A Class 4-like Chromosomal Integron Found in Aeromonas sp. Genomospecies paramedia Isolated from Human Feces.

Integrons are genetic elements that store, express and exchange gene cassettes. These elements are characterized by containing a gene that codes for an integrase (intI), a cassette integration site (attI) and a variable region holding the cassettes. Using bioinformatics and molecular biology methods, a functional integron found in Aeromonas sp. 3925, a strain isolated from diarrheal stools, is described. To confirm the integron class, a phylogenetic analysis with amino acid sequences was conducted. The integrase was associated to class 4 integrases; however, it is clearly different from them. Thus, we classified the associated element as a class 4-like integron. We found that the integrase activity is not under the control of the SOS or catabolic repression, since the expression was not increased in the presence of mitomycin or arabinose. The class-4-like integron is located on the chromosome and contains two well-defined gene cassettes: aadA1 that confers resistance to streptomycin and lpt coding for a lipoprotein. It also includes eight Open Reading frames (ORFs) with unknown functions. The strain was characterized through a Multilocus Phylogenetic Analyses (MLPA) of the gyrB, gyrA, rpoD, recA, dnaJ and dnaX genes. The phylogenetic results grouped it into a different clade from the species already reported, making it impossible to assign a species. We resorted to undertaking complete genome sequencing and a phylogenomic analysis. Aeromonas sp. 3925 is related to A. media and A. rivipollensis clusters, but it is clearly different from these species. In silico DNA-DNA hybridization (isDDH) and Average Nucleotide Identity (ANI) analyses suggested that this isolate belongs to the genomospecies paramedia. This paper describes the first class 4-like integron in Aeromonas and contributes to the establishment of genomospecies paramedia.

Prospecting of essential oils in combination with florfenicol against motile Aeromonas isolated from tambaqui (Colossoma macropomum).

Natural products with antimicrobial activity and their association with synthetic antimicrobials are a sustainable option in fish farming. The objective of this study was to determine antimicrobial activity, antibiofilm potential and synergism of five essential oils (EOs) with florfenicol against motile Aeromonas isolated from Amazonian Colossoma macropomum. As their major constituent, the EOs of the species of Aloysia triphylla, Croton cajucara (red and white morphotype), Cymbopongo citratus and Lippia gracilis present ß-pinene (22.1%), germacrene D (11.5%), linalool (23%), geranial (45.7%) and carvacrol (42.2%), respectively. The EOs of L. gracilis and C. citratus showed the best antimicrobial activities against the Aeromonas strains (5 mg mL-1). All EOs interfered with biofilm formation and consolidated biofilm. The EOs of A. triphylla, C. citratus and L. gracilis showed a synergistic effect with florfenicol, reducing the amount of the chemical into the water systems while treatment.

Aeromonas and Plesiomonas species from scarlet ibis (Eudocimus ruber) and their environment: monitoring antimicrobial susceptibility and virulence.

The present study aimed at evaluating the role of captive scarlet ibises (Eudocimus ruber) and their environment as reservoirs of Aeromonas spp. and Plesiomonas spp., and analyzing the in vitro antimicrobial susceptibility and virulence of the recovered bacterial isolates. Thus, non-lactose and weak-lactose fermenting, oxidase positive Gram-negative bacilli were recovered from cloacal samples (n = 30) of scarlet ibises kept in a conservational facility and from water samples (n = 30) from their environment. Then, the antimicrobial susceptibility, hemolytic activity and biofilm production of the recovered Aeromonas spp. and Plesiomonas shigelloides strains were assessed. In addition, the virulence-associated genes of Aeromonas spp. were detected. Ten Aeromonas veronii bv. sobria, 2 Aeromonas hydrophila complex and 10 P. shigelloides were recovered. Intermediate susceptibility to piperacillin-tazobactam and cefepime was observed in 2 Aeromonas spp. and 1 P. shigelloides, respectively, and resistance to gentamicin was observed in 4 P. shigelloides. The automated susceptibility analysis revealed resistance to piperacillin-tazobactam and meropenem among Aeromonas spp. and intermediate susceptibility to gentamicin among P. shigelloides. All Aeromonas isolates presented hemolytic activity, while 3 P. shigelloides were non-hemolytic. All Aeromonas spp. and 3/10 P. shigelloides were biofilm-producers, at 28 °C, while 10 Aeromonas spp. and 6/10 P. shigelloides produced biofilms, at 37 °C. The most prevalent virulence genes of Aeromonas spp. were asa1 and ascV. Scarlet ibises and their environment harbour potentially pathogenic bacteria, thus requiring monitoring and measures to prevent contamination of humans and other animals.

Impact of stress on Aeromonas diversity in tambaqui (Colossoma macropomum) and lectin level change towards a bacterial challenge.

Tambaqui (Colossoma macropomum) is among the most cultivated fish species in tropical countries. Stress is the main cause of disease in fish farms. The genus Aeromonas is a common causative agent of fish diseases. This work reports the identification of Aeromonas species colonizing gills of C. macropomum submitted or not to a confinement stress. We also evaluated changes in serum levels of lectins (carbohydrate-binding proteins that are components of fish immune system) in tambaqui submitted to a challenge using two isolated Aeromonas strains. Gill tissues from stressed and unstressed fishes were used to isolate Aeromonas. Then 72 Aeromonas strains were isolated, 97% being from stressed fishes. Among these, 63 were identified at species level and 6 were classified as atypical Aeromonas strains. The most prevalent species were Aeromonas bestiarum and Aeromonas caviae and their strains were used in bacterial challenges. The lectin serum levels significantly increased after 24 h of infection with A. bestiarum; however, no significant increase was found for infection with A. caviae. In conclusion, C. macropomum gills are susceptible to colonization by different Aeromonas species, mainly at confinement stressful conditions, and serum lectins may have a role in the acute immunological response towards infection by A. bestiarum.

Pro-inflammatory caspase-1 activation during the immune response in cells from rainbow trout Oncorhynchus mykiss (Walbaum 1792) challenged with pathogen-associated molecular patterns.

In response to pathogens, the higher vertebrate innate immune system activates pro-inflammatory caspase-1 which is responsible for the processing and secretion of several important cytokines involved in the host's defence against infection. To date, caspase-1 has been described in few teleost fish, and its activity has been demonstrated through substrate cleavage and inhibition by pharmacological agents. In this study, the detection of the active form of caspase-1 during the immune response in salmonid fish is described, where two antibodies were produced. These antibodies differentially recognize the structural epitopes of the inactive pro-caspase-1 and the processed active form of the caspase. Firstly, caspase-1 activation was demonstrated in vitro by ELISA, Western blotting and immunocytochemistry in rainbow trout macrophages exposed to different pathogen-associated molecular patterns plus the pathogen Aeromonas hydrophila. This activity was clearly abrogated by a caspase inhibitor and seems to be unrelated to IL-1ß secretion. Caspase-1 activation was then validated in vivo in gill cells from fish challenged with Aeromonas salmonicida. These results represent the first demonstration of caspase-1 activation in salmonids, and the first evidence of the putative regulatory role which this protease plays in inflammatory response in this fish group, as described for some other teleosts and mammals.

Chemical composition and antibacterial activity of Aloysia triphylla (L'Hérit) Britton extracts obtained by pressurized CO2 extraction

This study investigated the chemical composition of five different extracts of Aloysia triphylla and their activity against Aeromonas sp. The extracts were obtained from the dried leaves by pressurized CO2 extraction at 30, 50 and 70ºC, and 100, 150, and 200 bar, and analyzed by GC/FID and GC-MS. The antibacterial activity was assayed by the microdilution method. The tested microorganisms comprised seven Aeromonas isolates obtained from the kidney of infected silver catfish, Rhamdia quelen. The yield, chemical composition and antibacterial activity of the extracts were dependent on the extraction conditions. Mono and sesquiterpenoids were the major constituents of all the extracts and the highest extraction yield was obtained at 70ºC and 200 bar. A. triphylla presented moderate antibacterial activity against Aeromonas sp.

Ocorrência de Aeromonas sp. e microrganismos psicrotróficos e estimativa do prazo de validade comercial de filé de pintado (Pseudoplatystoma coruscans) mantidos sob refrigeração / Occurrence of Aeromonas sp. and psychrotrophic microorganisms and estimate the shelf life of "pintado" (Pseudoplatystoma coruscans) fillets kept under refrigeration

Verificou-se a ocorrência de bactérias do gênero Aeromonas e estimou-se o prazo de validade comercial de filés de pintado (Pseudoplatystoma coruscans) com pele, durante estocagem em refrigeração por meio da quantificação de microrganismos heterotróficos aeróbios psicrotróficos e análises físico-químicas para determinação do pH e detecção de amônia e gás sulfídrico. Foram utilizadas 45 amostras de filé de pintado, com aproximadamente 100 gramas cada, embaladas individualmente em polietileno de alta densidade e armazenadas em câmara frigorífica entre 0ºC e 3ºC. A cada dois a três dias de estocagem, três unidades de filés foram submetidas a análises microbiológicas e físico-químicas, totalizando 15 análises durante o período de estocagem. As contagens de Aeromonas sp. e microrganismos heterotróficos aeróbios psicrotróficos variaram de 1,89 a 9,47logUFC/g e 0 a 6,54logUFC/g, respectivamente. A variação do pH foi de 6,20 a 6,97, e as análises de amônia e gás sulfídrico foram negativas durante todo o período. O pH dos filés de pintado atingiu o limite máximo de 6,4 aos 23 dias de estocagem, e estimou-se o seu prazo de validade comercial.(AU)

This word studied the occurrence of bacteria from genus Aeromonas and estimate the shelf life of "pintado" fish fillets (Pseudoplatystoma coruscans), during cold storage, through the quantification of psychrotrophic aerobic microorganisms, physical and chemical analyses for presence of ammonia and gas sulphide (H2S) and pH as used in 45 samples of "pintado" fillets with approximately 100 grams each, individually packed in high density polyethylene and stored in cold storage (0ºC to 3ºC). Every 2-3 days of storage, 3 units of fillets were subjected to microbiological and physicochemical analysis for a total of 15 days during the storage period. The Aeromonas sp. and psychrotrophic microorganisms count varied from 1.89 to 9.47logCFU/g and 0 to 6.54logCFU/g, respectively. The pH variation was from 6.20 to 6.97 and ammonia and H2S analyses were negative during the whole period. According under Brazilian legislation (Brazil, 1981) estimated the commercial shelf life of "pintado" fillets being 23 days when the pH reached a value of 6.4. The pH of the "pintado" fillets reached the maximum limit of 6.4 at 23 days of storage, being its estimated commercial shelf life.(AU)

Ocorrência de Aeromonas sp. e microrganismos psicrotróficos e estimativa do prazo de validade comercial de filé de pintado (Pseudoplatystoma coruscans) mantidos sob refrigeração / Occurrence of Aeromonas sp. and psychrotrophic microorganisms and estimate the shelf life of "pintado" (Pseudoplatystoma coruscans) fillets kept under refrigeration

Verificou-se a ocorrência de bactérias do gênero Aeromonas e estimou-se o prazo de validade comercial de filés de pintado (Pseudoplatystoma coruscans) com pele, durante estocagem em refrigeração por meio da quantificação de microrganismos heterotróficos aeróbios psicrotróficos e análises físico-químicas para determinação do pH e detecção de amônia e gás sulfídrico. Foram utilizadas 45 amostras de filé de pintado, com aproximadamente 100 gramas cada, embaladas individualmente em polietileno de alta densidade e armazenadas em câmara frigorífica entre 0ºC e 3ºC. A cada dois a três dias de estocagem, três unidades de filés foram submetidas a análises microbiológicas e físico-químicas, totalizando 15 análises durante o período de estocagem. As contagens de Aeromonas sp. e microrganismos heterotróficos aeróbios psicrotróficos variaram de 1,89 a 9,47logUFC/g e 0 a 6,54logUFC/g, respectivamente. A variação do pH foi de 6,20 a 6,97, e as análises de amônia e gás sulfídrico foram negativas durante todo o período. O pH dos filés de pintado atingiu o limite máximo de 6,4 aos 23 dias de estocagem, e estimou-se o seu prazo de validade comercial.(AU)

This word studied the occurrence of bacteria from genus Aeromonas and estimate the shelf life of "pintado" fish fillets (Pseudoplatystoma coruscans), during cold storage, through the quantification of psychrotrophic aerobic microorganisms, physical and chemical analyses for presence of ammonia and gas sulphide (H2S) and pH as used in 45 samples of "pintado" fillets with approximately 100 grams each, individually packed in high density polyethylene and stored in cold storage (0ºC to 3ºC). Every 2-3 days of storage, 3 units of fillets were subjected to microbiological and physicochemical analysis for a total of 15 days during the storage period. The Aeromonas sp. and psychrotrophic microorganisms count varied from 1.89 to 9.47logCFU/g and 0 to 6.54logCFU/g, respectively. The pH variation was from 6.20 to 6.97 and ammonia and H2S analyses were negative during the whole period. According under Brazilian legislation (Brazil, 1981) estimated the commercial shelf life of "pintado" fillets being 23 days when the pH reached a value of 6.4. The pH of the "pintado" fillets reached the maximum limit of 6.4 at 23 days of storage, being its estimated commercial shelf life.(AU)

Ocorrência de Aeromonas sp. e microrganismos psicrotróficos e estimativa do prazo de validade comercial de filé de pintado (Pseudoplatystoma coruscans) mantidos sob refrigeração / Occurrence of Aeromonas sp. and psychrotrophic microorganisms and estimate the shelf life of "pintado" (Pseudoplatystoma coruscans) fillets kept under refrigeration

Verificou-se a ocorrência de bactérias do gênero Aeromonas e estimou-se o prazo de validade comercial de filés de pintado (Pseudoplatystoma coruscans) com pele, durante estocagem em refrigeração por meio da quantificação de microrganismos heterotróficos aeróbios psicrotróficos e análises físico-químicas para determinação do pH e detecção de amônia e gás sulfídrico. Foram utilizadas 45 amostras de filé de pintado, com aproximadamente 100 gramas cada, embaladas individualmente em polietileno de alta densidade e armazenadas em câmara frigorífica entre 0ºC e 3ºC. A cada dois a três dias de estocagem, três unidades de filés foram submetidas a análises microbiológicas e físico-químicas, totalizando 15 análises durante o período de estocagem. As contagens de Aeromonas sp. e microrganismos heterotróficos aeróbios psicrotróficos variaram de 1,89 a 9,47logUFC/g e 0 a 6,54logUFC/g, respectivamente. A variação do pH foi de 6,20 a 6,97, e as análises de amônia e gás sulfídrico foram negativas durante todo o período. O pH dos filés de pintado atingiu o limite máximo de 6,4 aos 23 dias de estocagem, e estimou-se o seu prazo de validade comercial.

This word studied the occurrence of bacteria from genus Aeromonas and estimate the shelf life of "pintado" fish fillets (Pseudoplatystoma coruscans), during cold storage, through the quantification of psychrotrophic aerobic microorganisms, physical and chemical analyses for presence of ammonia and gas sulphide (H2S) and pH as used in 45 samples of "pintado" fillets with approximately 100 grams each, individually packed in high density polyethylene and stored in cold storage (0ºC to 3ºC). Every 2-3 days of storage, 3 units of fillets were subjected to microbiological and physicochemical analysis for a total of 15 days during the storage period. The Aeromonas sp. and psychrotrophic microorganisms count varied from 1.89 to 9.47logCFU/g and 0 to 6.54logCFU/g, respectively. The pH variation was from 6.20 to 6.97 and ammonia and H2S analyses were negative during the whole period. According under Brazilian legislation (Brazil, 1981) estimated the commercial shelf life of "pintado" fillets being 23 days when the pH reached a value of 6.4. The pH of the "pintado" fillets reached the maximum limit of 6.4 at 23 days of storage, being its estimated commercial shelf life.

Molecular cloning, characterization and enzymatic properties of a novel βeta-agarase from a marine isolate Psudoalteromonas sp. AG52

An agar-degrading Pseudoalteromonas sp. AG52 bacterial strain was identified from the red seaweed Gelidium amansii collected from Jeju Island, Korea. A β-agarase gene which has 96.8 percent nucleotide identity to Aeromonas β-agarase was cloned from this strain, and was designated as agaA. The coding region is 870 bp, encoding 290 amino acids and possesses characteristic features of the glycoside hydrolase family (GHF)-16. The predicted molecular mass of the mature protein was 32 kDa. The recombinant β-agarase (rAgaA) was overexpressed in Escherichia coli and purified as a fusion protein. The optimal temperature and pH for activity were 55 ºC and 5.5, respectively. The enzyme had a specific activity of 105.1 and 79.5 unit/mg toward agar and agarose, respectively. The pattern of agar hydrolysis demonstrated that the enzyme is an endo-type β-agarase, producing neoagarohexaose and neoagarotetraose as the final main products. Since, Pseudoalteromonas sp. AG52 encodes an agaA gene, which has greater identity to Aeromonas β-agarase, the enzyme could be considered as novel, with its unique bio chemical characteristics. Altogether, the purified rAgaA has potential for use in industrial applications such as development of cosmetics and pharmaceuticals.

Molecular cloning, characterization and enzymatic properties of a novel ßeta-agarase from a marine isolate Psudoalteromonas SP. AG52.

An agar-degrading Pseudoalteromonas sp. AG52 bacterial strain was identified from the red seaweed Gelidium amansii collected from Jeju Island, Korea. A ß-agarase gene which has 96.8% nucleotide identity to Aeromonas ß-agarase was cloned from this strain, and was designated as agaA. The coding region is 870 bp, encoding 290 amino acids and possesses characteristic features of the glycoside hydrolase family (GHF)-16. The predicted molecular mass of the mature protein was 32 kDa. The recombinant ß-agarase (rAgaA) was overexpressed in Escherichia coli and purified as a fusion protein. The optimal temperature and pH for activity were 55 °C and 5.5, respectively. The enzyme had a specific activity of 105.1 and 79.5 unit/mg toward agar and agarose, respectively. The pattern of agar hydrolysis demonstrated that the enzyme is an endo-type ß-agarase, producing neoagarohexaose and neoagarotetraose as the final main products. Since, Pseudoalteromonas sp. AG52 encodes an agaA gene, which has greater identity to Aeromonas ß-agarase, the enzyme could be considered as novel, with its unique bio chemical characteristics. Altogether, the purified rAgaA has potential for use in industrial applications such as development of cosmetics and pharmaceuticals.

Molecular cloning, characterization and enzymatic properties of a novel eta-agarase from a marine isolate Psudoalteromonas sp. AG52

An agar-degrading Pseudoalteromonas sp. AG52 bacterial strain was identified from the red seaweed Gelidium amansii collected from Jeju Island, Korea. A -agarase gene which has 96.8% nucleotide identity to Aeromonas -agarase was cloned from this strain, and was designated as agaA. The coding region is 870 bp, encoding 290 amino acids and possesses characteristic features of the glycoside hydrolase family (GHF)-16. The predicted molecular mass of the mature protein was 32 kDa. The recombinant -agarase (rAgaA) was overexpressed in Escherichia coli and purified as a fusion protein. The optimal temperature and pH for activity were 55 ºC and 5.5, respectively. The enzyme had a specific activity of 105.1 and 79.5 unit/mg toward agar and agarose, respectively. The pattern of agar hydrolysis demonstrated that the enzyme is an endo-type -agarase, producing neoagarohexaose and neoagarotetraose as the final main products. Since, Pseudoalteromonas sp. AG52 encodes an agaA gene, which has greater identity to Aeromonas -agarase, the enzyme could be considered as novel, with its unique bio chemical characteristics. Altogether, the purified rAgaA has potential for use in industrial applications such as development of cosmetics and pharmaceuticals.

Patógenos isolados do trato gastrintestinal de cães saudáveis no Rio de Janeiro / Pathogens isolated from gastrointestinal tract of healthy dogs in Rio de Janeiro

A total of 30 stool samples from 13 healthy dogs were analyzed in order to detect microorganisms in their gastrointestinal tract. The samples were enriched in alkaline peptone water added with 1 per cent and 3 per cent sodium chloride and Rappaport-Vassiliadis enrichment broth (37ºC/18-24h). Then, they were streaked onto Pseudomonas-Aeromonas selective agar base, tiossulfate citrate bile sucrose agar, eosin methylene blue agar, and Hektoen enteric agar. After the biochemical characterization, the results showed 41 bacterial isolated strains, from which Escherichia coli, Vibrio fluvialis, and Aeromonas caviae were the main pathogens encountered.(AU)

Bactérias do gênero Aeromonas em água de matadouro bovino

The aim of this work was to verify the occurrence of Aeromonas bacteria in samples of water (supply and residual) collected at beef slaughterhouse. Water used at the internal facilities, water from corrals, drinking water, water for pre-hygiene and tranquilization of the animals and residual water from carcasses wash, were analyzed. From those 30 samples of each type, Aeromonas bacteria were isolated in 10 (33.3%) samples of corral water and in 10 (33.3%) samples of residual water from carcasses wash. None of the facilities treated water supply samples showed positive in isolation. The isolated species were Aeromonas hydrophila in two (2.2%) and Aeromonas caviae in 19 (21.1%) of the samples. One non-typical strain was isolated from corral water. The results demonstrated that corral water may be an important contamination source, mainly to the hide and, through it, Aeromonas sp. can reach the slaughter room.

Verificou-se a ocorrência de bactérias do gênero Aeromonas em amostras de água (abastecimento/residuária) obtidas em matadouro bovino. Analisaram-se a água utilizada nas dependências internas, a água dos currais, utilizada na dessedentação, pré-higienização e tranqüilização dos animais e a água residuária da lavagem das carcaças. Das 30 amostras representativas de cada tipo, bactérias do gênero Aeromonas foram isoladas em 10 (33,3%) amostras da água dos currais e em 10 (33,3%) amostras da água residuária da lavagem de carcaças. Nenhuma das amostras da água tratada de abastecimento das instalações revelou-se positiva no isolamento. As espécies isoladas foram Aeromonas hydrophila em duas (2,2%) e Aeromonas caviae em 19 (21,1%) amostras. Uma cepa considerada atípica foi isolada da água dos currais. Os resultados evidenciaram que a água dos currais pode ser uma importante fonte de contaminação, principalmente para a pele, e através dela as Aeromonas sp. podem chegar à sala de matança.

Prevalência de Aeromonas spp. em fezes diarréicas de crianças menores de 5 anos de idade na cidade de Goiânia, Goiás, no biênio 1995-1996 / The prevalence of Aeromonas spp. in the diarrheal feces of children under the age of 5 years in the city of Goiânia, Goiás in the 1995-1996 biennium

Foram analisadas 163 amostras de fezes de crianças com idade abaixo de 5 anos no período de 1995 a 1996, sendo 91 de fezes diarréicas e 72 de fezes não diarréicas. O material foi coletado em meio para transporte e submetido ao processo de enriquecimento a 4oC por 7 dias. Para o isolamento primário foi utilizado ágar amido ampicilina e incubado a 35oC por 18 a 24 horas. Foram isoladas 20 (21,9%) das seguintes espécies: Aeromonas A. caviae (7,7%), A. salmonicida salmonicida (6,6%), A. sobria (4,3%), A. hydrophila (2,2%) e Salmonicida achromogenes (1,1%). Nenhuma Aeromonas spp foi isolada dos 72 pacientes-controles. A susceptibilidade das amostras de Aeromonas spp aos antimicrobianos foi maior com a ciprofloxacina, diminuindo gradativamente com cloranfenicol, gentamicina, ampicilina e eritromicina.

From 1995 through 1996, 163 fecal specimens of children aged under 5 years were analysed, 91 being from diarrhea feces and 72 without diarrhea. The material was collected in transport medium and submitted to the enrichment procedure at 4 degrees C for 7 days. For the primary isolation starch ampicillin agar was used and incubated at 35 degrees C for 18 to 24 hours. Twenty (20.9%) from the following specimens were isolated: Aeromonas (A.) caviae (7.7%), A. salmonicida salmonicida (6.6%), A. sobria (4.3%), A. hydrophila (2.2%) and Salmonicida achromogenes (1.1%). No Aeromonas spp. was isolated from the 72 control subjects. The Aeromonas spp. susceptibility to antimicrobial was greater with ciprofloxacin, being this susceptibility gradually diminished with chloranphenicol, gentamicin, ampicillin and erythromycin.