Cytokines released from blood monocytes and expressed in mucocutaneous lesions of patients with paracoccidioidomycosis evaluated before and during trimethoprim-sulfamethoxazole treatment

Background Mucocutaneous lesions in paracoccidioidomycosis are granulomatous and result from tissue responses to Paracoccidioides brasiliensis, the aetiological agent.Objectives and methods In this study we investigate the expression of tumour necrosis factor (TNF)-alpha, interleukin (IL)-10 and transforming growth factor (TGF)-beta 1 by immunohistochemistry in skin and mucosa lesions from patients with the chronic form of paracoccidioidomycosis, evaluated before and at day 20 of trimethoprim-sulfamethoxazole treatment. Cytokine production by peripheral blood monocytes was also studied by enzyme immunoassay.Results Intense immunostaining for TNF-alpha was detected in mononuclear cells that infiltrated granulomas in all skin and mucosa lesions before treatment simultaneously with low IL-10 granular deposits in these cells. At day 20 of treatment, there was reduced TNF-alpha and IL-10 deposition. Immunoreactive TGF-beta 1 was observed diffusely in the dermis and generally in the cytoplasm of macrophages and giant cells, before treatment, and as increased TGF-beta 1 deposits in the fibrosis area at day 20 of treatment. Peripheral blood monocytes from patients with paracoccidioidomycosis, evaluated before treatment, produced high endogenous levels of TNF-alpha, TGF-beta 1 and IL-10 in relation to healthy controls. Lipopolysaccharide-stimulated monocytes from patients secreted lower levels of TNF-alpha in both periods of evaluation while no impairment in capacity of IL-10 and TGF-beta production was observed.Conclusions Trimethoprim-sulfamethoxazole therapy was effective in decreasing fungal load in the lesions, allowing patient immune response to control the infection leading to the healing of the lesions.

Linfonodo pulmonar na paracoccidioidomicose aguda infantil (relato de um caso).

The primary complex like Ghon was observed in a child's clinical roentgenographic study. C.S., white, male, 6 years old, was born in Curitiba (PR), Brazil and living in Guaratingueta (SP), Brazil, developed common cold, bimodal diary fever, chills, shake and sweats. Dyspnea, cough with general lymphadenopathy. Foot and right shoulder arthralgias. Six months ago visited a cave, equitation practice, dog and cat contacts and no transfusion, frontal sweats, fever (38.4 degrees C). T.A. was 8/6, tachycardia in generalized lymphadenopathy. Cardiopulmonary system was normal, mesogastric tumoral mass, hepatosplenomegaly and no ascites. Bone marrow with eosinophilia; nodule demonstrated presence of P. brasiliensis, hypoalbuminemia; hyperglobulinemia; anemia; leukocytosis with eosinophilia. Immunodiffusion with exoantigen 43 kd of P. brasiliensis was 1/32. Primary complex like Ghon was observed in interstitial pneumonia followed by mediastinal and mesogastric mass (35 to 40 days). Clavicular osteolytic lesions (45 to 60 days) appeared during paracoccidioidomycosis therapy. Recovery was observed 2 months after treatment of acute infantile paracoccidioidomycosis.

Comparative efficacies of antibiotics in a rat model of meningoencephalitis due to Listeria monocytogenes

The antibacterial activities of amoxicillin-gentamicin, trovafloxacin, trimethoprim-sulfamethoxazole (TMP-SMX) and the combination of trovafloxacin with TMP-SMX were compared in a model of meningoencephalitis due to Listeria monocytogenes in infant rats. At 22 h after intracisternal infection, the cerebrospinal fluid was cultured to document meningitis, and the treatment was started. Treatment was instituted for 48 h, and efficacy was evaluated 24 h after administration of the last dose. All tested treatment regimens exhibited significant activities in brain, liver, and blood compared to infected rats receiving saline (P < 0.001). In the brain, amoxicillin plus gentamicin was more active than all of the other regimens, and trovafloxacin was more active than TMP-SMX (bacterial titers of 4.1 +/- 0.5 log10 CFU/ml for amoxicillin-gentamicin, 5.0 +/- 0.4 log10 CFU/ml for trovafloxacin, and 5.8 +/- 0.5 log10 CFU/ml for TMP-SMX; P < 0.05). In liver, amoxicillin-gentamicin and trovafloxacin were similarly active (2.8 +/- 0.8 and 2.7 +/- 0.8 log10 CFU/ml, respectively) but more active than TMP-SMX (4.4 +/- 0. 6 log10 CFU/ml; P < 0.05). The combination of trovafloxacin with TMP-SMX did not alter the antibacterial effect in the brain, but it did reduce the effect of trovafloxacin in the liver. Amoxicillin-gentamicin was the most active therapy in this study, but the activity of trovafloxacin suggests that further studies with this drug for the treatment of Listeria infections may be warranted.

Population pharmacokinetics and pharmacodynamics of lumefantrine in young Ugandan children treated with artemether-lumefantrine for uncomplicated malaria

Background. The pharmacokinetics and pharmacodynamics of lumefantrine, a component of the most widely used treatment for malaria, artemether-lumefantrine, has not been adequately characterized in young children. Methods. Capillary whole-blood lumefantrine concentration and treatment outcomes were determined in 105 Ugandan children, ages 6 months to 2 years, who were treated for 249 episodes of Plasmodium falciparum malaria with artemether-lumefantrine. Results. Population pharmacokinetics for lumefantrine used a 2-compartment open model with first-order absorption. Age had a significant positive correlation with bioavailability in a model that included allometric scaling. Children not receiving trimethoprim-sulfamethoxazole with capillary whole blood concentrations <200 ng/mL had a 3-fold higher hazard of 28-day recurrent parasitemia, compared with those with concentrations >200 ng/mL (P =. 0007). However, for children receiving trimethoprim-sulfamethoxazole, the risk of recurrent parasitemia did not differ significantly on the basis of this threshold. Day 3 concentrations were a stronger predictor of 28-day recurrence than day 7 concentrations. Conclusions. We demonstrate that age, in addition to weight, is a determinant of lumefantrine exposure, and in the absence of trimethoprim-sulfamethoxazole, lumefantrine exposure is a determinant of recurrent parasitemia. Exposure levels in children aged 6 months to 2 years was generally lower than levels published for older children and adults. Further refinement of artemether-lumefantrine dosing to improve exposure in infants and very young children may be warranted. © 2016 The Author.

Use of a proposed antimicrobial susceptibility testing method for Haemophilus parasuis

The aim of this study was to examine the antimicrobial susceptibility of 97 Haemophilus parasuis cultured from Australian pigs. As there is no existing standard antimicrobial susceptibility technique available for H. parasuis, methods utilising the supplemented media, BA/SN for disc diffusion and test medium broth (TMB) for a microdilution technique, were initially evaluated with the reference strains recommended by the Clinical and Laboratory Standards Institute. The results of the media evaluation suggested that BA/SN and TMB can be used as suitable media for susceptibility testing of H. parasuis. The proposed microdilution technique was then used with 97 H. parasuis isolates and nine antimicrobial agents. The study found that Australian isolates showed elevated minimum inhibitory concentrations (MICs) for ampicillin (1%), penicillin (2%), erythromycin (7%), tulathromycin (9%), tilmicosin (22%), tetracycline (31%) and trimethoprim-sulfamethoxazole (40%). This study has described potential antimicrobial susceptibility methods for H. parasuis and has detected a low percentage of Australian H. parasuis isolates with elevated antimicrobial MICs.

First detection of extended-spectrum cephalosporin- and fluoroquinolone-resistant Escherichia coli in Australian food-producing animals

This study aimed to define the frequency of resistance to critically important antimicrobials (CIAs) [i.e. extended-spectrum cephalosporins (ESCs), fluoroquinolones (FQs) and carbapenems] among Escherichia coli isolates causing clinical disease in Australian food-producing animals. Clinical E. coli isolates (n = 324) from Australian food-producing animals [cattle (n = 169), porcine (n = 114), poultry (n = 32) and sheep (n = 9)] were compiled from all veterinary diagnostic laboratories across Australia over a 1-year period. Isolates underwent antimicrobial susceptibility testing to 18 antimicrobials using the Clinical and Laboratory Standards Institute disc diffusion method. Isolates resistant to CIAs underwent minimum inhibitory concentration determination, multilocus sequence typing (MLST), phylogenetic analysis, plasmid replicon typing, plasmid identification, and virulence and antimicrobial resistance gene typing. The 324 E. coli isolates from different sources exhibited a variable frequency of resistance to tetracycline (29.0–88.6%), ampicillin (9.4–71.1%), trimethoprim/sulfamethoxazole (11.1–67.5%) and streptomycin (21.9–69.3%), whereas none were resistant to imipenem or amikacin. Resistance was detected, albeit at low frequency, to ESCs (bovine isolates, 1%; porcine isolates, 3%) and FQs (porcine isolates, 1%). Most ESC- and FQ-resistant isolates represented globally disseminated E. coli lineages (ST117, ST744, ST10 and ST1). Only a single porcine E. coli isolate (ST100) was identified as a classic porcine enterotoxigenic E. coli strain (non-zoonotic animal pathogen) that exhibited ESC resistance via acquisition of blaCMY-2. This study uniquely establishes the presence of resistance to CIAs among clinical E. coli isolates from Australian food-producing animals, largely attributed to globally disseminated FQ- and ESC-resistant E. coli lineages.

Characterisation of antibiotic-resistant psychrotrophic bacteria in raw milk

Dynamics of raw milk associated bacteria during cold storage of raw milk and their antibiotic resistance was reviewed, with focus on psychrotrophic bacteria. This study aimed to investigate the significance of cold storage of raw milk on antibiotic-resistant bacterial population and analyse the antibiotic resistance of the Gram-negative antibiotic-resistant psychrotrophic bacteria isolated from the cold-stored raw milk samples. Twenty-four raw milk samples, six at a time, were obtained from lorries that collected milk from Finnish farms and were stored at 4°C/4 d, 6°C/3 d and 6°C/4 d. Antibiotics representing four classes of antibiotics (gentamicin, ceftazidime, levofloxacin and trimethoprim-sulfamethoxazole) were used to determine the antibiotic resistance of mesophilic and psychrotrophic bacteria during the storage period. A representative number of antibiotic-resistant Gram-negative isolates retrieved from the cold-stored raw milk samples were identified by the phenotypic API 20 NE system and a few isolates by the 16S rDNA gene sequencing. Some of the isolates were further evaluated for their antibiotic resistance by the ATB PSE 5 and HiComb system. The initial average mesophilic counts were found below 105 CFU/mL, suggesting that the raw milk samples were of good quality. However, the mesophilic and psychrotrophic population increased when stored at 4°C/4 d, 6°C/3 d and 6°C/4 d. Gentamicin- and levofloxacin-resistant bacteria increased moderately (P < 0.05) while there was a considerable rise (P < 0.05) of ceftazidime- and trimethoprim-sulfamethoxazole-resistant population during the cold storage. Of the 50.9 % (28) of resistant isolates (total 55) identified by API 20 NE, the majority were Sphingomonas paucimobilis (8), Pseudomonas putida (5), Sphingobacterium spiritivorum (3) and Acinetobacter baumanii (2). The analysis by ATB PSE 5 system suggested that 57.1% of the isolates (total 49) were multiresistant. This study showed that the dairy environment harbours multidrug-resistant Gramnegative psychrotrophic bacteria and the cold chain of raw milk storage amplifies the antibioticresistant psychrotrophic bacterial population.

Complexo Burkholderia cepacia em pacientes com fibrose cística: caracterização das espécies, avaliação do perfil de susceptibilidade aos antimicrobianos e da diversidade genética

O Complexo Burkholderia cepacia (CBc) é um grupo de 17 espécies intimamente relacionadas que estão associadas à deterioração pulmonar e aumento da mortalidade em pacientes com Fibrose Cística (FC). Essas espécies variam entre si em relação à prevalência, quadros clínicos e virulência. Pouco é conhecido em relação ao perfil de resistência aos antimicrobianos. Uma vez estabelecida a infecção, a abordagem terapêutica e as medidas de controle atualmente adotadas são baseadas no CBc, sem considerar cada espécie em particular. O objetivo deste estudo foi determinar a prevalência das espécies do CBc em pacientes atendidos em dois centros de referência no Rio de Janeiro, bem como estabelecer perfis de resistência a antimicrobianos e avaliar a diversidade molecular entre as espécies. Cem amostras do CBc isoladas de 38 pacientes com FC no período de janeiro de 2010 a fevereiro de 2012 foram identificadas por métodos fenotípicos e pelo sequenciamento do gene recA. As CIMs para amicacina, aztreonam, ceftazidima, trimetoprim/sulfametoxazol e tobramicina foram determinadas por microdiluição e a genotipagem das espécies foi realizada por PFGE com a enzima SpeI. B. vietnamiensis (44%) foi a espécie mais prevalente, seguida de B. cenocepacia IIIA (36%), B. multivorans (10%), B. cenocepacia IIIB (1%) e B. stabilis (1%). Cinco por cento das amostras não foram identificadas. B. vietnamiensis foi identificada em mais da metade dos pacientes (58,3%). Foram observadas diferenças no perfil de susceptibilidade entre as espécies do CBc. B. cenocepacia IIIA foi a espécie que apresentou as maiores taxas de resistência aos antimicrobianos, sobretudo para trimetoprim/ sulfametoxazol (80,5%), principal antimicrobiano utilizado no tratamento de infecções causadas pelo CBc. Amostras com perfis MDR ocorreram em todas as espécies, destacando-se o perfil A, resistente simultaneamente aos cinco antimicrobianos, observado em 58,8% das amostras de B.cenocepacia IIIA. A análise do polimorfismo genético mostrou que, apesar de B. vietnamiensis ter sido a espécie mais prevalente, a ocorrência de nove grupos clonais sugere que a aquisição dessas cepas tenha se dado a partir de uma fonte ambiental comum. Para B. cenocepacia IIIA, 52,9% das amostras foram atribuídas a um mesmo grupo clonal (BcA), compartilhado entre nove pacientes atendidos em um mesmo centro de referência. Oitenta por cento dessas amostras apresentaram ainda resistência a todos os antimicrobianos testados. Os dados mostram que, mesmo com o emprego de técnicas moleculares, é difícil a identificação do CBc em nível de espécie; que B. cenocepacia IIIA é caracterizada por índices de resistência superiores às outras espécies e que a transmissão cruzada entre os indivíduos aponta para a necessidade do estabelecimento de medidas de vigilância do CBc nos centros de referência.

Use of adhesion counts to help predict symptomatic infection and the ability of fluoroquinolones to penetrate bacterial biofilms on the bladder cells of spinal cord injured patients

There were three objectives to the present study: (1) compare the bladder infection rate and extent of biofilm formation for seven untreated spinal cord injured (SCI) patients and seven given prophylactic co-trimoxazole, (2) identify a level of bacterial adhesion to bladder cells which could be used to help predict symptomatic infection, and (3) determine from in vivo and in vitro studies whether fluoroquinolones were effective at penetrating bacterial biofilms. The results showed that the infection rate had not changed with the introduction of prophylaxis. However, the uropathogenic population had altered subsequent to the introduction of prophylaxis with E. coli being replaced by E. faecalis as the most common cause of infection. In 63% of the specimens from asymptomatic patients, the bacterial counts per cell were <20, while 81% of specimens from patients with at least one sign and one symptom of urinary tract infection (UTI) had > 20 adherent bacteria per bladder cell. Therefore, it is proposed that counts of > 20 bacteria adherent to sediment transitional epithelial bladder cells may be predictive of symptomatic UTI. Clinical data showed that fluoroquinolone therapy reduced the adhesion counts to <20 per cell in 63% of cases, while trimethoprim-sulfamethoxazole only did so in 44%. Further in vitro testing showed that ciprofloxacin (0.1, 0.5 and 1.0 micrograms/ml) partially or completely eradicated adherent biofilms from 92% of spinal cord injured patients' bladder cells, while ofloxacin did so in 71% cases and norfloxacin in 56%. These findings have important implications for the detection and treatment of bacteriuria in spinal cord injured patients.

Antimicrobic susceptibility and plasmid profile analysis as identity tests for multiple blood isolates of coagulase-negative staphylococci

We compared a disk diffusion antimicrobic susceptibility panel with plasmid DNA profiles as tests for identity of 106 isolates of coagulase-negative staphylococci cultured from the blood of 45 patients on multiple occasions. The antimicrobic panel included penicillin, oxacillin, clindamycin, trimethoprim-sulfamethoxazole, chloramphenicol, tetracycline, tobramycin, kanamycin, and gentamicin. Nineteen patterns of antimicrobic susceptibility were found. The most common pattern was present in 25% of the isolates, and at least one isolate from 31% of the patients had this pattern. Forty-seven distinct plasmid DNA profiles were found. The most common plasmid profile was present in 8.5% of the isolates, and at least one isolate from 15% of the patients had this profile. Twenty-eight patients had multiple isolates that were identical by plasmid profile analysis. Twenty-seven (96%) of these patients had isolates that were also identical by antimicrobic susceptibility. Nineteen patients had multiple isolates that were different by plasmid profile analysis. In 18 (95%) of these patients, the isolates were also different by antimicrobic susceptibility. Although plasmid DNA profile analysis is a more discriminating tool, these data confirm that a selected disk diffusion antimicrobic susceptibility panel may be used to screen multiple blood isolates of coagulase-negative staphylococci for identity or differences.

Perfil microbiológico, prevalencia de resistencia antibiótica y patrones de susceptibilidad en infección urinaria en una población pediátrica

Introducción: El incremento de la resistencia antibiótica se considera un problema de salud pública con consecuencias clínicas y económicas, por lo tanto se determinará la prevalencia de resistencia antibiótica en Infección del Tracto Urinario (ITU, el perfil microbiológico y los patrones de susceptibilidad en una población pediátrica atendida en la Fundación Cardioinfantil. Materiales y métodos: Estudio observacional de corte transversal, retrospectivo, entre 1 mes a 18 años de edad, con diagnóstico de ITU comunitaria atendidos entre Enero de 2011 y Diciembre de 2013. Se excluyeron pacientes con dispositivos en la vía urinaria, instrumentación quirúrgica previa, trayectos fistulosos entre la vía urinaria y sistema digestivo, ITU luego de 48 horas de hospitalización y recaída clínica en tratamiento. Se estableció la prevalencia de ITU resistente y se realizó un análisis descriptivo de la información. Resultados: Se evaluaron 385 registros clínicos, con una mediana de 1.08 años (RIQ 0.8 – 4.08), el 73.5% eran niñas. La fiebre predominó (76.5%), seguido de emesis (32.0%), disuria (23.7%) y dolor abdominal (23.1%). El uropatógeno más frecuente fue E.coli (75%), seguido de Proteus mirabilis (8.5%) y Klebsiella spp. (8.3%). La Ampicilina, el Trimetropim sulfametoxazol, la Ampicilina sulbactam y el ácido nalidixico tuvieron mayor tasa de resistencia. La prevalencia de BLEE fue 5.2% y AmpC 3.9%. La prevalencia de resistencia antimicrobiana fue de 11.9%. Conclusiones: La E.coli es el uropatogeno más frecuentemente aislado en ITU, con resistencia a la ampicilina en 60.2%, cefalosporinas de primera generación en 15.5%, trimetropin sulfametoxazol en 43.9%, cefepime 4.8%. La prevalencia de resistencia antimicrobiana fue de 11.9%.

Community-onset Escherichia coli infection resistant to expanded- spectrum cephalosporins in low-prevalence countries

Background
By global standards the prevalence of community onset expanded-spectrum cephalosporin resistant Escherichia coli (ESC-R-EC) remains low in Australia and New Zealand. Of concern, our countries are in a unique position with high extramural resistance pressure from close population and trade links to Asia-Pacific neighbours with high ESC-R-EC rates. We aim to characterize the risks and dynamics of community onset ESC-R-EC in our low-prevalence region.

Methods
A case-control methodology was used. Patients with ESC-R-EC or susceptible E. coli isolated from blood or urine were recruited at six geographically dispersed tertiary hospitals in Australia and New Zealand. Epidemiological data was prospectively collected and bacteria were retained for analysis.

Results
In total, 182 patients (91 cases and 91 controls) were recruited. Multivariate logistic regression identified risk factors for ESC-R amongst E. coli including birth on the Indian subcontinent (OR=11.13, 2.17-56.98, p=0.003), urinary tract infection in the past year (per infection OR=1.430, 1.13-1.82, p=0.003), travel to South East Asia, China, Indian subcontinent, Africa and the Middle East (OR=3.089, 1.29-7.38, p=0.011), prior exposure to trimethoprim+/-sulfamethoxazole &/or an expanded-spectrum cephalosporin (OR=3.665, 1.30-10.35, p=0.014) and healthcare exposure in the previous six months (OR=3.16, 1.54-6.46, p=0.02).

Amongst our ESC-R-EC the blaCTX-M ESBLs was dominant (83% of ESC-R-EC), and the worldwide pandemic clone ST-131 was frequent (45% of ESC-R-EC).

Conclusion
In our low prevalence setting, ESC-R amongst community onset E. coli may be associated with both ‘export’ from healthcare facilities into the community and direct ‘import’ into the community from high-prevalence regions.

Prevalence and antimicrobial resistance patterns of methicillin-resistant staphylococci (MRS) isolated in a Veterinary Teaching Hospital in Brazil

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Thermal decomposition of some chemotherapic substances

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Multidrug-resistant urinary tract isolates of Escherichia coli from Ribeirão Preto, São Paulo, Brazil

Multiple resistances to antimicrobial drugs arising in Escherichia coli isolates may complicate therapeutic management of urinary tract infection (UTI) by this organism. In order to assess the multidrug resistance (MDR) among urinary E. coli isolates, we have tested 11 antimicrobial drugs against 67 isolates from outpatients attended in a tertiary-care teaching hospital and of 78 isolates from a municipal health unit, respectively in Ribeirão Preto, State of São Paulo, Brazil. Seventy-six percent and 22% of the isolates from the tertiary-care hospital and the municipal unit, respectively, were resistant to three or more different classes of agents, and were considered to present MDR. Among the isolates from the hospital patients, 73.0%, 65.0%, 58.0%, 58.0% and 31.0% were resistant to tetracycline, ampicillin, cephalothin, trimethoprim-sulfamethoxazole (TMP/SMX) and norfloxacin, respectively; resistance from the municipal unit patients were 31.0%, 37.0%, 8.0%, 29.0% and 12.0% respectively, to the same drugs. The predominant phenotype among the MDR isolates presented is ampicillin, TMP/SMX and tetracycline resistance. The high prevalence of drug resistance among UTI patients calls for continuous surveillance to assure effective control of this infection. © 2007 by The Brazilian Journal of Infectious Diseases and Contexto Publishing. All rights reserved.