anaerobic, oxidase positive bacilli strains from different extraintestinal specimen (blood, ear exudates, infected wounds, conjunctive exudates, urine, and catheters, among others) sent by different provincial laboratories along the country. Aeromonas caviae, Aeromonas veronii bv sobria, Aeromonas jandaei, Vibrio cholerae no-O1, Vibrio vulnificus, Vibrio fluvialis and Plesiomonas shigelloides were the species most frequently found in the sample analysed.

Envolvimento de Aeromonas em surto de doenca diarréica aguda em São Bento do Una, Pernambuco / Aeromonas associated with an acute diarrhea outbreak in São Bento do Una, Pernambuco

No primeiro semestre de 2004, ocorreu um surto de diarréia em São Bento do Una, Pernambuco, registrando-se 2.170 casos. Nas 582 coproculturas realizadas, 145 (25 por cento) revelaram um enteropatógeno bacteriano, destacando 114 casos (19,5 por cento) com a participacão de Aeromonas, representadas por Aeromonas caviae (57/9,8 por cento), Aeromonas veronii biovar sobria (23/3,9 por cento), Aeromonas veronii biovar veronii (15/2,6 por cento) e outras espécies (19/3,2 por cento). Nos 31 episódios restantes (5,3 por cento), foram detectados: V. cholerae O1 Ogawa toxigênico (18/3,1 por cento), Salmonella spp (8/1,4 por cento), Shigella spp (3/0,5 por cento) e Vibrio cholerae não O1/não O139 (2/0,3 por cento).

Serogroups, K1 antigen, and antimicrobial resistance patterns of Aeromonas spp. strains isolated from different sources in Mexico

A total of 221 strains of Aeromonas species isolated in Mexico from clinical (161), environmental (40), and food (20) samples were identified using the automated system bioMérieux-Vitek®. Antisera for serogroups O1 to 044 were tested using the Shimada and Sakazaki scheme. The K1 antigen was examined using as antiserum the O7:K1C of Escherichia coli. Besides, we studied the antimicrobial patterns according to Vitek AutoMicrobic system. Among the 161 clinical strains 60 percent were identified as A. hydrophila, 20.4 percent as A. caviae, and 19.25 percent as A. veronii biovar sobria. Only A. hydrophila and A. veronii biovar sobria were found in food (55 and 90 percent respectively) and environmental sources (45 and 10 percent respectively). Using "O" antisera, only 42.5 percent (94/221) of the strains were serologically identified, 55 percent (121/221) were non-typable, and 2.5 percent (6/221) were rough strains. Twenty-two different serogroups were found, O14, O16, O19, O22, and O34 represented 60 percent of the serotyped strains. More than 50 percent of Aeromonas strain examined (112/221) expressed K1 encapsulating antigen; this characteristic was predominant among Aeromonas strains of clinical origin. Resistance to ampicillin/sulbactam and cephazolin was detected in 100 and 67 percent of Aeromonas strain tested for their susceptibility to antibiotics. In conclusion, antibiotic-resistant Aeromonas species that possess the K1 encapsulating antigen and represent serogroups associated with clinical syndrome in man are not uncommon among Aeromonas strains isolated from clinical, food and environmental sources in Mexico.

Diversidade e perfil plasmidial de bactérias do gênero Aeromonas isoladas de ambientes aquáticos do Estado de São Paulo / Diversity and plasmidial profile of bactéria of the genus Aeromonas isolated from aquatic evironments os State of São Paulo

Introdução: O gênero Aeromonas é ubiqüitário no ambiente aquático e já foi isolado de água doce, salgada, salobra, mineral, poluída ou tratada. Várias espécies de Aeromonas apresentam fatores de virulência e podem estar relacionadas a uma série de doenças para seu hospedeiro. Objetivo: Estudar a diversidade e caracterizar molecularmente através do perfil plasmidial, as diferentes espécies do gênero Aeromonas provenientes de corpos hídricos de São Paulo. Métodos: Foram coletadas 19 amostras de água, sendo 11 amostras de água doce, 1 de água salobra,e 7 de água salgada. As amostras foram concentradas através de membrana filtrante e enriquecidas em 100mL de água peptonada alcalina 1por cento. As amostras foram estriadas em placas de petri contendo diferentes meios de cultivo. As colônias típicas foram submetidas a provas bioquímicas para a identificação das espécies. Foi feito o levantamento bibliográfico do Índice de qualidade de Água (IQA) na CETESB, dos pontos de coleta analisados. Para análise do perfil plasmidial, foi utilizada a metodologia descrita por BIRNBORN e DOLU (1979). Resultados: As espécies de Aeromonas foram isoladas em 84,2por cento dos ambientes isolados; segundo os dados do CETESB, 58,3por cento das amostras apresentaram condições favoráveis de qualidade de água; 43,2por cento das cepas isoladas apresentaram pelo menos um plasmídio. Conclusões: Concluiu-se que esses organismos estão amplamente distribuídos no ambiente aquático e que as espécies de Aeromonas não apresentam o mesmo perfil plasmidial, Os organismos isolados dos ambientes estudados podem representar risco à saúde.

Prevalence of Aeromonas in different clinical and water samples with special reference to gastroenteritis.

A hospital based study was carried out in the Department of Microbiology, Tribhuvan University Teaching Hospital, Institute of Medicine with the aim to initiate the isolation and identification of Aeromonas spp. from the stool samples of gastroenteritic patients and to determine the prevalence of Aeromonas spp. in other clinical samples and water. Altogether 293 samples were investigated that include 172 stool samples, 60 pus/wound swabs, 20 body fluids and 41 water samples. The samples were collected and processed by standard microbiological techniques in order to isolate Aeromonas. Ampicillin blood agar (20 microg/ml) was used as selective medium for the isolation of Aeromonas. The specimen prevalence rate of Aeromonas spp. in stool was found to be 5.2% and the A. hydrophila (55.5%) was the predominant species followed by A. caviae (33.3%) and A. sobria (11.1%) in the stool samples. Likewise, 3.3% of pus sample showed positive growth of A. hydrophila. Aeromonas was not detected in any of the body fluids. Aeromonas spp. were isolated from 58.5% of water samples obtained from different hospitals. The commonest species was A. hydrophila (62.5%) followed by A. caviae (20.8%) and A. sobria (16.7%). In vitro susceptibility testing showed that the aminoglycosides and fluroquinolones were the effective antibiotics against Aeromonas. It was found that 88.9% of Aeromonas spp. isolated from stool samples were sensitive to gentamicin, 77.8% to ciprofloxacin, norfloxacin, tetracycline and ceftriaxone and 66.6% to nalidixic acid whereas cent percent Aeromonas spp. from water samples were sensitive to gentamicin, ciprofloxacin, norfloxacin, tetracycline and ceftriaxone and 75.0% to nalidixic acid. The enteric Aeromonas isolates were more resistant to antimicrobial agents than the water isolates.

Correlation of the suicide phenomenon in Aeromonas species with virulence and enteropathogenicity.

Certain strains of mesophilic Aeromonads like A. hydrophila, A. veronii biotype sobria and A. caviae when grown in broth containing 0.5% glucose, undergo growth inhibition concomitant with acetate accumulation. As these strains become nonviable after 24 h, this phenomenon is termed suicide. We investigated suicidal strains of Aeromonas species as means of understanding animal virulence and enteropathogenicity. Non suicidal strains of A. Hydrophila showed and overall 88.8% lethality rate and non suicidal strains of A. veronii biotype sobria showed 83.3% lethality rate and was nil for its suicidal part. Of the two suicidal A. caviae strains tested, none were lethal. The present data suggest that the suicide phenomenon may explain strain specific [A. veronii biotype sobria, A. hydrophila] and species specific [A. caviae] virulence and enteropathogenicity.

Typing of Aeromonas isolates from children with diarrhoea & water samples by randomly amplified polymorphic DNA polymerase chain reaction & whole cell protein fingerprinting.

BACKGROUND & OBJECTIVES: Aeromonas spp. are water-borne organisms, often associated with childhood diarrhoea. The present study was conducted to examine the epidemiological relationship among the Aeromonas spp. isolated from water and children with acute diarrhoea in Chennai. METHODS: Thirty six Aeromonas isolates inclusive of 16 from children with diarrhoea, 15 from domestic water samples and 5 reference strains were studied by randomly amplified polymorphic DNA polymerase chain reaction (RAPD-PCR). Twenty eight Aeromonas isolates, 15 from children with diarrhoea, 10 from domestic water samples and three reference strains were analysed by SDS-PAGE for their whole cell protein profiles. RESULTS: The 36 Aeromonas isolates examined by RAPD-PCR generated RAPD fingerprints with majority of the bands ranging from about 250 to 2800 bp. The RAPD fingerprints did not correspond with the phenospecies and varied greatly among the strains within the phenospecies. Cluster analysis revealed two major groups at 75 per cent hierarchical level, comprising 18 Aeromonas isolates, mainly recovered from domestic water samples, while the clinical isolates were scattered in different hierarchical levels in the dendrogram. The whole cell protein fingerprints examined by SDS-PAGE did not correspond with the phenospecies. Only four isolates of A. caviae were found to produce similar protein fingerprints allowing them to form a cluster at about 90 per cent hierarchical level, while the rest of the isolates were scattered at various hierarchical levels in the dendrogram. INTERPRETATION & CONCLUSIONS: In the present study, RAPD fingerprinting was found to be useful in distinguishing Aeromonas isolates recovered from clinical and domestic water supplies. However, RAPD-PCR could not distinguish the phenospecies of the genus Aeromonas. Whole cell protein fingerprinting and cluster analysis could neither differentiate isolates from clinical and domestic water sources nor the phenospecies of the genus Aeromonas.

Aplicación de la actividad pirazinamidasa para la identificación en especies de Aeromonas / Application of the pyrazinamidase activity for the identification of Aeromonas species

Se ubicaron en especies 70 cepas de Aeromonas aisladas de pacientes con enfermedad diarreica aguda, utilizando las pruebas del factor CAMP y la actividad pirazinamidasa. Se destaca el alto tanto por ciento de coincidencia entre ambas pruebas de fácil realización en los laboratorios de microbiología clínica

Estudio comparativo de 4 métodos para la identificación de especies del género Aeromonas / Comparative study of 4 methods for the identification of species of the genus Aeromonas

Se exponen brevemente los resultados obtenidos con la aplicación de los métodos AEROKEY II, AEROKEY II + esquema de Abbott, Api 20 NE, Sistema Biolog. El estudio se llevó a cabo con 38 cepas de Aeromonas aisladas de niños menores de 5 años con enfermedad diarreica aguda (EDA), el AEROKEY II + esquema de Abbott resultó el mejor método de identificación