Evaluation of bacterial growth inhibition by mercaptopropionic acid in metallo-β-lactamase detection on multidrug-resistant Acinetobacter baumannii

INTRODUCTION: Metallo-β-lactamase (MBL) has been reported all over the world. METHODS: The inhibitory effect of mercaptopropionic acid (MPA) on bacterial growth was evaluated by comparison between disk diffusion and broth dilution methodology with determination of the minimum inhibitory concentration (MIC) for multidrug-resistant Acinetobacter baumanni strains. RESULTS: MPA significantly inhibited growth of the strains. CONCLUSIONS: The use of MPA can affect the results in phenotypic methods of MBL detection.

Bacterial meningitis and living conditions

INTRODUCTION: Bacterial meningitis has great social relevance due to its ability to produce sequelae and cause death. It is most frequently found in developing countries, especially among children. Meningococcal meningitis occurs at a high frequency in populations with poor living conditions. This study describes the temporal evolution of bacterial meningitis in Salvador, Brazil, 1995-2009, and verifies the association between its spatial variation and the living conditions of the population. METHODS: This was an ecological study in which the areas of information were classified by an index of living conditions. It examined fluctuations using a trend curve, and the relationship between this index and the spatial distribution of meningitis was verified using simple linear regression. RESULTS: From 1995-2009, there were 3,456 confirmed cases of bacterial meningitis in Salvador. We observed a downward trend during this period, with a yearly incidence of 9.1 cases/100,000 population and fatality of 16.7%. Children aged <5 years old and male were more affected. There was no significant spatial autocorrelation or pattern in the spatial distribution of the disease. The areas with the worst living conditions had higher fatality from meningococcal disease (β = 0.0078117, p < 0.005). CONCLUSIONS: Bacterial meningitis reaches all social strata; however, areas with poor living conditions have a greater proportion of cases that progress to death. This finding reflects the difficulties for ready access and poor quality of medical care faced by these populations.

Avaliação da monitorização diária da proteína C-Reativa no doente crítico

RESUMO: O objectivo desta Tese de Doutoramento foi estudar o valor da Proteína CReactiva(PCR) como marcador de infecção e sepsis. Por definição, um marcador da infecção não está presente se o doente não está infectado, deve aparecer concomitantemente ou idealmente preceder a instalação da infecção, deve desaparecer com a instituição de terapêutica antimicrobiana adequada e permanecer elevado se a infecção for refractária ao tratamento. Do ponto de vista biológico, a PCR é o protótipo das proteínas de fase aguda, com uma marcada elevação da sua concentração sérica em resposta a diversos estímulos inflamatórios em particular infecções bacterianas. A sua concentração sérica depende apenas da intensidade do estímulo e da velocidade de síntese hepática, não sendo influenciada por nenhum factor ou tratamento a não ser que este tenha influência directa sobre o estímulo desencadeante, o que a torna um marcador de infecção com grande potencial. Nesta Tese comparou-se a PCR com marcadores clássicos de infecção, temperatura e contagem leucocitária, em diversas situações clínicas analisando doentes com infecções documentadas e doentes controlos, sem infecção. Globalmente os resultados dos trabalhos desta Tese mostram que a PCR é um bom marcador de infecção de acordo com a definição previamente apresentada. Em conjunto com a restante avaliação clínica e laboratorial, a monitorização diária da PCR nos doentes sem infecção mostrou ser útil como sentinela da infecção, isto é, apresenta valores baixos nos doentes sem infecção e sobe precocemente nos doentes que desenvolvem uma infecção. Nos doentes com infecção documentada revelou um ser bom marcador de resposta à terapêutica e evolução clínica, diminuindo naqueles que melhoravam e persistindo elevada nos que tinham mau prognóstico, bem assim como identificar diferentes perfis evolutivos. Em suma, a monitorização diária da PCR mostrou utilidade ao longo de todo o internamento na Unidade de Cuidados Intensivos, quer na presença quer na ausência de infecção. Deste todo, a monitorização diária da PCR pode a possibilitar uma utilização mais racional e judiciosa da terapêutica antimicrobiana, contribuindo dessa forma para uma diminuição da toxicidade e da pressão antibiótica, menor risco de emergência de resistências e finalmente diminuição dos custos. Uma vez que, os doentes internados nas Unidades de Cuidados Intensivos apresentam as mesmas doenças que os restantes doentes admitidos no hospital apenas se distinguindo pela sua maior gravidade, poder-se-á extrapolar que a PCR também é potencialmente um bom marcador de infecção nestes doentes. ----------------ABSTRACT: The aim of this PhD Thesis was to assess the value of C-Reactive Protein (CRP) as a marker of infection and sepsis. A marker of infection should be absent in a non-infected patient, should increase alongside or ideally precede the development of an infection, and finally should assess the therapeutic response, that is to say decrease or even disappear with adequate antimicrobial therapy or on the opposite remain elevated if the infection is refractory to the prescribed treatment. The biology of CRP makes it the prototype of acute phase proteins, with marked and sharp elevations of its serum concentration in response to several inflammatory stimulus in particular bacterial infections. Besides, CRP level depends only of the intensity of the stimulus and the rate of hepatic synthesis. Its concentration is not modified by any therapy or intervention. Only those interventions affecting the inflammatory process responsible for the acute phase reaction can change the CRP level. These properties make CRP a potentially good marker of infection. In this Thesis the value of CRP was studied in comparison to traditional markers of infection, like temperature and white cell count, in different clinical situations analysing patients with documented infections and a control group without infection. The aggregated results of the analysis presented in this Thesis illustrate that CRP could be used as a marker of infection. In conjunction with other clinical and laboratory manifestations of sepsis, daily CRP measurement in patients without infection was useful in prediction of infection as its concentration remains low in patients without infection whereas if an infection appears its levels raise markedly. In addition, in patients with documented infections CRP was useful as a marker of therapeutic response and follow-up, with marked decreases in patients with good outcome and remaining elevated in those with poor prognosis, as well as the recognition of different patterns of evolution. In summary, daily CRP measurement was helpful in critical ill patients along the entire Intensive Care Unit stay, both in the presence and in the absence of infection. As a result, daily CRP measurement can assure a better and more rational use of antibiotics and consequently contribute to a decrease in the antibiotic toxicity and demand, reducing the risks of emergence of resistant strains aas well as costs. Provided that patients admitted to an Intensive Care Unit presented the same clinical diagnosis as those admitted to the wards but with higher severity, one can speculate that CRP is also a potentially good marker of infection in these of patients.

Ants in a hospital environment and their potential as mechanical bacterial vectors

Introduction We studied the richness and abundance of ant species, their bacteria and the bacteria isolated from patient clinical samples. Methods Ants were collected with baited traps at 64 sites in a public hospital in São Luis, State of Maranhão, Brazil. Results In total, 1,659 ants from 14 species were captured. The most frequent species were Crematogaster victima, Solenopsis saevissima, Tapinoma melanocephalum, Camponotus vittatus and Paratrechina fulva. Forty-one species of bacteria were isolated from the ants and 18 from patients. Conclusions Ants are potential vehicles for pathogenic and opportunistic bacteria, and they can represent a risk factor in nosocomial infections.

Performance of the Vitek 2 system software version 5.03 in the bacterial identification and antimicrobial susceptibility test: evaluation study of clinical and reference strains of Gram-positive cocci

Introduction. The genera Enterococcus, Staphylococcus and Streptococcus are recognized as important Gram-positive human pathogens. The aim of this study was to evaluate the performance of Vitek 2 in identifying Gram-positive cocci and their antimicrobial susceptibilities. Methods. One hundred four isolates were analyzed to determine the accuracy of the automated system for identifying the bacteria and their susceptibility to oxacillin and vancomycin. Results. The system correctly identified 77.9% and 97.1% of the isolates at the species and genus levels, respectively. Additionally, 81.8% of the Vitek 2 results agreed with the known antimicrobial susceptibility profiles. Conclusion. Vitek 2 correctly identified the commonly isolated strains; however, the limitations of the method may lead to ambiguous findings.

Colistin and anti-Gram-positive bacterial agents against Acinetobacter baumannii

Introduction Acinetobacter baumannii has attained an alarming level of resistance to antibacterial drugs. Clinicians are now considering the use of older agents or unorthodox combinations of licensed drugs against multidrug-resistant strains to bridge the current treatment gap. We investigated the in vitro activities of combination treatments that included colistin with vancomycin, norvancomycin or linezolid against multidrug-resistant Acinetobacter baumannii. Methods The fractional inhibitory concentration index and time-kill assays were used to explore the combined effects of colistin with vancomycin, norvancomycin or linezolid against 40 clinical isolates of multidrug-resistant Acinetobacter baumannii. Transmission electron microscopy was performed to evaluate the interactions in response to the combination of colistin and vancomycin. Results The minimum inhibitory concentrations (MICs) of vancomycin and norvancomycin for half of the isolates decreased below the susceptibility break point, and the MIC of linezolid for one isolate was decreased to the blood and epithelial lining fluid concentration using the current dosing regimen. When vancomycin or norvancomycin was combined with subinhibitory doses of colistin, the multidrug-resistant Acinetobacter baumannii test samples were eradicated. Transmission electron microscopy revealed that subinhibitory doses of colistin were able to disrupt the outer membrane, facilitating a disruption of the cell wall and leading to cell lysis. Conclusions Subinhibitory doses of colistin significantly enhanced the antibacterial activity of vancomycin, norvancomycin, and linezolid against multidrug-resistant Acinetobacter baumannii.

Post-splenectomy infections in chronic schistosomiasis as a consequence of bacterial translocation

INTRODUCTION : Bacterial translocation is the invasion of indigenous intestinal bacteria through the gut mucosa to normally sterile tissues and internal organs. Schistosomiasis may cause alterations in the immune system and damage to the intestines, portal system and mesenteric lymph nodes. This study investigated bacterial translocation and alterations in the intestinal microbiota and mucosa in schistosomiasis and splenectomized mice. METHODS : Forty female 35-day-old Swiss Webster mice were divided into the following four groups with 10 animals each: schistosomotic (ESF), splenectomized schistosomotic (ESEF), splenectomized (EF) and control (CF). Infection was achieved by introduction of 50 Schistosoma mansoni (SLM) cercariae through the skin. At 125 days after birth, half of the parasitized and unparasitized mice were subjected to splenectomy. Body weights were recorded for one week after splenectomy; then, the mice were euthanized to study bacterial translocation, microbiota composition and intestinal morphometry. RESULTS : We observed significant reductions in the weight increases in the EF, ESF and ESEF groups. There were increases of at least 1,000 CFU of intestinal microbiota bacteria in these groups compared with the CF. The EF, ESF and ESEF mice showed decreases in the heights and areas of villi and the total villus areas (perimeter). We observed frequent co-infections with various bacterial genera. CONCLUSIONS : The ESEF mice showed a higher degree of sepsis. This finding may be associated with a reduction in the immune response associated with the absence of the spleen and a reduction in nutritional absorption strengthened by both of these factors (Schistosoma infection and splenectomy).

Aerobic bacterial profile and antibiotic resistance in patients with diabetic foot infections

ABSTRACTINTRODUCTION: This study aimed to determine the frequencies of bacterial isolates cultured from diabetic foot infections and assess their resistance and susceptibility to commonly used antibiotics.METHODS: This prospective study included 41 patients with diabetic foot lesions. Bacteria were isolated from foot lesions, and their antibiotic susceptibility pattern was determined using the Kirby-Bauer disk diffusion method and/or broth method [minimum inhibitory concentration (MIC)].RESULTS: The most common location of ulceration was the toe (54%), followed by the plantar surface (27%) and dorsal portion (19%). A total of 89 bacterial isolates were obtained from 30 patients. The infections were predominantly due to Gram-positive bacteria and polymicrobial bacteremia. The most commonly isolated Gram-positive bacteria were Staphylococcus aureus, followed by Staphylococcus saprophyticus, Staphylococcus epidermidis, Streptococcus agalactiae, and Streptococcus pneumoniae. The most commonly isolated Gram-negative bacteria were Proteus spp. and Enterobacterspp., followed by Escherichia coli, Pseudomonasspp., and Citrobacterspp. Nine cases of methicillin-resistant Staphylococcus aureus (MRSA) had cefoxitin resistance, and among these MRSA isolates, 3 were resistant to vancomycin with the MIC technique. The antibiotic imipenem was the most effective against both Gram-positive and Gram-negative bacteria, and gentamicin was effective against Gram-negative bacteria.CONCLUSIONS: The present study confirmed the high prevalence of multidrug-resistant pathogens in diabetic foot ulcers. It is necessary to evaluate the different microorganisms infecting the wound and to know the antibiotic susceptibility patterns of the isolates from the infected wound. This knowledge is crucial for planning treatment with the appropriate antibiotics, reducing resistance patterns, and minimizing healthcare costs.

Bacterial cellulose as a novel substrate for retinal pigment epithelium cells transplantation in age-related macular degeneration

Tese de Doutoramento em Engenharia Química e Biológica.

The influence of biofilm formation by Gardnerella vaginalis and other anaerobes on bacterial vaginosis

Bacterial vaginosis (BV) is the worldwide leading vaginal disorder in women of reproductive age. BV is characterized by the replacement of beneficial lactobacilli and the augmentation of anaerobic bacteria. Gardnerella vaginalis is a predominant bacterial species, however, BV is also associated with other numerous anaerobes, such as Atopobium vaginae, Mobiluncus mulieris, Prevotella bivia, Fusobacterium nucleatum and Peptoniphilus sp.. Currently, the role of G. vaginalis in the etiology of BV remains a matter of controversy. It is however known that, in BV patients, a biofilm is usually formed on the vaginal epithelium and G. vaginalis is typically the predominant species. So, the current paradigm is that the establishment of a biofilm plays a key role in the pathogenesis of BV. This review provides background on the influence of biofilm formation by G. vaginalis and other anaerobes in the polymicrobial etiology of BV, through its initial adhesion until biofilm formation and discusses the commensal and synergic interactions established between them to understand the phenotypic shift of G. vaginalis' biofilms into BV establishment.

Engineering modular viral scaffolds for targeted bacterial population editing

Bacteria are central to human health and disease, but existing tools to edit microbial consortia are limited. For example, broad-spectrum antibiotics are unable to precisely manipulate bacterial communities. Bacteriophages can provide highly specific targeting of bacteria, but assembling well-defined phage cocktails solely with natural phages can be a time-, labor- and cost-intensive process. Here, we present a synthetic biology strategy to modulate phage host ranges by engineering phage genomes in Saccharomyces cerevisiae. We used this technology to redirect Escherichia coli phage scaffolds to target pathogenic Yersinia and Klebsiella bacteria, and conversely, Klebsiella phage scaffolds to target E. coli by modular swapping of phage tail components. The synthetic phages achieved efficient killing of their new target bacteria and were used to selectively remove bacteria from multi-species bacterial communities with cocktails based on common viral scaffolds. We envision this approach accelerating phage biology studies and enabling new technologies for bacterial population editing.

Bacterial vaginosis biofilms: challenges to current therapies and emerging solutions

Bacterial vaginosis (BV) is the most common genital tract infection in women during their reproductive years and it has been associated with serious health complications, such as preterm delivery and acquisition or transmission of several sexually transmitted agents. BV is characterized by a reduction of beneficial lactobacilli and a significant increase in number of anaerobic bacteria, including Gardnerella vaginalis, Atopobium vaginae, Mobiluncus spp., Bacteroides spp. and Prevotella spp.. Being polymicrobial in nature, BV etiology remains unclear. However, it is certain that BV involves the presence of a thick vaginal multi-species biofilm, where G. vaginalis is the predominant species. Similar to what happens in many other biofilm-related infections, standard antibiotics, like metronidazole, are unable to fully eradicate the vaginal biofilm, which can explain the high recurrence rates of BV. Furthermore, antibiotic therapy can also cause a negative impact on the healthy vaginal microflora. These issues sparked the interest in developing alternative therapeutic strategies. This review provides a quick synopsis of the currently approved and available antibiotics for BV treatment while presenting an overview of novel strategies that are being explored for the treatment of this disorder, with special focus on natural compounds that are able to overcome biofilm-associated antibiotic resistance.