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Background Mobile phones in hospital settings have been identified as an important source of cross-contamination because of the low frequency with which mobile phones are cleaned by health workers and cyclical contamination of the hands and face. The aim of this study was to investigate whether the mobile phones of the anesthesia team at a teaching hospital are potential reservoirs of nosocomial bacteria. In addition, differences in device sanitization and hand hygiene habits between attending and resident anesthesiologists were correlated with mobile phone colonization. Methods A prevalence study was conducted over a 6-month period from 2017 to 2018 that involved the collection of samples from the mobile phones of the anesthesiology team and culturing for surveillance. A questionnaire was administered to assess the mobile phone sanitization and hand washing routines of the anesthesia team in specific situations. Results Bacterial contamination was detected for 86 of the 128 mobile phones examined (67.2%). A greater presence of Micrococcus spp. on devices was correlated with a higher frequency of mobile phone use (p = 0.003) and a lower frequency of sanitization (p = 0.003). The presence of bacteria was increased on the mobile phones of professionals who did not perform handwashing after tracheal intubation (p = 0.003). Conclusion Hand hygiene and device sanitization habits were more important than the use behavior, as a higher presence of bacteria correlated with poorer hygiene habits. Furthermore, handwashing is the best approach to prevent serious colonization of mobile devices and the possible transmission of pathogens to patients under the care of anesthesiologists.

Infections are an important cause of mortality among patients receiving maintenance hemodialysis. Staphylococcus aureus is a frequent etiological agent and previous nasal colonization is a risk factor for infection. Repeated antimicrobial decolonization reduces infection in this population but can induce resistance. We compared photodynamic therapy, a promising bactericidal treatment that does not induce resistance, to mupirocin treatment among nasal carriers of Staphylococcus aureus.A randomized, controlled follow-up pilot study.34 patients receiving maintenance hemodialysis with nasal Staphylococcus aureus.Decolonization with photodynamic therapy (λ= 660 nm, 400 mW/cm2, 300 seconds, methylene blue 0.01%, single application) or with topical mupirocin (twice a day for 5 days).Nasal swabs were collected at T0 (when carrier state was identified), T1 (immediately after intervention), T2 (1 month after treatment), and T3 (3 months after treatment). Bacterial isolates were subjected to proteomic analysis to identify the species present and were characterized using an antibiogram.All 17 participants randomized to photodynamic therapy and 13 of 17 (77%) randomized to mupirocin were adherent to treatment. Among these participants, 12 (71%) had negative T1 cultures compared with 13 (77%) of the 17 participants treated with mupirocin (P=1.000; RR 0.92, CI 0.61 to 1.38). Moreover, 67% of patients who had negative cultures at T1 were recolonized within 3 months. There were no adverse events in the photodynamic therapy group.Restricted to nasal colonization and infectious complications were not assessed.Photodynamic therapy is a feasible approach to treating nasal carriage of Staphylococcus aureus. Future larger studies should be conducted to determine if photodynamic therapy is equivalent to the standard of care with mupirocin.

Polymyxins are still used mainly in treating infections caused by carbapenem-resistant Klebsiella pneumoniae worldwide. The most frequent mechanism of acquired resistance to polymyxins in Gram-negative bacilli is the occurrence of mutations in chromosomal genes regulating operons responsible for lipopolysaccharide modification. As we observed at Santa Casa de São Paulo hospital the occurrence of infections caused by isolates resistant to polymyxins in patients previously treated with this antimicrobial, and new infections caused by the same polymyxin-susceptible species, in this study, we aimed to determine the clonality of consecutive K. pneumoniae isolates from the same patients and characterize the molecular determinants of polymyxin resistance in paired or clonal isolates. A total of 24 pairs and one trio of K. pneumoniae isolates were included in this study. Species identification was achieved by mass spectrometry and multiplex PCR. Polymyxin B minimal inhibitory concentrations were determined by broth microdilution. Clonality was evaluated using pulsed-field gel electrophoresis. The presence of insertions in mgrB gene was tested by PCR, and mutations on pmrA, pmrB, phoP, and phoQ were evaluated by PCR and complete nucleotide sequencing. A fraction of 23.8% of strains resistant to polymyxin B had an insertion in mgrB. Amino acid substitution F204L in PmrB may be implicated in polymyxin resistance. Substitutions T246A and R256G in PmrB were not implicated in polymyxin resistance. In this study, polymyxin resistance after a first susceptible isolate was detected was most frequently due to an infection caused by a distinct clone. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s42770-021-00600-5.

Objective: Respiratory syncytial virus is a pathogen frequently involved in nosocomial outbreaks. Although several studies have reported nosocomial outbreaks in neonatal intensive care units, molecular epidemiology data are scarce. Here, the authors describe two consecutive respiratory syncytial virus outbreaks caused by genotypes ON-1 and NA-2 in a neonatal intensive care unit in São Paulo, Brazil. Methods: A prospective search for respiratory syncytial virus was performed after diagnosing the index case and four other symptomatic newborns in the neonatal intensive care unit. Nasopharyngeal aspirate samples of all patients in the neonatal intensive care unit were tested for 17 respiratory viruses using real-time reverse transcriptase polymerase chain reaction. Genotyping was performed using nucleotide sequencing. Results: From May to August 2013, two different outbreaks were detected in the neonatal intensive care unit. A total of 20 infants were infected with respiratory syncytial virus-A (ten and 14 with ON-1 and NA-2 genotypes, respectively). The mean age of the infants was 10 days, mean birth weight was 1,961 g, and the mean gestational age was 33 weeks. Risk factors (heart disease, lung disease, and prematurity) were present in 80% and 85.7% of infants in the ON-1 and NA-2 groups, respectively. In total, 45.8% of infants were asymptomatic and 20.8% required mechanical ventilation. Coinfections were not detected during the outbreaks. Conclusions: Infants in a neonatal intensive care unit who develop abrupt respiratory symptoms should be tested for respiratory viruses, especially respiratory syncytial virus. Even in the absence of severe symptoms, respiratory syncytial virus detection can prevent nosocomial transmission through infection control measures. A better understanding of respiratory syncytial virus molecular epidemiology is essential for developing new vaccines and antiviral drugs against respiratory syncytial virus. Resumo: Objetivo: O vírus sincicial respiratório é um patógeno frequentemente envolvido em surtos nosocomiais. Embora vários estudos tenham relatado tais surtos em unidades de terapia intensiva neonatal, os dados epidemiológicos moleculares são escassos. Neste artigo, descrevemos dois surtos consecutivos de vírus sincicial respiratório causados pelos genótipos ON-1 e NA-2 em uma unidade de terapia intensiva neonatal em São Paulo, Brasil. Métodos: Uma busca prospectiva por vírus sincicial respiratório foi realizada após o diagnóstico do caso índice e outros quatro recém-nascidos sintomáticos na unidade de terapia intensiva neonatal. Amostras de aspirado nasofaríngeo de todos os pacientes da unidade de terapia intensiva neonatal foram testadas para 17 vírus respiratórios com reação em cadeia da polimerase via transcriptase reversa em tempo real. A genotipagem foi realizada utilizando sequenciamento de nucleotídeos. Resultados: De maio a agosto de 2013, foram detectados dois surtos diferentes na unidade de terapia intensiva neonatal. Vinte e quatro crianças foram infectadas com vírus sincicial respiratório-A (10 e 14 com os genótipos ON-1 e NA-2, respectivamente). A média da idade dos lactentes era de 10 dias, o peso médio ao nascer foi de 1961 g e a idade gestacional média de 33 semanas. Fatores de risco (doença cardíaca, doença pulmonar e prematuridade) estavam presentes em 80% e 85,7% dos bebês nos grupos ON-1 e NA-2, respectivamente. No total, 45,8% dos lactentes eram assintomáticos e 20,8% necessitaram de ventilação mecânica. Não foram detectadas coinfecções durante os surtos. Conclusões: Bebês em unidade de terapia intensiva neonatal que desenvolvem sintomas respiratórios abruptos devem ser testados para vírus respiratórios, especialmente o vírus sincicial respiratório. Mesmo na ausência de sintomas graves, a detecção de vírus sincicial respiratório pode prevenir a transmissão nosocomial através de medidas de controle de infecção. Um melhor entendimento da epidemiologia molecular do vírus sincicial respiratório é essencial para o desenvolvimento de novas vacinas e drogas antivirais contra o vírus sincicial respiratório. Keywords: Respiratory syncytial virus, ON-1, NA-2, Outbreak, Neonatal intensive care unit, Palavras-chave: Vírus sincicial respiratório, ON-1, NA-2, Surto, Unidade de terapia intensiva neonatal

Objective: This study aimed to correlate amplitude-integrated electroencephalography findings with early outcomes, measured by mortality and neuroimaging findings, in a prospective cohort of infants at high risk for brain injury in this center in Brazil. Methods: This blinded prospective cohort study evaluated 23 preterm infants below 31 weeks of gestational age and 17 infants diagnosed with hypoxic-ischemic encephalopathy secondary to perinatal asphyxia, with gestational age greater than 36 weeks, monitored with amplitude-integrated electroencephalography in a public tertiary center from February 2014 to January 2015. Background activity (classified as continuous, discontinuous high-voltage, discontinuous low-voltage, burst-suppression, continuous low-voltage, or flat trace), presence of sleep-wake cycling, and presence of seizures were evaluated. Cranial ultrasonography in preterm infants and cranial magnetic resonance imaging in infants with hypoxic-ischemic encephalopathy were performed. Results: In the preterm group, pathological trace or discontinuous low-voltage pattern (p = 0.03) and absence of sleep-wake cycling (p = 0.019) were associated with mortality and brain injury assessed by cranial ultrasonography. In patients with hypoxic-ischemic encephalopathy, seizure patterns on amplitude-integrated electroencephalography traces were associated with mortality or brain lesion in cranial magnetic resonance imaging (p = 0.005). Conclusion: This study supports previous results and demonstrates the utility of amplitude-integrated electroencephalography for monitoring brain function and predicting early outcome in the studied groups of infants at high risk for brain injury. Resumo: Objetivo: Este estudo visou correlacionar os achados do eletroencefalograma de amplitude integrada (aEEG) com resultados precoces, medidos por mortalidade e achados de neuroim- agem, em uma coorte prospectiva de neonatos com risco elevado de lesão cerebral em nosso centro no Brasil. Métodos: O estudo prospectivo de coorte cego avaliou 23 neonatos prematuros abaixo de 31 semanas de idade gestacional (IG) e 17 neonatos diagnosticados com Encefalopatia Hipóxico-Isquêmica (EHI) secundária à asfixia perinatal, com IG superior a 36 semanas, monitorados com aEEG em um centro terciário público de fevereiro de 2014 a janeiro de 2015. Foram avaliadas a atividade de base (classificada como padrão contínuo, descontínuo de alta voltagem, descontínuo de baixa voltagem, supressão de explosão, contínuo de baixa voltagem ou traço plano), a presença de ciclo do sono-vigília e a presença de convulsões. Foram feitas a ultrassonografia craniana em prematuros e a ressonância magnética (RM) craniana em neonatos com EHI. Resultados: No grupo de prematuros, o traço patológico ou padrão descontínuo de baixa voltagem (p = 0,03) e a ausência de ciclo do sono-vigília (p = 0,019) foram associados a mortalidade e lesão cerebral avaliada por ultrassonografia craniana. Em pacientes com EHI, os padrões de convulsão nos traçados do aEEG foram associados a mortalidade ou lesão cerebral na RM craniana (p = 0,005). Conclusão: Este estudo corrobora os resultados anteriores e demonstra a utilidade do aEEG no monitoramento da função cerebral e na predição de alterações precoces nos grupos de neonatos estudados com risco elevado de lesão cerebral. Keywords: Newborn, Brain injury, Amplitude-integrated EEG, Early outcome, Palavras-chave: Recém-nascido, Lesão cerebral, EEG de amplitude integrada, Resultado precoce

Introduction Febrile neutropenia is one of the most serious treatment-related complications in cancer patients. Susceptible to rapidly progressing infections, which result in prolonged hospitalization and use of broad-spectrum antibiotics, neutropenic patients are subject to colonization by multiresistant agents, which enhances the risk of infections. Methods In this study we included samples collected with nasal, oropharyngeal and anal swabs from hospitalized children with febrile neutropenia following chemotherapy, between March 2014 and 2015, aiming to elucidate colonization by S. aureus and Enterococcus spp., as well as their resistance profile. Results S. aureus was found in 22% of the patients and 14% of the events. Methicillin-resistant S. aureus colonized 13.6% of patients. Including anal swabs in the screening increased the identification of colonized patients by 20%. Enterococcus spp. was found in 27% of patients and 17% of episodes. Enterococcal isolates resistant to vancomycin, accounting for 25% of the total, were not isolated in anal swabs at any time, with the oropharyngeal site being much more important. The rate of infection by Enterococcus spp. was 4.5% of all patients and 16% among the colonized patients. Conclusion Especially in this population, colonization studies including more sites can yield a higher chance of positive results. Establishing the colonization profile in febrile neutropenic children following chemotherapy may help to institute an empirical antibiotic treatment aimed at antibiotic adequacy and lower induction of resistance, thereby decreasing the risk of an unfavorable clinical outcome.

Aims This study investigates the antimicrobial activity in Staphylococcus aureus isolates (methicillin-sensitive S. aureus (MSSA) and methicillin-resistant S. aureus (MRSA)) and antioxidant activity of green propolis, Baccharis dracunculifolia DC extracts and Artepillin C™. Methods and Results The amount of Artepillin C in different extracts was determined by high performance liquid chromatography analysis. Minimum inhibitory concentration 90 (MIC90) was determined using 40 isolates of S. aureus inoculated in Mueller–Hinton agar culture medium containing the green propolis and B. dracunculifolia DC extracts. PVEE (green propolis ethanolic extract) and BDEH (B. dracunculifolia hexanic extract) showed the greatest antimicrobial activity with MIC90 values of 246·3 and 295·5 μg ml−1 respectively. Green propolis ethanolic and hexanic extracts (PVEE and PVEH respectively) showed the greatest antioxidant activity assessed by DPPH (1,1-diphenyl-2-picryl hydrazyl radical) with IC50 values of 13·09 and 95·86 μg ml−1 respectively. Conclusions Green propolis ethanolic displays better antimicrobial and antioxidant activities compared to other extracts. These activities may be related to the presence of Artepillin C in synergism with the other constituents of the extracts. Significance and Impact of the Study In this study, the antimicrobial activity of the extracts of green propolis and B. dracunculifolia DC demonstrated in MRSA and MSSA clinical isolates indicated that they can be important tools to treat infections caused by these bacteria.

Hospital-acquired infections account for high mortality rates and hospital costs. We analyzed pediatric data from a tertiary teaching hospital and found that most of the cases occurred in the intensive care unit and had significant association with invasive devices. Bloodstream infections were the main site of infection, and Gram-negative bacteria were the predominant etiology.

From July 2009 to July 2015, Staphylococcus aureus isolated from pediatric sterile sites were selected. Polymerase chain reaction was used to detect mecA and lukS-PV/lukF-PV genes. The rate of methicillin-resistant Staphylococcus aureus was 37.7%. Ten isolates had the lukS-PV/lukF-PV genes, 2 of which were methicillin-resistant Staphylococcus aureus. Skin and soft tissues infections were significantly associated with lukS-PV/lukF-PV positive isolates, P = 0.008.

BACKGROUND: Clindamycin has become an important antimicrobial option for the treatment of Staphylococcus aureus. However, little is known about the current patterns of clindamycin-susceptibility in S. aureus invasive isolates, both in our country and in other developing countries in the world. AIMS: The aim of this study was to determine the prevalence of constitutive and inducible clindamycin resistance in methicillin-susceptible S. aureus (MSSA) and methicillin-resistant S. aureus (MRSA) blood culture isolates in São Paulo, Brazil. MATERIALS AND METHODS: From July 2011 to June 2012, all S. aureus isolates from blood cultures collected at our hospital were included in the study. Antimicrobial susceptibility testing was performed according to recommendations of the Clinical and Laboratory Standards Institute. RESULTS: Total prevalence of clindamycin resistance was 68%, including 7.2% with inducible resistance. In MRSA resistance rate was 90.8% whereas in MSSA the rate was 32.7%. CONCLUSIONS: Our high prevalence of clindamycin resistance highlights the importance of performing D-test in a routine base, as well of maintaining continued surveillance for the prevalence of clindamycin resistance.

Spontaneous bacterial peritonitis is an uncommon manifestation of invasive pneumococcal disease and frequently occurs when an underlying hepatic disease is present. Bacterial identification through culture can be particularly challenging in patients with prior or concurrent antimicrobial use. DNA amplification detects very few copies of target DNA under ideal conditions in CSF or pleural effusion and, therefore, can be useful in selected infections. A culture-negative spontaneous pneumococcal peritonitis without preexisting peritoneal disease diagnosed by qPCR is herein described. Keywords: Streptococcus pneumoniae, Invasive pneumococcal disease, Spontaneous peritonitis, Real-time PCR, lytA primer

We studied 70 pediatric clinical isolates of Staphylococcus aureus to perform a comparison of different methods for detection of methicillin-resistant S. aureus (MRSA) using polymerase chain reaction for mecA as the “gold standard” assay. The isolates were tested with oxacillin and cefoxitin discs, oxacillin agar screening plate and E-test. Of the 70 strains, 43 were mecA-positive and 27 mecA-negative. Oxacillin agar screening plate and E-test had 100% sensitivity and specificity for the presence of mecA gene. We found a specificity of 96% for both disc-diffusion methods, and sensitivities of 95% (oxacillin disc) and 93% (cefoxitin disc). However, with the new Clinical and Laboratory Standards Institute 2007 cefoxitin breakpoints the sensitivity of this method would be increased to 100%, without any decrease in specificity. These results show how important is the evaluation of the available microbiological methods by different laboratories in different settings.

To the Editor: Infections caused by carbapenem-resistant Enterobacteriaceae have been associated with higher death rates than infections caused by carbapenem-susceptible strains, and resistant infections are mostly treated with polymyxins (1). Several outbreaks caused by carbapenem- and polymyxin-resistant Klebsiella pneumoniae (CPRKp) have been reported, mainly from Europe, and represent an emerging threat.

To identify the onset time of bacterial proliferation in mechanical ventilator circuits in a pediatric intensive care unit (PICU) and the colonizing agents involved, as well to verify any possible pre-use contamination, we conducted a prospective microbiological analysis of the circuits of 30 patients, with a total of 342 cultures. Bacterial colonization of the mechanical ventilator circuits in the studied PICU was confirmed. Microorganisms were present in 39% of expiratory branch cultures: K. pneumoniae (77%), A. baumannii (65%), and S. aureus (42%). The inspiratory branch registered a smaller number of positive culture: A. baumannii (51%), S. aureus, coagulase-negative (38%) and K. pneumoniae (32%). As to the colonization onset time, 23% of the cultures collected on the 1 day from the inspiratory branch were positive, versus 30% of those collected from the expiratory branch, which lead us to conclude that bacteria can be present within the first hours of use. The sequential analysis of bacterial colonization over the course of circuit usage and the observation that the number of positive cultures becomes progressively higher on the 5t, 6 and 7 days in the expiratory branch both suggest that the systems should be replaced more often. Original Research Article British Microbiology Research Journal, 4(6): 598-606, 2014 599

Traditional methods for microbial identification are often very laborious and time consuming. A new mass spectrometry based technique, matrix-assisted laser desorption ionization-time of flight (MALDI-TOF), has been described as a rapid, practical and low-cost method for this purpose. In this article, primary and possible future applications of this tool are briefly discussed. Os métodos tradicionais para identificação de microrganismos no laboratório clínico muitas vezes são trabalhosos e demorados. Uma nova metodologia, com base em espectrometria de massas, a matrix-assisted laser desorption ionization-time of flight (MALDI-TOF), é extremamente promissora para utilização na rotina microbiológica, sendo rápida, prática e pouco custosa. Neste artigo, são expostas, de forma breve, as principais aplicações atuais do método, assim como as perspectivas futuras.

BACKGROUND: After the dissemination of penicillin and oxacillin resistance in Staphylococcus aureus, vancomycin-intermediate and vancomycin resistant isolates have been reported. Even between isolates with minimum inhibitory concentrations (MICs) within the susceptible range, some authors have demonstrated that higher MICs correlate with higher lethality. METHODS: To test this hypothesis in our setting, we compared vancomycin MICs evaluated by two methods and clinical outcomes in hospitalized patients with S. aureus bacteremia. RESULTS: We compared lethality in patients infected with isolates that had MICs under or over 2 mg/L. Among patients infected with isolates that had microdilution MICs

Mupirocin is a topical antimicrobial agent and part of most Staphylococcus aureus decolonization regimens. Thus, knowing the mupirocin susceptibility profile of colonizing S. aureus is paramount for the proper use of this agent. We evaluated S. aureus isolates from 128 colonized children, using disc diffusion (with 5 mcg and 200 mcg discs) and Etest. None were low-level or high-level mupirocin-resistant. Since mupirocin will be increasingly needed for the control of S. aureus infection, continuous monitoring of its susceptibility status is necessary.

To the editor, We read with interest the paper authored by Preoţescu LL and Streinu-Cercel O,1 and would like to input more data into their brief review on the subject of Staphylococcus aureus colonization in children. We have previously performed and published2 a colonization study, from March to November 2009, in 197 children in Sao Paulo, Brazil. Samples were obtained using swabs from both the anterior nares and the oropharynx of each individual. The overall prevalence of S. aureus was 65% and prevalence of methicillin-resistant isolates was 5.9%. In 51 patients (25.9%), S. aureus was detected in both sites. Oropharyngeal colonization without nasal colonization occurred in 42 cases (21.3%), whereas nasal colonization without colonization of the oropharynx was the case for 35 children (17.8%). Of the eleven MRSA isolates, six (3.2%) were found only in the oropharynx and three (1.6%) only in nares. Our results have shown, in addition to a high prevalence of colonization, the importance of investigating other sites in addition to the anterior nares.

Methicillin-resistant Staphylococcus aureus (MRSA) are now a worldwide problem. Cystic fibrosis (CF) patients are commonly colonized and infected by MRSA. Accurate oxacillin susceptibility testing is mandatory for the adequate management of these patients. We performed a comparison of the accuracy of different tests in CF isolates, including methicillin-susceptible S. aureus and MRSA with different SCCmec types, and using the mecA gene as the gold-standard. The sensitivity and specificity of oxacillin disc, Etest, and oxacillin agar screening plate were 100%. Sensitivity of the cefoxitin disc was 85% and specificity was 100%. For clinically relevant isolates, laboratories may consider the use of a combination of two phenotypic methods.

Twenty seven S. aureus isolates were obtained from cystic fibrosis (CF) patients at a tertiary care hospital in Brazil. Nineteen (70.4%) were methicillin-susceptible S. aureus (MSSA) and eight (29.6%) methicillin-resistant S. aureus (MRSA). Of the MRSA isolates, four had SCCmec type III and four had SCCmec type IV. PVL genes were not detected in any of the MSSA or MRSA isolates. New studies are necessary to evaluate the exact impact of these different MRSA clones in CF patients.

Apesar das diversas recomendações disponíveis na literatura sobre o diagnóstico laboratorial da resistência à oxacilina em Staphylococcus aureus, a verdade é que ainda não existe consenso absoluto sobre qual ou quais métodos utilizar e quando utilizar cada um deles. Segundo a literatura, a maioria dos métodos disponíveis apresenta um índice não desprezível de erros diagnósticos. Em casos de isolados de processos infecciosos invasivos, ou de infecções persistentes e/ou refratárias aos antimicrobianos, o uso de um método adicional ou confirmatório pode ser importante. Além disso, a contínua reavaliação desses testes é necessária. Despite guidelines published in the literature, the optimal routine phenotypic method for detecting methicillin resistance in Staphylococcus aureus remains controversial. Most available methods have a considerable rate of errors, according to the literature. When testing invasive isolates or isolates from persistent/refractory infections, using an additional or confirmatory method could be important. In addition, the continual evaluation of these tests is necessary.

We evaluated the performance of several methods for the detection of methicillin resistance in Staphylococcus aureus using 101 clinical S. aureus isolates from pediatric patients in a tertiary hospital in Brazil; 50 isolates were mecA-positive and 51 were mecA-negative. The Etest and oxacillin agar screening plates were 100% sensitive and specific for mecA presence. Oxacillin and cefoxitin disks gave sensitivities of 96 and 92%, respectively, and 98% specificity. Alterations of CLSI cefoxitin breakpoints increased sensitivity to 98%, without decreasing specificity. Our results highlight the importance of a continuing evaluation of the recommended microbiological methods by different laboratories and in different settings. If necessary, laboratories should use a second test before reporting a strain as susceptible, especially when testing strains isolated from invasive or serious infections. With the new (2007) CLSI breakpoints, the cefoxitin-disk test appears to be a good option for the detection of methicillin resistance in S. aureus. Dept Pediat, Div Pediat Infect Dis, Sao Paulo, Brazil Santa Casa Univ Hosp, Dept Pathol, Microbiol Lab, Sao Paulo, Brazil FMUSP, Hosp Clin, LIM 03, Sao Paulo, Brazil Web of Science

Goud et al 1 , in their very interesting article regarding the prevalence of Staphylococcus aureus in their community, state that “the anterior nares are the most common site for colonization”. Recent data suggest that the oropharynx could be more frequently colonized by S. aureus than the anterior nares 2-4 . Furthermore, even in some populations where nasal colonization is more common than oropharyngeal colonization, the use of a throat swab would increase significantly the rate of detection of the carriers 5 . In addition, another matter of discussion would be the chosen screening method for vancomycin resistance. The authors used disk-diffusion, which is not considered accurate and reliable enough for the detection of decreased susceptibility of S. aureus to this antimicrobial agent. It is of paramount importance to remember that, according to the current guidelines issued by the Clinical and Laboratory Standards Institute (CLSI, formerly NCCLS), minimum inhibitory concentration (MIC) tests should be performed to determine the susceptibility of all isolates of staphylococci to vancomycin. The disk test, although detecting accurately S. aureus containing vanA (vancomycin resistant isolates), does not differentiate vancomycin-susceptible from vancomycin-intermediate isolates 6 .

O perfil de resistência, que algumas das espécies do complexo Klebsiella pneumoniae podem expressar, representa uma grande ameaça à saúde humana, particularmente quando resistentes aos carbapenêmicos, que são amplamente utilizados no tratamento de infecções graves em pacientes hospitalizados. O principal mecanismo de resistência aos carbapenêmicos é a produção de carbapenemases, particularmente dos tipos KPC e NDM. Um dos compostos desenvolvidos para o tratamento de infecções causadas por cepas produtoras de KPC é a combinação ceftazidimaavibactam (CAZ-AVI), mas que não tem atividade inibitória sobre metalo-betalactamases, a exemplo das NDMs. Os objetivos deste trabalho foram determinar a frequência das espécies do complexo K. pneumoniae e da coprodução de KPC, avaliar a clonalidade dos isolados, a sensibilidade ao aztreonam-avibactam (ATM-AVI), o desempenho do disco de meropenem (MEM) com inibidores para detecção de coprodução de NDM e KPC e desenvolver um teste de triagem para prever a sensibilidade ao ATM-AVI. Um total de 113 isolados do complexo K. pneumoniae produtoras de NDM ou coprodutoras de NDM e KPC, provenientes da coleção de bactérias do Grupo Fleury, coletadas períodos pré e pós início do uso de CAZ-AVI no Brasil, foram utilizadas neste estudo. A identificação da espécie e a presença dos genes blaNDM e blaKPC foi confirmada por PCR multiplex. A clonalidade dos isolados foi avaliada por eletroforese em campos pulsados (PFGE) após clivagem com XbaI. A produção de carbapenemases foi confirmada utilizando-se o teste Blue Carba. O desempenho dos discos de meropenem e CAZ-AVI contendo um ou mais inibidores de carbapenemases foi comparado com o teste molecular. A pré-difusão combinada foi realizada pré-incubando-se o ágar não inoculado com disco de CAZ-AVI, e a seguir aplicando-se o inóculo bacteriano e um disco de ATM após remover o disco de CAZ-AVI. Após incubação, os halos foram aferidos e correlacionados com a concentração inibitória mínima para ATM-AVI. As CIMs para ATM e ATM-AVI foram determinadas segundo o EUCAST. A identificação das espécies por PCR evidenciou as seguintes frequências: K. pneumoniae 75,2% (n=85); K. quasipneumoniae 16,8% (n=19), e K. variicola 8% (n=9). Uma fração de 12,4% (n=14) dos isolados apresentaram os genes blaNDM e blaKPC e 87,6% (n=99) apenas blaNDM. A análise dos perfis de PFGE de K. pneumoniae evidenciou a presença de cinco grupos clonais predominantes. Isolados do principal grupo clonal Ap (n=15) foram detectados nas cidades de São Paulo e Porto Alegre durante todo o período analisado. O grupo clonal Lp foi detectado nas cidades de São Paulo e Recife em 2019. Os dois principais grupos clonais no período pré-CAZ-AVI continham maior número de isolados do que aqueles no período de uso do CAZ-AVI. Os perfis de PFGE de K. quasipneumoniae evidenciaram quatro grupos clonais predominantes, e presentes apenas no estado de São Paulo, com persistência do grupo clonal Aq desde 2017. Quanto à K. variicola, foram observados dois grupos clonais predominantes Av e Bv, o primeiro presente apenas em São Paulo desde 2018 e o segundo em Porto Alegre apenas em 2019. Calculando-se a diferença entre os diâmetros de halo do disco MEM contendo EDTA e ácido fenilborônico (AFB) e o maior dos halos obtidos para MEM com EDTA ou AFB, observou-se que todos os isolados com coexpressão de KPC e NDM apresentaram diferença ≥ 5 mm. Uma fração de 42,3% dos isolados positivos apenas para blaNDM apresentaram sensibilidade para ATM (CIM ≤ 4 mg/L). Todos os isolados testados apresentaram CIM para ATM-AVI ≤ 1/4 mg/L, sendo a CIM90 0,125/4 mg/l. No teste de pré-difusão combinada, o menor diâmetro de halo obtido foi de 23 mm. A espécie predominante na amostragem foi K. pneumoniae. A disseminação clonal, observada neste estudo, contrasta com a diversidade clonal descrita em outros locais do mundo para produtores de NDM, exceto Grécia e China. Considerando os pontos de corte atuais para ATM, é provável que haja resposta clínica adequada no uso de ATM-AVI no tratamento de infecções causadas por isolados produtores de NDM e coprodutores de KPC e NDM. Utilizando-se o valor de corte de ≤ 5 mm para a diferença entre halos de inibição, de MEM com AFB e EDTA e o segundo maior halo com inibidor, a sensibilidade foi de 100% e a especificidade foi de 96,1,0%. O método de pré-difusão com CAZ-AVI e ATM é um método simples e o diâmetro ≥ 23 mm tem excelente correlação com a CIM para ATM-AVI ≤ 1/4 mg/L. The resistance profile, which some species of the Klebsiella pneumoniae complex may express, represent a great threat to human health, particularly when resistant to carbapenems, which are widely used in the treatment of severe infections in hospitalized patients. The main mechanism of resistance to carbapenems is the production of carbapenemases, particularly KPCs and NDMs. One of the compounds developed for the treatment of infections caused by KPC-producing strains is the combination ceftazidime-avibactam (CAZ-AVI), but which has no inhibitory activity on metallobetalactamases, as is the case for NDMs. The objectives of this work were to determine the frequency of K. pneumoniae complex species and KPC co-production, evaluate the clonality of isolates, the susceptibility to aztreonam-avibactam (ATM-AVI), the performance of meropenem (MEM) disks with inhibitors for detecting NDM co-production and KPC and develop a screening test to predict sensitivity to ATM-AVI. A total of 113 NDM-producing or NDM and KPC co-producing K. pneumoniae complexes, from the Fleury Group\'s bacteria collection, collected in the pre- and post-starting periods of CAZ-AVI use in Brazil, were used in this study. Species identification and the presence of the blaNDM and blaKPC genes were confirmed by multiplex PCR. The clonality of the isolates was evaluated by pulsed field electrophoresis (PFGE) after cleavage with XbaI. Carbapenemase production was confirmed using the Blue Carba test. The performance of MEM and CAZ-AVI disks containing one or more carbapenemase inhibitors was compared with the molecular test. Combined pre-diffusion was performed by preincubating the uninoculated agar with a CAZ-AVI disk, and then applying the bacterial inoculum and na ATM disk after removal of the CAZ-AVI disk. After incubation, halos were measured and correlated with the minimum inhibitory concentration (MIC) for ATM-AVI. ATM and ATM-AVI MICs were determined according to EUCAST. The identification of species by PCR evidenced the following frequencies: K. pneumoniae 75.2% (n=85); K. quasipneumoniae 16.8% (n=19), and K. variicola 8% (n=9). A fraction of 12.4% (n=14) of the isolates had the blaNDM and blaKPC genes and 87.6% (n=99) had only blaNDM. The analysis of the PFGE profiles of K. pneumoniae evidenced the presence of five predominant clonal groups. Isolates from the main clonal group Ap (n=16) were detected in the cities of São Paulo and Porto Alegre throughout the analyzed period. The clonal group Lp was detected in the cities of São Paulo and Recife 2019. The PFGE profiles of K. quasipneumoniae showed four predominant clonal groups, present only in the state of São Paulo, with persistence of the clonal group Aq since 2017. As for K. variicola, two predominant clonal groups Av and Bv were observed, the first present only in São Paulo since 2018 and the second in Porto Alegre only in 2019. Calculating the difference between the inhibition zone diameters of the MEM disk containing EDTA and phenylboronic acid (AFB) and the largest of the inhibition zone diameters obtained for MEM with EDTA or AFB, it was observed that all isolates with co-expression of KPC and NDM showed a difference 5 ≥mm. A fraction of 42.3% of isolates positive only for blaNDM showed sensitivity to ATM (MIC ≤ 4 mg/L). All tested isolates presented MIC for ATM-AVI ≤ 1/4 mg/L, being the MIC90 0.125/4 mg/l. In the combined pre-diffusion test, the smallest inhibition zone diameter obtained was 23 mm. The predominant species in the sample was K. pneumoniae, but a significant fraction of the other species in the complex was also observed in the sample. The clonal spread observed in this study contrasts with the clonal diversity described elsewhere in the world for NDM-producing isolates, except Greece and China. Considering the current cut-off points for ATM, it is likely that there is an adequate clinical response in the use of ATM-AVI in infections caused by NDM-producing and KPC-NDM co-producing isolates in Brazil. Using the cutoff value of 5 mm for the difference between inhibition zones, of MEM with AFB and EDTA and the second largest zone of MEM with inhibitor, the sensitivity was 100% and the specificity was 96.1%. The pre-diffusion method with CAZ-AVI and ATM is a simple method and the diameter ≥ 23 mm has excellent correlation with the MIC for ATM-AVI ≤ 1/4 mg/L.

As micobactérias não tuberculosas (NTM) podem ser encontradas em diversos ambientes, como solo, sistemas aquáticos naturais, sistemas de abastecimento de água potável e também em eucariotos. As micobactérias de crescimento rápido são um subgrupo de NTM que têm prevalência significativa em infecções relacionadas a traumas cutâneos, procedimentos cirúrgicos vídeo-assistidos e em infecções pulmonares, sendo Mycobacterium abscessus a espécie mais relevante desse grupo. Durante um pseudo-surto de infecções pulmonares pós-broncoscopia, ocorrido na cidade de São Paulo em 2007, cepas de M. abscessus subsp. bolletii (\"M. massiliense\") pertencentes ao clone epidêmico BRA100 foram detectadas em amostras coletadas do único broncoscópio usado e em amostras de lavado broncoalveolar. Todas as cepas, com exceção da F1660, apresentaram tolerância ao glutaraldeído, uma característica marcante do clone BRA100. Foi realizado o sequenciamento completo do genoma utilizando-se o sistema Illumina com metodologia de pares casados e as sequências obtidas foram comparadas, com enfoque na análise de genes potencialmente envolvidos na tolerância ao glutaraldeído. Encontramos em ambas as cepas cromossomos com 4,6 Mpb, com 64% de conteúdo GC. As sequências de nucleotídeos possuem 99% de identidade entre si. Ambos os cromossomos possuem 4.616 sequências codificantes de proteínas. Elementos de profago, como proteínas de bacteriófagos, estão presentes nos cromossomos. Bem como diversos genes associados à resistência a antibióticos e biocidas. Foram localizados operons mce, associados com a capacidade de sobrevivência em amebas e invasão de macrófagos e componentes da família T7SS, relacionadas à virulência em diversas espécies de micobactérias, como M. tuberculosis e também à transferência genética horizontal em M. smegmatis. Adicionalmente, foi detectada em ambas as cepas uma estrutura extracromossômica circular, de 97 Kbp, com 62,7% de conteúdo GC e 121 sequências codificantes. Embora a maioria das proteínas preditas tenha função desconhecida, foi possível encontrar um bloco de genes que pertencem ao sistema de secreção do tipo sete (T7SS), similar ao encontrado em plasmídeos de micobactérias de crescimento lento, sugerindo a troca de material genético entre essas espécies. Apenas na cepa tolerante ao glutaraldeído - F1725 - foi detectado um plasmídeo do grupo IncP, designado pBRA100, que contém genes que codificam resistência à kanamicina, amônio quaternário, sulfonamidas e estreptomicina, e um novo gene fosI, descrito neste estudo, que confere resistência à fosfomicina. Não foram encontradas nos cromossomos diferenças que possam justificar a tolerância ao glutaraldeído. A diferença marcante entre os dois genomas é a presença do plasmídeo pBRA100 na cepa tolerante ao glutaraldeído Non-tuberculous mycobacteria (NTM) can be found in diverse environments as soil, water plant, natural water systems as well as in Eukaryotes. Among the NTM group, rapid growth mycobacteria (RGM) have a substantial prevalence in infections following cutaneous trauma, lung infection and after video assisted surgeries. Mycobacterium abscessus is considered to be the most relevant pathogen pertaining to the RGM group. During a pseudooutbreak of lung infections occurred in the city of São Paulo in 2007, strains from M. abscessus subsp. bolletii (\"M. massiliense\") pertaining to the BRA100 clone were detected in samples from the biopsy channel of the only bronchoscope in use as well as from bronchoalveolar lavage. All the strains but F1660 exhibited tolerance to glutaraldehyde, which is a remarking characteristic of the BRA100 clone. We report here the complete genome sequences for the strains F1725 and F1660, respectively tolerant and susceptible to glutaraldehyde. The genomes of both strains consist of a 4.6 Mpb chromosome with 4,616 coding sequences and high GC content of 64%. The chromosomes encode diverse proteins related to antibiotic and biocide resistance. Operons involved in virulence, as mce, are present as well as components from T7SS family, related to virulence in Mycobacterium tuberculosis and in horizontal gene transfer in M. smegmatis. Both strains harbored a circular extra-chromosomal structure, with T7SS system elements related to those found in plasmids from slow growing mycobacteria. It is circular, has 97 kbp and 121 open reading frames. Additionally, only F1725 strain has an IncP multidrug resistant plasmid, named pBRA100. It confers resistance to kanamycin, quaternary ammonium, sulfamethoxazole and streptomycin and also has new fosfomycin resistance gene fos I. No differences were found in the chromosomes that could justify tolerance to glutaraldehyde. The remarkable difference between the two genomes is the presence of plasmid pBRA100 in the glutaraldehyde tolerant strain