Implication for Functions of the Ectopic Adipocyte Copper Amine Oxidase (AOC3) from Purified Enzyme and Cell-Based Kinetic Studies

AOC3 is highly expressed in adipocytes and smooth muscle cells, but its function in these cells is currently unknown. The in vivo substrate(s) of AOC3 is/are also unknown, but could provide an invaluable clue to the enzyme's function. Expression of untagged, soluble human AOC3 in insect cells provides a relatively simple means of obtaining pure enzyme. Characterization of enzyme indicates a 6% titer for the active site 2,4,5-trihydroxyphenylalanine quinone (TPQ) cofactor and corrected kcat values as high as 7 s−1. Substrate kinetic profiling shows that the enzyme accepts a variety of primary amines with different chemical features, including nonphysiological branched-chain and aliphatic amines, with measured kcat/Km values between 102 and 104 M−1 s−1. Km(O2) approximates the partial pressure of oxygen found in the interstitial space. Comparison of the properties of purified murine to human enzyme indicates kcat/Km values that are within 3 to 4-fold, with the exception of methylamine and aminoacetone that are ca. 10-fold more active with human AOC3. With drug development efforts investigating AOC3 as an anti-inflammatory target, these studies suggest that caution is called for when screening the efficacy of inhibitors designed against human enzymes in non-transgenic mouse models. Differentiated murine 3T3-L1 adipocytes show a uniform distribution of AOC3 on the cell surface and whole cell Km values that are reasonably close to values measured using purified enzymes. The latter studies support a relevance of the kinetic parameters measured with isolated AOC3 variants to adipocyte function. From our studies, a number of possible substrates with relatively high kcat/Km have been discovered, including dopamine and cysteamine, which may implicate a role for adipocyte AOC3 in insulin-signaling and fatty acid metabolism, respectively. Finally, the demonstrated AOC3 turnover of primary amines that are non-native to human tissue suggests possible roles for the adipocyte enzyme in subcutaneous bacterial infiltration and obesity

Serotypes, virulence factors, antibiotic sensitivity, beta-lactamase activity and plasmid analysis of Salmonella from children with diarrhea in Tripoli (Libya)

A total of 21 Salmonella strains isolated in Libya (16 from children with diarrhea and 5 from healthy controls) were serotyped and studied for their cell invasive ability, production of cytotoxin, antibiotic susceptibility, β-lactamase activity and plasmid profiles. Eight different serotypes of Salmonella were identified: 6 S. saintpaul , 4 S. wien (1 from control), 2 S. newport , 2 S. muenchen (1 from contr ol), 2 S. typhimurium (1 from control) , 2 S. hadar (1 from control), 2 S. reading (1 from control), 1 S. kottbus . Twenty (95%) were positive in the invasiveness assay using HeLa cells, and all (100%) were negative for cytotoxin production in HT29 cells. More than 40% were resistant to ampicillin, cefalexin, cefam andole, cefoperazone, chloramphenicol, gentamicin, mezlocillin and trimethoprim- sulphamethoxazole and 100% were susceptible to the new quinolones. Most (67%) of the strains harbored plasmids and 43% produced β-lactamase. A strong association was observed between the presence of more than one plasmid, β-lactamase activity, and multiple-resistance to antimicrobial agents and serotypes S. saintpa ul and S. wien . Curing experiments with acridine orange showed that 2 plasmids (33 and 1.4 megadaltons ) might be responsible for the resistance to chloramphenicol and gentamicin. The present study demonstrated that multiple-resistant salmonellae are widesp read in Libya and the resistance is mainly plasmid mediated

Identificação e resistência a antimicrobianos de espécies de Aeromonas móveis isoladas de peixes e ambientes aquáticos Identification and antimicrobial resistance of motile Aeromonas isolated from fish and aquatic environment

Com o objetivo de se verificar a diversidade de espécies de Aeromonas móveis e seu perfil de susceptibilidade a antimicrobianos em pisciculturas comerciais, foram selecionadas oito tilapiculturas localizadas na região do Alto Rio Grande, Minas Gerais. De cada propriedade foram coletadas três amostras de peixes em estádio de pré-abate (vivos e saudáveis), uma amostra de água do tanque e uma amostra da água de abastecimento do sistema. De cada peixe foram coletadas amostras de lavado superficial e do parênquima renal. Diluições seriadas adequadas de cada amostra foram plaqueadas em TSA-ampicilina (10 mg/l) e as amostras de rim em Ágar Sangue de cavalo a 5%. A partir de colônias isoladas positivas para o teste da oxidase foram realizados testes para identificação do gênero (testes presuntivos) e das espécies de Aeromonas (testes bioquímicos). O perfil de antibiograma foi obtido pelo teste de difusão de discos de antibióticos em Ágar Mueller Hinton. Foram obtidos 75 isolados diferenciados em nove espécies de Aeromonas: A. jandaei, A. hydrophila, A. trota, A. caviae, A. sobria, A. eucrenophila, A. veronii bt veronii, A. schubertii, A. media, além de amostras classificadas como Aeromonas atípicas. Do total isolado, oito amostras foram provenientes da superfície corpórea de peixes, 14 da água de abastecimento e 53 da água do tanque. Não houve isolamento a partir dos espécimes de parênquima renal. Em relação ao perfil de resistência, 93% dos isolados foram resistentes à eritromicina, 36% à tetraciclina, 13% ao ác. nalidíxico, 9% à gentamicina, 8% à nitrofurantoína, 8% à canamicina, 5% à norfloxacina, 4% ao cloranfenicol e 3% às sulfonamidas. Dentre os isolados analisados, 43% apresentaram índice de múltipla resistência a antimicrobianos (MAR) igual ou superior a 22%, ou seja, resistência a dois ou mais drogas das nove testadas. Os dados apontam para um risco iminente, tanto pelo isolamento de amostras potencialmente patogênicas a seres humanos quanto pelo perfil de multirresistência dos isolados.<br>This experiment was done with the objective to verify the diversity of motile Aeromonas species and their antibiotic susceptibility profiles. A total of eight commercial producers of tilapia located in the Alto Rio Grande region of Minas Gerais state were selected for this experiment. The collected samples were three fish, one sample of pond water and one sample of water used in filling the pond. Suitable dilutions of the washed surface of the fish and water samples were scattered in TSA supplemented with ampicilin (10mg/l). Kidney samples were streaked in Horse Blood Agar. Isolated colonies oxidase positive, were submitted of tests for identification of the species of Aeromonas (presumptive and biochemical tests). The antibiogram profile of the isolated samples was obtained through a diffusion test of antibiotic disks in Mueller Hinton Agar. Were obtained 75 different isolates in nine species of Aeromonas: A. jandaei, A. hydrophila, A. trota, A. caviae, A. sobria, A. eucrenophila, A. veronii bv veronii, A. schubertii, A. media, and samples classified like atypical Aeromonas. Of the total isolations, eight samples were taken from the corporal surface of the fish, 14 from the water source and 53 from the pond water. There was no isolation of the renal tissue. In relation to the resistance profile, 93% of the isolated samples were resistant to erythromycin, 36% to tetracycline, 13% to nalidixic acid, 9% to gentamicin, 8% to nitrofurantoin, 8% to kanamycin, 5% to norfloxacin, 4% to chloramphenicol and 3% to sulfonamides. Among the isolated samples analyzed, 42,66% presented a MAR index (Multiple Antibiotic Resistance Index) equal to or superior to 22,2%. It was concluded that the isolation of potentially pathogenic bacterial species, with multiresistant antibiotic traits may be a risk to human food chain

The occurrence of Aeromonas spp. in the bottled mineral water, well water and tap water from the municipal supplies

The aim of this work was to study the occurrence of Aeromonas sp in the bottled mineral water, well water and tap water from the municipal supplies. Positive samples were found for Aeromonas spp. 12.7% from the mineral water, 8.3% from the artesian water and 6.5% from the tap water. The recovery of Aeromonas spp. was significantly higher in the bottled mineral and artesian water than in the tap water from municipal supplies. The occurrence of the Aeromonas spp. did not correlate significantly with the contamination indicator bacteria (i.e. total coliforms) in the artesian water samples. However, a significant correlation was found between Aeromonas spp. and total coliforms in the both mineral water and tap water samples. The presence or absence of a correlation between the indicator bacteria and Aeromonas could reflect the occasional appearance of the pathogen in the drinking water and the different rates of survival and recovery of these agents compared with those fecal indicators. The finding that 41.6, 14.8 and 9.0 % of the artesian water, bottled mineral water and tap water, respectively, sampled in the current study failed to meet the Brazilian standard for total coliforms in the drinking water should therefore be of concern.<br>A porcentagem de amostras positivas para Aeromonas foi de 12.7% para água mineral, 8.3% para água de poço artesiano e 6.5% para água do sistema público de abastecimento. O isolamento de Aeromonas spp. foi significativamente maior em água mineral e água de poço artesiano do que em água do sistema público. A ocorrência de Aeromonas spp. não teve correlação significativa com os indicadores de contaminação tradicionalmente utilizados (coliformes totais) em amostras de água de poço artesiano. No entanto, esta correlação foi positiva e significativa em água mineral e água do sistema público. A presença ou ausência de correlação entre bactérias indicadoras e a presença de Aeromonas pode refletir o aparecimento ocasional de patógenos em águas para consumo humano e as diferentes taxas de sobrevivência e isolamento destes agentes comparados com os indicadores fecais de contaminação. A constatação de que 41.6%, 14.8% e 9.0% respectivamente amostras de água de poço, água mineral e água do abastecimento público utilizadas neste estudo apresentaram índices de coliformes maiores do que os aceitáveis pela legislação brasileira é um fato preocupante. Estes números mostram a necessidade de melhoria nos sistemas de monitoramento para a indústria de águas minerais e o sistema público de abastecimento. As cepas isoladas pertencentes ao gênero Aeromonas foram identificadas ao nível de espécie como A. hydrophila e A. jandaei. A significância do grande número de isolamentos de espécies de Aeromonas em saúde pública ainda não está clara. É necessário o estudo dos efeitos de cepas específicas utilizando modelos animais de infecção. Estes resultados podem contribuir para a reavaliação dos critérios empregados para a análise da qualidade microbiológica da água e a definição de limites de densidades para o gênero Aeromonas em águas destinadas ao consumo humano

Global Review of the Age Distribution of Rotavirus Disease in Children Aged < 5 Years Before the Introduction of Rotavirus Vaccination

International audienceWe sought datasets with granular age distributions of rotavirus-positive disease presentations among children <5 years of age, before the introduction of rotavirus vaccines. We identified 117 datasets and fit parametric age distributions to each country dataset and mortality stratum. We calculated the median age and the cumulative proportion of rotavirus gastroenteritis events expected to occur at ages between birth and 5.0 years. The median age of rotavirus-positive hospital admissions was 38 weeks (interquartile range [IQR], 25-58 weeks) in countries with very high child mortality and 65 weeks (IQR, 40-107 weeks) in countries with very low or low child mortality. In countries with very high child mortality, 69% of rotavirus-positive admissions in children <5 years of age were in the first year of life, with 3% by 10 weeks, 8% by 15 weeks, and 27% by 26 weeks. This information is critical for assessing the potential benefits of alternative rotavirus vaccination schedules in different countries and for monitoring program impact