Assessment of central venous catheter-associated infections using semi-quantitative or quantitative culture methods

Semiquantitative (Maki) and quantitative (Brun- Buisson) culture techniques were employed in the diagnosis of catheter-related bloodstream infections (CRBSI) in patients who have a short-term central venous catheter (inserted for 30 days). The diagnosis of CRBSI was based on the results of semiquantitative and quantitative culture of material from the removed catheters. Catheter tips (118) from 100 patients were evaluated by both methods. Semiquantitative analysis revealed 34 catheters (28.8%) colonized by ≥15 colonyforming units (cfu), while quantitative cultures (34 catheters, 28.8%) showed the growth of ≥103 cfu/mL. Bacteremia was confirmed in four patients by isolating microorganisms of identical species from both catheters and blood samples. Using the semiquantitative culture technique on short-term central venous catheter tips, we have shown that with a cut-off level of ≥15 cfu, the technique had 100.0% sensitivity, specificity of 68.4%, 25.0% positive predictive value (PPV) and 100.0% negative predictive value (NPV), efficiency of 71.4% and a prevalence of 9.5%. The quantitative method, with a cut-off limit of ≥103 cfu/mL, gave identical values: the sensitivity was 100.0%, specificity 68.4%, positive predictive value (PPV) 25.0%, negative predictive value (NPV) 100.0%, efficiency 71.4% and prevalence 9.5%. We concluded that the semiquantitative and quantitative culture methods, evaluated in parallel, for the first time in Brazil, have similar sensitivity and specificity. Keywords: central venous catheter; semi-quantitative culture; quantitative culture; catheter-related bacteremia.

Diagnostic accuracy of semi-quantitative and quantitative culture techniques for the diagnosis of catheter-related infections in newborns and molecular typing of isolated microorganisms

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

Cough-generated aerosols of Pseudomonas aeruginosa and other Gram Negative Bacteria from cystic fibrosis patients.

Background: Pseudomonas aeruginosa is the most common bacterial pathogen in cystic fibrosis (CF) patients. Current infection control guidelines aim to prevent transmission via contact and respiratory droplet routes and do not consider the possibility of airborne transmission. We hypothesized that with coughing, CF subjects produce viable, respirable bacterial aerosols. Methods: Cross-sectional study of 15 children and 13 adults with CF, 26 chronically infected with P. aeruginosa. A cough aerosol sampling system enabled fractioning of respiratory particles of different size, and culture of viable Gram negative non-fermentative bacteria. We collected cough aerosols during 5 minutes voluntary coughing and during a sputum induction procedure when tolerated. Standardized quantitative culture and genotyping techniques were used. Results: P. aeruginosa was isolated in cough aerosols of 25 (89%) subjects of whom 22 produced sputum samples. P. aeruginosa from sputum and paired cough aerosols were indistinguishable by molecular typing. In 4 cases the same genotype was isolated from ambient room air. Approximately 70% of viable aerosols collected during voluntary coughing were of particles ≤ 3.3 microns aerodynamic diameter. P. aeruginosa, Burkholderia cenocepacia Stenotrophomonas maltophilia and Achromobacter xylosoxidans were cultivated from respiratory particles in this size range. Positive room air samples were associated with high total counts in cough aerosols (P=0.003). The magnitude of cough aerosols were associated with higher FEV1 (r=0.45, P=0.02) and higher quantitative sputum culture results (r=0.58, P=0.008). Conclusion: During coughing, CF patients produce viable aerosols of P. aeruginosa and other Gram negative bacteria of respirable size range, suggesting the potential for airborne transmission.

Cough-generated aerosols of Pseudomonas aeruginosa and other Gram-negative bacteria from patients with cystic fibrosis

BACKGROUND: Pseudomonas aeruginosa is the most common bacterial pathogen in patients with cystic fibrosis (CF). Current infection control guidelines aim to prevent transmission via contact and respiratory droplet routes and do not consider the possibility of airborne transmission. It was hypothesised that subjects with CF produce viable respirable bacterial aerosols with coughing.

METHODS: A cross-sectional study was undertaken of 15 children and 13 adults with CF, 26 chronically infected with P aeruginosa. A cough aerosol sampling system enabled fractioning of respiratory particles of different sizes and culture of viable Gram-negative non-fermentative bacteria. Cough aerosols were collected during 5 min of voluntary coughing and during a sputum induction procedure when tolerated. Standardised quantitative culture and genotyping techniques were used.

RESULTS: P aeruginosa was isolated in cough aerosols of 25 subjects (89%), 22 of whom produced sputum samples. P aeruginosa from sputum and paired cough aerosols were indistinguishable by molecular typing. In four cases the same genotype was isolated from ambient room air. Approximately 70% of viable aerosols collected during voluntary coughing were of particles <or=3.3 microm aerodynamic diameter. P aeruginosa, Burkholderia cenocepacia, Stenotrophomonas maltophilia and Achromobacter xylosoxidans were cultivated from respiratory particles in this size range. Positive room air samples were associated with high total counts in cough aerosols (p = 0.003). The magnitude of cough aerosols was associated with higher forced expiratory volume in 1 s (r = 0.45, p = 0.02) and higher quantitative sputum culture results (r = 0.58, p = 0.008).

CONCLUSION: During coughing, patients with CF produce viable aerosols of P aeruginosa and other Gram-negative bacteria of respirable size range, suggesting the potential for airborne transmission.

Perfil de resistencia microbiológico en ciudados intensivos adultos en la fundación Santa Fe de Bogotá año 2014

Se realizó un estudio descriptivo, retrospectivo; se usó la base de datos de los aislamientos microbiológicos documentados en las UCI de la Fundación Santa fe de Bogotá para el año 2014. La prevalencia de bacterias resistentes en los aislamientos de la FSFB no es baja, por lo que se requiere una terapia empírica acertada acorde con la flora local. Se requieren estudios analíticos para evaluar factores asociados al desarrollo de gérmenes multi resistentes y mortalidad por sepsis

Nutrição parenteral como fator de risco para infecção relacionada a cateter venoso central

O uso de cateteres venosos centrais (CVC) para fins diagnósticos e terapêuticos está incorporado à prática médica diária. Complicações sérias, com elevada morbidade e mortalidade, como a sepse, estão associadas a este procedimento. O diagnóstico das infecções relacionadas a cateter é fundamentado em sinais clínicos e laboratoriais. Os fatores de risco para infecção devem ser considerados por ocasião da utilização de CVC e estão relacionados ao paciente, ao cateter, ao tipo de solução administrada, ao profissional que manipula o cateter e ao agente etiológico. A identificação destes fatores permite a intervenção precoce sobre os mesmos e o manejo adequado do CVC e das complicações clínicas relacionadas.

Prevalência de bacteriúria assintomática em crianças durante a idade pré-escolar no município de Araraquara-SP

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Diagnóstico das infecções relacionadas a cateter em recém-nascidos e epidemiologia molecular dos Staphylococcus spp. isolados

Pós-graduação em Biologia Geral e Aplicada - IBB

Comparação das técnicas semi-quantitativa e quantitativa nas infecções relacionadas a cateter em neonatos e determinação da resistência à oxacilina nos Staphylococcus spp

Study the semi-quantitative and quantitative technique in the diagnosis of catheter-related infections in newborns and to determine oxacillin resistance in Staphylococcus isolated. It was analyzed 353 catheter tips from 273 newborns in the Neonatal Unit of Hospital FMB. To confirm the diagnosis of infection, were analyzed the clinical data of newborns, the presence of at least one positive blood culture and growth of ≥ 1000 CFU/mL on quantitative culture and/or ³15 UFC on semiquantitative culture, with the same microorganism isolation (species and drug sensitivity) in blood culture and no other focus of infection except the catheter. The disk diffusion technique was used to check similarity of strains and resistance to oxacillin. Of the 353 tips analyzed, 39 were included in this study as the inclusion criteria. The semiquantitative culture was positive in 26 (66.7%) catheters and quantitative culture was positive in 24 (61.5%). Of 273 patients, 19 (6.9%) had a diagnosis of catheter-related bloodstream Infection (CR-BSI). Of the 19 episodes of CR-BSI, S. epidermidis was the predominant etiological agent (84.2%). The resistance to the antibiotic methicillin was found in 14 (73.7%) strains of Staphylococcus. The semiquantitative method was more sensitive (79%) compared with the quantitative method (63%). The use of antibiotics may have influenced the sensitivity of the quantitative method as the microorganisms present in the lumen are exposed to higher concentrations of antibiotics administered via the catheter

Comparison of real-time PCR assays for detection, quantification, and differentiation of campylobacter jejuni and campylobacter coli in broiler neck skin samples

We tested the use of multiplex real-time PCR for detection and quantification of Campylobacter jejuni and Campylobacter coli on broiler carcass neck skin samples collected during 2008 from slaughterhouses in Switzerland. Results from an established TaqMan assay based on two different targets (hipO and ceuE for C. jejuni and C. coli, respectively) were corroborated with data from a newly developed assay based on a single-nucleotide polymorphism in the fusA gene, which allows differentiation between C. jejuni and C. coli. Both multiplex real-time PCRs were applied simultaneously for direct detection, differentiation, and quantification of Campylobacter from 351 neck skin samples and compared with culture methods. There was good correlation in detection and enumeration between real-time PCR results and quantitative culture, with real-time PCR being more sensitive. Overall, 251 (71.5%) of the samples were PCR positive for Campylobacter, with 211 (60.1%) in the hipO-ceuE assays, 244 (69.5%) in the fusA assay, and 204 (58.1%) of them being positive in both PCR assays. Thus, the fusA assay was similarly sensitive to the enrichment culture (72.4% positive); however, it is faster and allows for quantification. In addition, real-time PCR allowed for species differentiation; roughly 60% of positive samples contained C. jejuni, less than 10% C. coli, and more than 30% contained both species. Real-time PCR proved to be a suitable method for direct detection, quantification, and differentiation of Campylobacter from carcasses, and could permit time-efficient surveillance of these zoonotic agents.

Comparison of conventional PCR, quantitative PCR, bacteriological culture and the Warthin Starry technique to detect Leptospira spp. in kidney and liver samples from naturally infected sheep from Brazil

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

Validation of a new proposal of quantitative sperm culture for industrialized bull semen

The Brazilian Ministry of Agriculture (MAPA) has discussed the mandatory culture of industrialized semen, both to ensure biosefety, and to prevent in vitro fertilization problems caused by oocyte contamination with ubiquitous and opportunistic bacteria from preputial microbiota. Pour plate, a quantitative technique recommended by the World Organization for Animal Health (OIE), is operationally difficult and costly for routine analysis in Artificial Insemination Centers (AICs). The objectives of this study were to evaluate and validate viable superficial bacteria counts (VSBC), in CFU/mL, compared with pour plate counts, in industrialized bull semen samples from AICs. Semen straws from Projeto Hungria - MAPA bulls were used. VSBC and pour plate were carried out in parallel in serial dilutions of the samples, from 10(-1) to 10(-5). CFU/mL means or medians recorded in each dilution and technique were compared, and no statistical differences were observed between the two techniques regarding the quantification of bacteria in CFU/mL, suggesting that it may be possible to replace pour plate for CBSV, a cheaper and more practical technique.

A quantitative approach for understanding small-scale human mesenchymal stem cell culture - implications for large-scale bioprocess development

Human mesenchymal stem cell (hMSC) therapies have the potential to revolutionise the healthcare industry and replicate the success of the therapeutic protein industry; however, for this to be achieved there is a need to apply key bioprocessing engineering principles and adopt a quantitative approach for large-scale reproducible hMSC bioprocess development. Here we provide a quantitative analysis of the changes in concentration of glucose, lactate and ammonium with time during hMSC monolayer culture over 4 passages, under 100% and 20% dissolved oxgen (dO2), where either a 100%, 50% or 0% growth medium exchange was performed after 72h in culture. Yield coefficients, specific growth rates (h-1) and doubling times (h) were calculated for all cases. The 100% dO2 flasks outperformed the 20% dO2 flasks with respect to cumulative cell number, with the latter consuming more glucose and producing more lactate and ammonium. Furthermore, the 100% and 50% medium exchange conditions resulted in similar cumulative cell numbers, whilst the 0% conditions were significantly lower. Cell immunophenotype and multipotency were not affected by the experimental culture conditions. This study demonstrates the importance of determining optimal culture conditions for hMSC expansion and highlights a potential cost savings from only making a 50% medium exchange, which may prove significant for large-scale bioprocessing. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

The relationship of E. coli and total coliform culture to the quantitative molecular detection of various fecal indicators, sources, and pathogens in private drinking water wells

Water remains a predominant vector for human enteric pathogens not just for developing countries but also developed nations, where numerous infectious disease outbreaks, linked to the contamination of drinking water have been documented. Private drinking water wells are a source of drinking water that is largely unstudied even though a significant percentage of the population in Ontario relies on wells as their primary water source. As there exists little to no systematic surveillance for enteric infections or outbreaks related to well water sources, these individuals may be at higher risk of waterborne infectious diseases. The relationships between various fecal indicators in the water of private drinking water wells, including E. coli, Total Coliforms (TC) and Bacteroides, and enteric pathogens, including Campylobacter jejuni, Salmonella spp., and Shiga toxin producing E. coli, were studied. Convenience private well water samples collected from various regions of interest during the summer of 2014 underwent membrane filtration and culture to determine quantities of E. coli and TC colony forming units. 289 E. coli positive and 230 TC-only waters were successfully analyzed by individual qPCR assays for the aforementioned enteric pathogens. Microbial source tracking methods targeted to specific Bacteroides were used to determine the source of fecal contamination as either human or bovine. The source of fecal contamination varied by geographic region and is thought to be due to such things as differences in septic tank density and underlying geology, among others. Fecal indicators, E. coli and Bacteroides, were significantly correlated. E. coli as measured by qPCR was more strongly correlated to both total and human-specific Bacteroides genetic markers than culturable E. coli. Lastly, 1.9% of samples showed molecular evidence of contamination with enteric pathogens. Although low, this finding is significant given the limited volume of water available for testing, and suggests a potential health risk to consumers. Knowing the extent of contamination, as well as the biologic source, can better inform risk assessment and the development of potential intervention strategies for private well water in specific regions of Ontario.