Resistência aos antimicrobianos e diversidade genética de variantes fenotípicas de Pseudomonas aeruginosa não sensíveis aos carbapenêmicos recuperadas de pacientes com fibrose cística com infecção pulmonar crônica: uma análise temporal / Antimicrobial resistance and genetic diversity of phenotypic variants of carbapenem non susceptible Pseudomonas aeruginosa recovered from patients with cystic fibrosis with chronic lung infection: a temporal analysis

Pseudomonas aeruginosa, bactéria ubíqua e versátil, pode se comportar como um patógeno oportunista, com ampla capacidade adaptativa, por múltiplos fatores de virulência e resistência. Como agente patogênico nas infecções pulmonares em pacientes com fibrose cística (FC), é motivo de prognóstico ruim, aumento de hospitalizações e altas taxas de morbimortalidade, sendo quase impossível a sua erradicação, ao evoluírem para a cronicidade. Globalmente, é notável o aumento nos índices de amostras não sensíveis aos carbapenêmicos e a múltiplos antimicrobianos, essenciais à terapêutica. Assim, avaliamos temporalmente a susceptibilidade aos antimicrobianos e a presença de amostras hipermutáveis (HPM) em P. aeruginosa de diferentes morfotipos, não sensíveis aos carbapenêmicos (PANSC), obtidas de pacientes FC com infecção pulmonar crônica, acompanhados em dois centros de referência no Rio de Janeiro. De 2007 a 2016, a análise retrospectiva, através dos resultados obtidos no teste de disco-difusão (TDD), permitiu selecionar amostras de PANSC incluídas neste trabalho. Usando os resultados obtidos no TDD, foi definida a susceptibilidade a outros antimicrobianos, bem como os fenótipos de resistência, multi-(MDR), extensivo-(XDR) e pandroga resistentes (PDR). Adicionalmente, determinou-se a concentração inibitória mínima (CIM) para imipenem (IPM), meropenem (MEM), doripenem (DOR) e polimixina (POL). Através de teste fenotípico, foi calculada a frequência de mutação espontânea e as amostras hipermutáveis foram caracterizadas. O sequenciamento de genoma total (SGT) foi realizado em seis amostras de diferentes morfotipos, incluindo uma variante fenotípica rara, a small colony variant (SCV). Essas amostras foram recuperadas em dois episódios de exacerbação do paciente. Foram investigadas a clonalidade, resistência a antimicrobianos e virulência. Das 143 amostras, de 18 pacientes (9 pediátricos e 9 adultos), os resultados do TDD apontaram taxas de não susceptibilidade superiores a 44% para gentamicina, amicacina, tobramicina e ciprofloxacina, e maiores de 30 % para POL. Pela determinação da CIM, quase a totalidade (96%) das amostras foram não sensíveis a IMP, seguidos de 56% para MEM e 44% para DOR. Analisando-se a distribuição dos valores da CIM50 e CIM90 nos dois grupos de pacientes, os valores para IMP foram maiores entre as amostras dos pacientes pediátricos, equivalendo a 32 µg/mL e 64 µg/mL, respectivamente. Cerca de 25%, 37% e 6% eram MDR, XDR e PDR, respectivamente. Aproximadamente 12% eram HPM, e mais da metade destas foram XDR. Após o SGT, as seis amostras, recuperadas do caso clínico foram classificadas em um novo sequence type (ST2744), com a presença de genes de resistência adquiridos blaPAO, blaOXA-50, aph(3')-Iib, fosA, catB7 e crpP, apresentando mutações em genes codificadores de porinas e bombas de efluxo. Entretanto, não foram observados marcadores genéticos clássicos exclusivos para os fenótipos SCV e HPM. Este é o primeiro relato de P. aeruginosa SCV na FC, no Brasil. A vigilância epidemiológica de P. aeruginosa é crucial para a conduta terapêutica, bem como para o sucesso da resposta do paciente e erradicação da infecção pulmonar, justificando o uso de técnicas fenotípicas e moleculares na detecção dos mecanismos de resistência e virulência desse microrganismo na FC.

Pseudomonas aeruginosa, a ubiquitous and versatile bacterium, can behave as an opportunistic pathogen, with strong adaptive capacity, due to multiple virulence and resistance factors. As a pulmonary infection pathogen in patients with cystic fibrosis (CF), it is related with poor prognosis, increased hospitalizations and high rates of morbidity and mortality, and the eradication is almost impossible, especially after chronicity. The increase rates of isolates non-susceptible to carbapenem and multiple antimicrobials, essentials to therapy, have been observed worldwide. Therefore, we assessed the antimicrobial susceptibility and the presence of hypermutability (HPM) in non-susceptible to carbapenem P. aeruginosa (PANSC) isolates from different morphotypes, obtained from CF patients with chronic pulmonary infection, followed at two reference centers in Rio de Janeiro. Using the results obtained by disk-diffusion test (DDT) between 2007 to 2016, we select 143 PANSC and susceptibility to other antimicrobials was defined, as well as the resistance phenotypes, multi- (MDR), extensive- (XDR) and pandrug resistant (PDR). Additionally, the minimum inhibitory concentration (MIC) for imipenem (IPM), meropenem (MEM), doripenem (DOR) and polymyxin (POL) was determined. Hypermutable isolates were characterized by determination of mutation frequency. Whole genome sequencing (WGS) was performed in six morphotypes isolates, including the small colony variant (SCV), a rare variant phenotype. These isolates were recovered in two exacerbation episodes. Clonality, antimicrobial resistance and virulence were investigated. Of the total (143 isolates) isolated from 18 patients (9 pediatric and 9 adults), non-susceptibility rates above than 44% for gentamicin, amikacin, tobramycin and ciprofloxacin, and more than 30% for POL were observed. Almost all (96%) of the isolates were non-susceptible to IPM by MIC determination, followed by 56% for MEM and 44% for DOR. MIC50 (32 µg/mL) and MIC90 (64 µg/mL) rates for IPM were higher among pediatric patient isolates and 25%, 37% and 6% were MDR, XDR and PDR, respectively. 12% of all isolates were classified as HPM and more than half were categorized as XDR. Using WGS, the six isolates recovered from the clinical case, were identified as a new sequence type (ST2744). Acquired resistance genes blaPAO, blaOXA-50, aph (3')-Iib, fosA, catB7 and crpP and mutations in encoding genes for porins and efflux pumps, was annotated. None exclusive classic genetic markers related to SCV and HPM phenotypes were not observed. This is the first Brazilian report of P. aeruginosa SCV in CF. Our results highlight the importance of epidemiological surveillance in P. aeruginosa. The application of phenotypic and molecular techniques to investigate resistance and virulence mechanisms, can contribute to therapeutic success in CF.

A modified gentamicin protection assay for detecting invasive phenotype of Salmonella enterica serovar Enteritidis / 生物工程学报

Salmonella enterica serovar Enteritidis (SE) is one of the most important zoonotic pathogens that cause enteritis and systemic infection in animals and human. Understanding invasive capacities of SE isolates is of vital importance to elucidate pathogenesis of Salmonella infection. To improve the throughput capacity and repeatability of classical gentamicin protection assay (GPA), a modified PGA was developed by taking high-throughput advantage of 96-well cell plates and multichannel pipettes. In addition, drop plate technique rather than spread plate method was applied in the modified GPA protocol for bacterial enumeration. The modified GPA protocol was evaluated by phenotyping intracellular replication of a high virulent and a low virulent SE isolates, JL228 and LN248, in a phagocytic cell line RAW264.7. The protocol was then applied in invasive phenotype determination of 16 SE strains to non-phagocytes (HT-29) and the intracellular replication of 43 SE strains to phagocytes (RAW264.7). Significant lower intra-group and inter-group coefficient of variations of the modified GPA was observed, implying good repeatability and reproducibility over traditional protocol. Further, replication phenotypes were also correlated with those from direct observation by confocal microscopy. Collectively, the improved GPA protocol had advantages of high throughput capacity, good repeatability and reliability, it was also noticed that the protocol also represented a fast and labor-saving alternative scheme for the invasive phenotype determination of Salmonella Enteritidis, and providing reliable phenotype profiles for Salmonella-host interplay interpretation.

Actividad sinérgica in vitro de los alcaloides Crotsparina y Esparsiflorina aislados de Croton bomplandianum frente a Pseudomonas aeruginosa / In vitro synergic activity of the alkaloids Crotsparine and Sparsiflorine, isolated from Croton bomplandianum, against Pseudomonas aeruginosa

Se evaluó la actividad sinérgica de los alcaloides crotsparina y esparsiflorina, aislados de Croton bomplandianum Baill. con los antibacterianos gentamicina y ciprofloxacina frente a Pseudomonas aeruginosa, microorganismo frecuentemente responsable de infecciones intrahospitalarias. Se empleó el método del "tablero de damas". Se encontraron combinaciones que presentaban efecto sinérgico, logrando la reducción de 87,5% de la CMI de gentamicina, mientras que para ciprofloxacina se logró una reducción del 25,0%. Esto abre interesantes perspectivas sobre el uso combinado de productos naturales puros y fármacos en uso clínico para el tratamiento de infecciones producidas por este microrganismo(AU)

Nephroprotective property of Cinnamomum zeylanicum and its antibacterial activity in combination with gantamicin

Cinnamomum zeylanicum has strong antioxidant properties and has been presented to have nephroprotective effects. Present work was aimed to study the nephroprotective property of the plant extract through urinary enzymes excretion, to confirm its protective effects and to observe the antibacterial activities of gentamicin in combination with the plant extract. 200mg/kg/day of the plant extracts were administered alone and as co-therapy with gentamicin. Urinary lactate dehydrogenase [LDH] and Urinary alkaline phospatase [ALP] excretions were observed through reagents kits with the help of Power-Lab 300. Antibacterial activities were assessed for gentamicin alone and in combination with the extract. Present study showed that the plant extract have excess quantity of flavonoids, which may responsible for attenuating the excessive excretion of urinary LDH. However, Urinary ALP excretion was found remained same throughout the study period in all experimental groups; might be detected in acute damage. Further, the plant also proved to have no decreasing impact on the antibacterial activities of gentamicin

Staphylococcus aureus resistance to topical antimicrobials in atopic dermatitis

Abstract: Background: Topical antimicrobial drugs are indicated for limited superficial pyodermitis treatment, although they are largely used as self-prescribed medication for a variety of inflammatory dermatoses, including atopic dermatitis. Monitoring bacterial susceptibility to these drugs is difficult, given the paucity of laboratory standardization. Objective: To evaluate the prevalence of Staphylococcus aureus topical antimicrobial drug resistance in atopic dermatitis patients. Methods: We conducted a cross-sectional study of children and adults diagnosed with atopic dermatitis and S. aureus colonization. We used miscellaneous literature reported breakpoints to define S. aureus resistance to mupirocin, fusidic acid, gentamicin, neomycin and bacitracin. Results: A total of 91 patients were included and 100 S. aureus isolates were analyzed. All strains were methicillin-susceptible S. aureus. We found a low prevalence of mupirocin and fusidic acid resistance (1.1% and 5.9%, respectively), but high levels of neomycin and bacitracin resistance (42.6% and 100%, respectively). Fusidic acid resistance was associated with more severe atopic dermatitis, demonstrated by higher EASI scores (median 17.8 vs 5.7, p=.009). Our results also corroborate the literature on the absence of cross-resistance between the aminoglycosides neomycin and gentamicin. Conclusions: Our data, in a southern Brazilian sample of AD patients, revealed a low prevalence of mupirocin and fusidic acid resistance of S. aureus atopic eczema colonizer strains. However, for neomycin and bacitracin, which are commonly used topical antimicrobial drugs in Brazil, high levels of resistance were identified. Further restrictions on the use of these antimicrobials seem necessary to keep resistance as low as possible.

A Simple, Reproducible, Inexpensive, Yet Old-Fashioned Method for Determining Phagocytic and Bactericidal Activities of Macrophages

Macrophages (Mphi) play a pivotal role in the protection system by recognizing and eliminating invading pathogenic bacteria. Phagocytosis and the killing of invading bacteria are major effector functions of Mphi. Although the phagocytic and bactericidal activities of Mphi have been analyzed via several methods using a light microscope, a fluorescence microscope, or a fluorescence-activated cell sorter, expensive materials and equipment are usually required, and the methods are rather complicated. Moreover, it is impossible to determine both the phagocytic and bactericidal activities of Mphi simultaneously using these methods. In this review, we describe a simple, reproducible, inexpensive, yet old-fashioned method (antibiotic protection assay) for determining the phagocytic and bactericidal activities of Mphi.

Clinical pharmacology of gentamicin in neonates: regimen, toxicology and pharmacokinetics / Farmacologia clínica da gentamicina em neonatos: modalidades de uso, toxicologia e farmacocinética

Gentamicin is an aminoglycoside antibiotic. It kills bacteria by inhibiting protein synthesis and to some extent by lysing the cell envelope. Gentamicin is frequently the first choice drug because of its reliability, but also because of the long experience with its use. In combination with β-lactam antibiotics it is recommended for the treatment of sepsis or pneumonia and is active against P. aeruginosa, Enterobacter, Klebsiella and Serratia. However, gentamicin is ototoxic and nephrotoxic. The human mitochondrial genetic variant m.1555A > G has been reported to be an important cause of non-syndromic hereditary hearing dysfunction and may cause permanent hearing loss. Even short courses of gentamicin therapy in healty newborn infants can lead to abnormalities of auditory function. It is active against very resistant bacteria at peak concentrations (> 10 mg/l) that are high enough to be potentially toxic. For safe therapeutic efficacy, peak plasma concentrations of gentamicin should range from 4 to 10 mg/l; but trough concentrations, immediately before a new drug administration, must be lower that 2 mg/L to avoid toxic effects. Pharmacokinetic parameters vary considerably in infants. Half-life ranges from 5.4 to 10.0 hours, clearance 0.50 to 1.71 ml/h/kg and distribution volume from 0.4 to 0.7 l/kg. Preterm infants have a longer half-life than full-term infants. Thus, it is mandatory to monitor gentamicin serum concentrations whenever infants are treated for 48 hours or more.

A gentamicina é antibiótico do grupo dos aminoglicosídeos. Destrói bactérias por inibição de síntese proteica e, em certa medida, por lise do envelope celular. A gentamicina é a droga de primeira escolha por causa de sua atividade confiável e em virtude de longa experiência com seu uso. Em combinação com antibióticos β-lactâmicos é recomendada para o tratamento de septicemia ou pneumonia e é ativa contra P. aeruginosa, Enterobacter, Klebsiella e Serratia. No entanto, a gentamicina é ototóxica e nefrotóxica. A variante genética mitocondrial humana m.1555A > G é tida como importante causa de disfunção auditiva hereditária não-sindrômica e pode causar perda permanente da audição. Até mesmo procedimentos terapêuticos de gentamicina de curta duração em recém-nascidos sadios podem levar a anormalidades da função auditiva. É ativa contra algumas espécies de bactérias apenas em concentrações de pico (> 10 mg/l) que são suficientemente altas para produzirem efeitos tóxicos. A gentamicina deve cair a concentrações mínimas menores que 2 mg/l para evitar efeitos tóxicos. Para produzir efeitos terapêuticos, as concentrações plasmáticas máximas de gentamicina deve variar de 4 a 10 mg/l. Os parâmetros farmacocinéticos variam consideravelmente em lactentes. A meia-vida varia entre 5,4-10,0 horas, o "clearance" varia entre 0,50 e 1,71 ml/h/kg e volume de distribuição de 0,4-0,7 l/kg. Em prematuros a meia-vida é mais longa do que a de crianças nascidas a termo. Por esses motivos, sempre que lactentes são tratadas durante 48 horas ou mais, monitorizar as concentrações séricas de gentamicina é essencial.

Invasin gimB found in a bovine intestinal Escherichia coli with an adherent and invasive profile

The invasin gimB (genetic island associated with human newborn meningitis) is usually found in ExPEC (Extraintestinal Pathogenic Escherichia coli) such as UPEC (uropathogenic E. coli), NMEC (neonatal meningitis E. coli) and APEC (avian pathogenic E. coli). In NMEC, gimB is associated with the invasion process of the host cells. Due to the importance of E. coli as a zoonotic agent and the scarce information about the frequency of gimB-carrying strains in different animal species, the aim of this study was to investigate the presence of gimB in isolates from bovine, swine, canine and feline clinical samples. PCR was conducted on 196 isolates and the identity of the amplicons was confirmed by sequencing. Of the samples tested, only E. coli SB278/94 from a bovine specimen was positive (1/47) for gimB, which represents 2.1% of the bovine isolates. The ability of SB278/94 to adhere to and invade eukaryotic cells was confirmed by adherence and gentamicin-protection assays using HeLa cells. This is the first study that investigates for gimB in bovine, canine and feline E. coli isolates and shows E. coli from the intestinal-bovine samples harboring gimB.

Effect of Trimetaphosphate and Fluoride Association on Hydroxyapatite Dissolution and Precipitation In Vitro

The present study analyzed the action of sodium trimetaphosphate (TMP) and/or fluoride on hydroxyapatite. Hydroxyapatite powder was suspended in different solutions: deionized water, 500 µg F/mL, 1,100 µg F/mL, 1%TMP, 3%TMP, 500 µg F/mL plus 1%TMP and 500 µg F/mL plus 3%TMP. The pH value of the solutions was reduced to 4.0 and after 30 min, raised to 7.0 (three times). After pH-cycling, the samples were analyzed by X-ray diffraction and infrared spectroscopy. The concentrations of calcium fluoride, fluoride, calcium and phosphorus were also determined. Adding 1% or 3% TMP to the solution containing 500 µg F/mL produced a higher quantity of calcium fluoride compared to samples prepared in a 1,100 µg F/mL solution. Regarding the calcium concentration, samples prepared in solutions of 1,100 µg F/mL and 500 µg F/mL plus TMP were statistically similar and showed higher values. Using solutions of 1,100 µg F/mL and 500 µg F/mL plus TMP resulted in a calcium/phosphorus ratio close to that of hydroxyapatite. It is concluded that the association of TMP and fluoride favored the precipitation of a more stable hydroxyapatite.

O presente estudo avaliou a ação do trimetafosfato de sódio (TMP) e/ou fluoreto sobre a hidroxiapatita. Pó de hidroxiapatita foi suspenso em diferentes soluções: água deionizada, 500 µg F/mL, 1100 µg F/mL, 1%TMP, 3%TMP, 500 µg F/mL adicionado a 1%TMP e 500 µg F/mL associado a 3%TMP. O pH das soluções foi reduzido para 4,0 e depois de 30 min, elevado para 7,0 (três vezes). Depois do processo de ciclagem de pH, as amostras foram analisadas por difração de raios-X e espectroscopia por infravermelho. As concentrações de fluoreto de cálcio, fluoreto, cálcio e fósforo também foram determinadas. A adição de 1% ou 3% TMP na solução contendo 500 µg F/mL produziu uma maior quantidade de fluoreto de cálcio comparado às amostras tratadas com uma solução de 1100 µg F/mL. A respeito da concentração de cálcio, amostras tratadas com soluções de 1100 µg F/mL e 500 µg F/mL adicionado ao TMP foram estatisticamente similares e mostraram maiores valores. Soluções de 1100 µg F/mL e 500 µg F/mL adicionado ao TMP resultaram em uma proporção molar Ca/P mais próxima à da hidroxiapatita. Conclui-se que a associação de TMP e F favoreceu a precipitação de uma hidroxiapatita mais estável.

Apoptosis in HEp-2 cells infected with Ureaplasma diversum

BACKGROUND: Bacterial pathogens have many strategies for infecting and persisting in host cells. Adhesion, invasion and intracellular life are important features in the biology of mollicutes. The intracellular location ofUreaplasma diversum may trigger disturbances in the host cell. This includes activation or inhibition of pro and anti-apoptotic factors, which facilitate the development of host damage. The aim of the present study was to associate U. diversum infection in HEp-2 cells and apoptosis induction. Cells were infected for 72hs with four U. diversum clinical isolates and an ATCC strain. The U. diversuminvasion was analyzed by Confocal Laser Scanning Microscopy and gentamicin invasion assay. The apoptosis was evaluated using pro-apoptotic and anti-apoptotic gene expression, and FITC Annexin V/Dead Cell Apoptosis Kit. RESULTS: The number of internalized ureaplasma in HEp-2 cells increased significantly throughout the infection. The flow cytometry analysis with fluorochromes to detect membrane depolarization and gene expression for caspase 2, 3 and 9 increased in infected cells after 24 hours. However, after 72 hours a considerable decrease of apoptotic cells was observed. CONCLUSIONS: The data suggests that apoptosis may be initially induced by some isolates in association with HEp-2 cells, but over time, there was no evidence of apoptosis in the presence of ureaplasma and HEp-2 cells. The initial increase and then decrease in apoptosis could be related to bacterial pathogen-associated molecular pattern (PAMPS). Moreover, the isolates of U. diversum presented differences in the studied parameters for apoptosis. It was also observed that the amount of microorganisms was not proportional to the induction of apoptosis in HEp-2 cells.

High level of gentamicin resistance [HLGR] among enterococcus strains isolated from clinical specimens

Enterococci are pathogens that can cause nosocomial infections and acquire resistance properties via several molecular mechanisms. The aac [6']Ie-aph[2"]Ia gene plays a significant role in the emergence of high-level gentamicin-resistant [HLGR] strains. The screening of resistant strains and the provision of appropriate antibiotic therapy can decide the outcome of serious nosocomial infections. In the present study, 142 enterococci were isolated from patients, and the species were identified using standard methods. An antimicrobial susceptibility test was performed using the disc diffusion method, and the minimum inhibition concentration [MIC] of gentamicin was determined according to the broth micro-dilution method. Additionally, PCR was utilized to detect the aac[6']Ie-aph[2"]Ia gene, the presence of which was confirmed by digestion with Sca1 and sequencing. Of the 142 isolates, 62 [43.7%] were found to exhibit the HLGR phenotype. All except one of the HLGR isolates contained the aac[6']Ie-aph[2"]Ia gene. The prevalence of resistance to other antibiotics and multi-drug resistance [MDR] was higher among the HLGR isolates compared to the non-HLGR isolates. Our results indicate that high prevalence rates of MDR and HLGR enterococci are an important problem associated with medical treatment. Furthermore, the presence of the aacaacaac[6']Ie-aph[2"]Ia gene was shown to correspond to the presence of the HLGR phenotype among enterococci

Use of In Vivo and In Vitro Systems to Select Leishmania amazonensis Expressing Green Fluorescent Protein

Various Leishmania species were engineered with green fluorescent protein (GFP) using episomal vectors that encoded an antibiotic resistance gene, such as aminoglycoside geneticin sulphate (G418). Most reports of GFP-Leishmania have used the flagellated extracellular promastigote, the stage of parasite detected in the midgut of the sandfly vector; fewer studies have been performed with amastigotes, the stage of parasite detected in mammals. In this study, comparisons were made regarding the efficiency for in vitro G418 selection of GFP-Leishmania amazonensis promastigotes and amastigotes and the use of in vivo G418 selection. The GFP-promastigotes retained episomal plasmid for a prolonged period and G418 treatment was necessary and efficient for in vitro selection. In contrast, GFP-amastigotes showed low retention of the episomal plasmid in the absence of G418 selection and low sensitivity to antibiotics in vitro. The use of protocols for G418 selection using infected BALB/c mice also indicated low sensitivity to antibiotics against amastigotes in cutaneous lesions.

Clinical Features, Risk Factors and Outcomes of Bacteremia due to Enterococci with High-Level Gentamicin Resistance: Comparison with Bacteremia due to Enterococci without High-Level Gentamicin Resistance

High-level gentamicin resistance (HLGR) in enterococci has increased since the 1980s, but the clinical significance of the resistance and its impact on outcome have not been established. One hundred and thirty-six patients with bacteremia caused by enterococci with HLGR (HLGR group) were compared with 79 patients with bacteremia caused by enterococci without HLGR (non-HLGR group). Hematologic malignancy, neutropenia, Enterococcus faecium infection, nosocomial infection and monomicrobial bacteremia were more common in the HLGR group than the non-HLGR group, and APACHE II scores were also higher (P<0.05, in each case). Neutropenia, monomicrobial infection, stay in intensive care at culture, and use of 3rd generation cephalosporin, were independent risk factors for acquisition of HLGR enterococcal bacteremia. Fourteen-day and 30-day mortalities were higher in the HLGR group than the non-HLGR group in univariate analysis (37% vs. 15%, P=0.001; 50% vs. 22%, P<0.001). However, HLGR was not an independent risk factor for mortality due to enterococcal bacteremia in multivariate analysis. Therefore, HLGR enterococcal bacteremia is associated with more severe comorbid conditions and higher mortality than non-HLGR enterococcal bacteremia but the HLGR itself does not contribute significantly to mortality.

Efecto de trimetazidina en la nefrotoxicidad por gentamicina / Trimetazidine effect on nephrotoxicity by gentamicyn

La Trimetazidina (TMZ) es una droga utilizada como cardioprotector, ya que previene la muerte celular secundaria a la isquemia miocárdica. Algunos investigadores le atribuyeron efecto reno-protector, actividad antioxidante y scavenger de radicales libres del oxígeno. El objetivo del presente trabajo es mostrar el efecto citoprotector de TMZ en las alteraciones inducidas por Gentamicina (G) a nivel de la célula del túbulo renal. Se diseñaron esquemas en animales de experimentación tratados con ambas drogas. Ratas macho Wistar de 180 a 200 g de peso fueron distribuidas en 5 grupos (n=8) y tratadas con: dieta estándar (A); suplementada con 20 mg/Kg/día de TMZ durante 27 días (B); suplementada con 50 mg/Kg/día de G durante 7 días(C); pretratadas 20 días con 20 mg/Kg/día de TMZ y los últimos 7 días con G (D) y tratadas simultáneamente durante 7 días con 20 mg/Kg/día de TMZ y 50 mg/Kg/día de G(E). Se midieron los compuestos nitrogenados urea y creatinina, la excreción de gamma glutamiltranspeptidasa urinaria y se efectuaron estudios estructurales con tinción de hematoxilina-eosina y ultraestructurales. Se utilizó el grupo C como testigo de nefrotoxicidad inducida por G. El pretratamiento durante 20 días con TMZ demostró el efecto protector para la nefrotoxicidad inducida, sin cambios bioquímicos-funcionales, ni alteración de la histoarquitectura, ni de la ultraestructura. El tratamiento simultáneo con TMZ y G no mostró efecto protector. Se concluye que en el modelo de ratas macho Wistar se demuestra el efecto citoprotector de TMZ en tratamiento previo por 21 días. El estudio histológico del tejido renal, bajo estas condiciones, presenta histoarquitectura conservada y función renal normal. Se infiere que el efecto citoprotector de TMZ que impide la nefrotoxicidad inducida por G se debe a la inhibición de la reabsorción y acumulación de Gentamicina en la célula del túbulo proximal del nefrón.

Trimetazidine (TMZ) is a drug used as a cardioprotector since it prevents cell death secondary to myocardial ischemia. Some investigators have attributed protective effect, antioxidant activity and oxygen free radical scavenging abilityt to TMZ. The aim of the present work is to show the cytoprotective effect of TMZ on Gentamicin (G)-induced alterations at the level of the renal tubular cell. Schemes were designed in experimental animals treated with both drugs. Male Wistar rats weighing 200 to 260 g were divided into 5 groups (n=8) and treated with: standard diet (A); standard diet supplemented with 20 mg/Kg/day of TMZ for 27 days (B); standard diet supplemented with 50 mg/Kg/day of G for 7 days (C), pretreated for 20 days with 20 mg/Kg/day of TMZ and for the last 7 days with G (D), and treated simultaneously for 7 days with 20 mg/Kg/day of TMZ and 50 mg/Kg/day of G (E). The nitrogen compounds urea and creatinine were measured and so was the excretion of urinary gamma-glutamyl transpeptidase. Structural studies with hematoxilin and eosin staining and ultrastructural studies were also performed. Gentamicin was used as a control for nephrotoxicity (group C). Pretreatment with TMZ showed a protective effect against induced nephrotoxicity, with no biochemical changes or alterations in the histoarchitecture. Simultaneous treatment with TMZ and G (group E) showed no protective effect. Conclusions: the cytoprotective effect of TMZ on G-induced nephrotoxicity would take place at the level of the proximal tubular cell of the brush border by inhibiting G reabsorption and accumulation.

Gentamicin-induced preconditioning of proximal tubular LLC-PK1 cells stimulates nitric oxide production but not the synthesis of heat shock protein

Nephrotoxicity is the main side effect of antibiotics such as gentamicin. Preconditioning has been reported to protect against injuries as ischemia/reperfusion. The objective of the present study was to determine the effect of preconditioning with gentamicin on LLC-PK1 cells. Preconditioning was induced in LLC-PK1 cells by 24-h exposure to 2.0 mM gentamicin (G/IU). After 4 or 15 days of preconditioning, cells were again exposed to gentamicin (2.0 mM) and compared to untreated control or G/IU cells. Necrosis and apoptosis were assessed by acridine orange and HOESCHT 33346. Nitric oxide (NO) and endothelin-1 were assessed by the Griess method and available kit. Heat shock proteins were analyzed by Western blotting. After 15 days of preconditioning, LLC-PK1 cells exhibited a significant decrease in necrosis (23.5 ± 4.3 to 6.5 ± 0.3%) and apoptosis (23.5 ± 4.3 to 6.5 ± 2.1%) and an increase in cell proliferation compared to G/IU. NO (0.177 ± 0.05 to 0.368 ± 0.073 ìg/mg protein) and endothelin-1 (1.88 ± 0.47 to 2.75 ± 0.53 pg/mL) production significantly increased after 15 days of preconditioning compared toG/IU. No difference in inducible HSP 70, constitutive HSC 70 or HSP 90 synthesis in tubular cells was observed afterpreconditioning with gentamicin. The present data suggest that preconditioning with gentamicin has protective effects on proximal tubular cells, that involved NO synthesis but not reduction of endothelin-1 or production of HSP 70, HSC 70, or HSP 90. We conclude that preconditioning could be a useful tool to prevent the nephrotoxicity induced by gentamicin.

Outbreaks of Imipenem Resistant Acinetobacter Baumannii Producing OXA-23 beta-Lactamase in a Tertiary Care Hospital in Korea

PURPOSE: Since November 2006, imipenem-resistant Acinetobacter baumannii isolates have increased in Kyung Hee University Hospital in Seoul, Korea. The purpose of this study was to determine the genetic basis and molecular epidemiology of outbreak isolates. MATERIALS AND METHODS: Forty-nine non-repetitive isolates of the 734 IRAB strains were investigated in order to determine their characteristics. The modified Hodge and the ethylenediaminetetraacetic acid (EDTA)-disk synergy test were performed for the screening of carbapenemase and metallo-beta-lactamase production. Multiplex polymerase chain reaction (PCR) assays were performed for the detection of genes encoding for OXA-23-like, OXA-24-like, OXA-58-like and OXA-51-like carbapenemase. Pulsed-field gel electrophoresis (PFGE) was performed for strain identification. RESULTS: All isolates showed 100% resistance to ciprofloxacin and gentamicin, 97.9% resistance to cefepime, piperacillin/tazobactam, aztreonam, ceftazidime and piperacillin, 93.9% resistance to tobramycin and 57.1% resistance to amikacin. All of the 49 isolates (100%) showed positive results in the modified Hodge test and negative results in the EDTA-disk synergy test. They all (100%) possessed the encoding gene for an intrinsic OXA-51-like carbapenemase and an acquired OXA-23-like carbapenemase in the multiplex PCR assay. PFGE patterns revealed that all isolates were clonally related from A1 to A14. CONCLUSION: It is concluded that all of the 49 IRAB isolates acquired resistance to imipenem by producing OXA-23 carbapenemase and they might have originated from a common source.

In vitro activity of gentamicin and amikacin against Salmonella enterica serovar Typhi: a search for a treatment regimen for typhoid fever

The emergence of Salmonella enterica serovar Typhi isolates resistant to ciprofloxacin and 3rd-generation cephalosporins is a concern for physicians in developing countries. This study assessed the in vitro activity of gentamicin and amikacin against 464 S. enterica serovar Typhi isolates obtained from blood of patients clinically suspected of enteric fever who attended the Calcutta School of Tropical Medicine from 1991 to 2003. The isolates were sensitive to gentamicin and amikacin, showing minimum inhibitory concentrations 0.01-4 microg/mL and 0.005-3.5 microg/mL respectively. Both agents showed bactericidal activity at concentrations of 2 microg/mL after incubation for 6 hours. Aminoglycoside antibiotics such as gentamicin and amikacin may thus be introduced as a treatment regimen for typhoid fever

Clinical, epidemiological, and microbiological characteristics of bacteremia caused by high-level gentamicin-resistant Enterococcus faecalis

Enterococcus spp bacteremia is associated with high mortality and the appearance of high-level gentamicin resistance (HLGR) created additional challenges for the treatment of these infections. We evaluated the epidemiological and clinical characteristics of patients with bacteremias caused by HLGR and non_HLGR Enterococcus faecalis isolates at a teaching hospital in the State of São Paulo, Brazil. Patients with bacteremia due to E. faecalis diagnosed between January 1999 and December 2003 were included in the study. We collected clinical, epidemiological, and microbiological data from medical records. Banked isolates were typed using pulsed-field gel electrophoresis. We identified 145 cases of E. faecalis bacteremia: 66 (45.5 percent) were caused by HLGR isolates and 79 (54.5 percent) by non_HLGR. In the univariate analysis, patients with HLGR infection were older, had higher rates of bladder catheterization, and more often had treatment with cephalosporin, quinolone, and/or carbapenem compared with patients with non_HLGR infection (P < 0.05). Multivariate analysis indicated that older age, hematological malignancy, and previous use of vancomycin were independently associated with HLGR (P < 0.05). Mortality rates were not significantly different among patients with HLGR (50 percent) and non_HLGR (43 percent) infections (P = 0.40). Of the 32 genotyped isolates, 16 were distributed into 6 main electrophoresis patterns and 16 others had distinct patterns. E. faecalis bacteremia is associated with high mortality and is frequently caused by HLGR isolates at this teaching hospital. The variability among genotyped isolates suggests that endogenous infections, rather than patient-to-patient transmission of E. faecalis, are more common at this institution.