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1.
Western Pac Surveill Response J ; 11(1): 41-46, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32963890

RESUMO

Problem: Emerging bacterial antimicrobial (antibiotic) resistance (AMR) is a global threat to human health. However, most lower income countries do not have microbiological diagnostic testing for prompt, reliable confirmation of bloodstream infection and identification of AMR. Context: Clinicians in Pacific island nations are increasingly challenged by patients who have infection due to antimicrobial-resistant bacteria. Treatment of infection remains empirical because of a lack of diagnostic testing capacity and may follow guidelines that were formulated without reference to local measures of AMR prevalence. There is limited understanding among clinicians of microbiology testing and test interpretation. Action: Examine the lessons learnt from pilot laboratory development programmes in two Pacific island nations that focused on establishing standard procedures for micrological diagnostics and antimicrobial susceptibility testing (AST) and on improving the training of clinicians to increase their use of laboratory services. Outcome: The pilot programmes addressed a range of logistical difficulties and evaluated two blood culture systems. They also examined and improved internal QC implementation and evaluated the prevalence of AMR. Discussion: Continued development of microbiological diagnostic capability in the Pacific region is paramount. Pacific Island nations need to develop the capability of at least one central laboratory to culture AMR pathogens and subject them to quality-controlled AST or arrange for suitable referral to a nearby country. Discussion: This study demonstrated a persistently high prevalence of three major bacterial STIs across four countries in WHO's Western Pacific Region during nearly two decades. Further strengthening of strategies to control and prevent STIs is warranted.


Assuntos
Antibacterianos/farmacologia , Técnicas Bacteriológicas/normas , Farmacorresistência Bacteriana , Testes de Sensibilidade Microbiana/normas , Técnicas de Diagnóstico Molecular/normas , Humanos , Ilhas do Pacífico , Projetos Piloto , Controle de Qualidade
2.
PLoS One ; 15(8): e0237655, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32810167

RESUMO

BACKGROUND: Several tests are available for plague confirmation but bacteriological culture with Yersinia pestis strain isolation remains the gold standard according to the World Health Organization. However, this is a time consuming procedure; requiring specific devices and well-qualified staff. In addition, strain isolation is challenging if antibiotics have been administered prior to sampling. Here, we developed a loop-mediated isothermal amplification (LAMP) technique, a rapid, simple, sensitive and specific technique that would be able to detect Y. pestis in human biological samples. METHODS: LAMP primers were designed to target the caf1 gene which is specific to Y. pestis. The detection limit was determined by testing 10-fold serial dilution of Y. pestis DNA. Cross-reactivity was tested using DNA extracts from 14 pathogens and 47 residual samples from patients suffering from non-plague diseases. Specificity and sensitivity of the LAMP caf1 were assessed on DNA extracts of 160 human biological samples. Then, the performance of the LAMP caf1 assay was compared to conventional PCR and bacteriological culture. RESULTS: The detection limit of the developed Y. pestis LAMP assay was 3.79 pg/µl, similar to conventional PCR. The result could be read out within 45 min and as early as 35 minutes in presence of loop primer, using a simple water bath at 63°C. This is superior to culture with respect to time (requires up to 10 days) and simplicity of equipment compared to PCR. Furthermore, no cross-reactivity was found when tested on DNA extracts from other pathogens and human biological samples from patients with non-plague diseases. Compared to the gold standard, LAMP sensitivity and specificity were 97.9% (95% CI: 89.1%-99.9%) and 94.6% (95% CI: 88.6%-97.9%), respectively. CONCLUSION: LAMP detected Y. pestis effectively with high sensitivity and specificity in human plague biological samples. It can potentially be used in the field during outbreaks in resource limited countries such as Madagascar.


Assuntos
Técnicas Bacteriológicas/métodos , DNA Bacteriano/isolamento & purificação , Técnicas de Amplificação de Ácido Nucleico/métodos , Peste/diagnóstico , Yersinia pestis/isolamento & purificação , Técnicas Bacteriológicas/economia , Estudos de Viabilidade , Humanos , Limite de Detecção , Madagáscar , Técnicas de Amplificação de Ácido Nucleico/economia , Peste/microbiologia , Fatores de Tempo , Yersinia pestis/genética
3.
PLoS One ; 15(8): e0236700, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32750088

RESUMO

Mycobacterial culture remains the gold standard for the diagnosis of tuberculosis. However, an appropriate digestion and decontamination method (DDM) is essential for the effective recovery of tubercle bacilli in culture. Therefore, the current study was designed to compare the performance of papain-cetylpyridinium chloride [papain-CPC] and pepsin-cetylpyridinium chloride [pepsin-CPC] DDMs against N-acetyl L-Cysteine-sodium hydroxide (NALC-NaOH) DDM for recovery of mycobacteria from clinically suspected pulmonary tuberculosis cases. To evaluate papain-CPC, pepsin-CPC and NALC-NaOH DDMs, sputum samples (N = 1381) were cultured on Löwenstein-Jensen medium and the results were compared. The papain-CPC DDM showed sensitivity, specificity, positive predictive value, and negative predictive value of 100%, 93.27%, 71.7%, and 100%, respectively as compared to NALC-NaOH DDM. Similarly, pepsin-CPC DDM demonstrated sensitivity, specificity, positive predictive value and negative predictive value of 98.94%, 94.7%, 76.11%, and 99.81%, respectively. In summary, both papain-CPC and pepsin-CPC DDMs are highly sensitive and specific techniques for recovery of mycobacteria as compared to NALC-NaOH DDM. However, when the overall performances of all DDMs compared, papain-CPC DDM isolated increased number of mycobacterial isolates with comparatively higher numbers of colonies on LJ media than both pepsin-CPC and NALC-NaOH DDMs, indicating its potential to replace the NALC-NaOH DDM for recovery of mycobacteria from sputum samples.


Assuntos
Técnicas Bacteriológicas/métodos , Meios de Cultura/química , Mycobacterium tuberculosis/isolamento & purificação , Escarro/microbiologia , Tuberculose Pulmonar/microbiologia , Acetilcisteína/química , Cetilpiridínio/química , Humanos , Papaína/química , Pepsina A/química , Sensibilidade e Especificidade , Manejo de Espécimes
4.
Rev Bras Epidemiol ; 23: e200079, 2020.
Artigo em Inglês, Português | MEDLINE | ID: mdl-32696931

RESUMO

OBJECTIVE: This study aimed to investigate the factors associated with the outcomes of recovery and abandonment in the incarcerated population with tuberculosis. METHODS: A quantitative and observational analytical study was performed with data from the Notification Disease Information System (Sinan), tuberculosis data from the incarcerated population in the state of Paraiba from 2007 to 2016; Notifications of individuals over the age of 18, reported as "new cases" and the outcome, "recovery" or "abandonment" status were included. Those people who until December 2016 had no outcome information were excluded. Analyses were performed using bivariate and multivariate statistics from the Poisson regression. RESULTS: Of the 614 notifications, most were male (93.8%). In the bivariate analysis, there was a statistically relevant association of outcomes with Acquired Immunodeficiency Syndrome (p = 0.044), Human Immunodeficiency Virus (HIV) serology (p = 0.048) and lack of completion of follow-up bacilloscopy (p = 0.001). In the adjusted multivariate analysis, Acquired Immunodeficiency Syndrome (RR = 1.998; 95%CI 1.078 - 3.704; p = 0.028) and lack of completion of follow-up bacilloscopy (RR = 5.251; 95%CI 2.158 - 12.583; p <0.001*) remained significantly associated with the dropout outcome. CONCLUSION: Recovery and abandonment outcomes were mainly associated with whether the follow-up bacilloscopy was performed or not and Acquired Immunodeficiency Syndrome.


Assuntos
Pacientes Desistentes do Tratamento/estatística & dados numéricos , Prisioneiros/estatística & dados numéricos , Tuberculose/terapia , Síndrome de Imunodeficiência Adquirida/epidemiologia , Adulto , Técnicas Bacteriológicas/estatística & dados numéricos , Brasil/epidemiologia , Feminino , Seguimentos , Humanos , Masculino , Resultado do Tratamento
5.
Sheng Wu Gong Cheng Xue Bao ; 36(5): 908-919, 2020 May 25.
Artigo em Chinês | MEDLINE | ID: mdl-32567274

RESUMO

A multiplex PCR method was developed to detect the main pathogens of Qinghai Tibetan sheep endometritis. First, the genomes of five standard bacterial strains were extracted and specific primers were selected; the multiplex PCR method was established by using the genome of the standard strain as a template. The samples were collected by sterile cotton swab from Tibetan sheep uterus, and then placed in LB medium and numbered. After 48 h, the genomes of cultured bacteria were extracted and detected by single PCR method, then the positive samples were recorded. The positive samples detected by single PCR were selected for multiplex PCR detection and recorded again. The coincidence rate between these two methods was calculated to measure the accuracy of multiplex PCR. In order to identify the species of the pathogen, 30 positive samples verified by single and multiplex PCR were randomly selected for bacterial isolation and identification. In the 600 samples, the infected ratio of Streptococcus agalactiae (GBS) was 47.33%, Escherichia coli 34.83%, Staphylococcus aureus 6.5%, Salmonella and Trueperella pyogenes were negatively detected. Among the positive samples detected by multiplex PCR, the positive ratio of GBS was 45.50%, E. coli 33.50%, S. aureus 6.5%. Comparison of two detection results, Multiplex PCR detection coincidence rate is more than 95%. The isolated pathogens were identified as E. coli, GBS and S. aureus, which was consistent with the results of two methods. The multiplex PCR method was successfully established and the main pathogens of endometritis in Qinghai Tibetan sheep were GBS, E. coli and S. aureus.


Assuntos
Bactérias , Técnicas Bacteriológicas , Endometrite , Reação em Cadeia da Polimerase Multiplex , Doenças dos Ovinos , Animais , Bactérias/genética , Bactérias/isolamento & purificação , Técnicas Bacteriológicas/métodos , Endometrite/microbiologia , Endometrite/veterinária , Feminino , Reação em Cadeia da Polimerase Multiplex/normas , Reação em Cadeia da Polimerase/veterinária , Sensibilidade e Especificidade , Ovinos , Doenças dos Ovinos/microbiologia , Tibet
6.
Cochrane Database Syst Rev ; 6: CD012431, 2020 06 04.
Artigo em Inglês | MEDLINE | ID: mdl-32497279

RESUMO

BACKGROUND: Sore throat is a common condition caused by viruses or bacteria, and is a leading cause of antibiotic prescription in primary care. The most common bacterial species is group A streptococcus ('strep throat'). Between 50% to 70% of pharyngitis cases are treated with antibiotics, despite the majority of cases being viral in origin. One strategy to reduce antibiotics is to use rapid tests for group A streptococcus to guide antibiotic prescriptions. Rapid tests can be used alone or in combination with a clinical scoring system. OBJECTIVES: To assess the efficacy and safety of strategies based on rapid tests to guide antibiotic prescriptions for sore throat in primary care settings. SEARCH METHODS: We searched CENTRAL, MEDLINE, Embase, CINAHL, Web of Science, and LILACS, as well as the trial registries ClinicalTrials.gov and the WHO ICTRP on 5 June 2019. SELECTION CRITERIA: We included randomised controlled trials (RCTs) comparing rapid tests with management based on clinical grounds to guide the prescription of antibiotics for people with a sore throat in ambulatory care settings. We included trials that randomised individuals, as well as cluster-RCTs in which individual practitioners (or practices) or emergency departments were randomised. DATA COLLECTION AND ANALYSIS: Two review authors independently extracted data on the primary outcomes (number of participants provided with an antibiotic prescription; number of participants with an antibiotic dispensed) and secondary outcomes (duration of sore throat symptoms; duration of other symptoms; quality of life measures; number of participants with a complication attributed to the index infection; number of participants in need of re-consultation by the end of follow-up; number of participants in need of hospital admission by the end of follow-up; number of satisfied participants; number of participants with an adverse event attributed to the rapid test). We assessed the risk of bias of all included trials and used GRADE to assess the certainty of the evidence. We performed meta-analyses and sensitivity analyses when feasible. MAIN RESULTS: We included five trials (2891 children and adult participants in total; 2545 participants after adjusting for clustering). Management in the intervention group was as follows: in three trials rapid tests were used in combination with a clinical scoring system; in one trial, some physicians were asked to use rapid tests alone, while others were asked to use rapid tests in combination with a clinical scoring system; in one trial, rapid tests were used alone. Based on data from five trials (2545 participants), a large reduction in prescribed antibiotics was found in the rapid test group (481/1197) versus management based on clinical grounds (865/1348), for a summary risk difference (RD) of -25%, 95% confidence interval (CI) -31% to -18%; I2 = 62%; moderate-certainty evidence. Estimates of effect on antibiotic prescription rates were stable in various sensitivity analyses. Based on data from two trials (900 people) originating from the same overarching study, the evidence suggests that rapid tests may not reduce dispensed antibiotic treatments: rapid test group (156/445) versus management based on clinical grounds (197/455); summary RD -7%, 95% CI -17% to 2%; I2 = 53%; low-certainty evidence. Four trials (2075 participants) reported data on the number of participants with a complication attributed to the index infection; the summary odds ratio (OR) was 0.85, 95% CI 0.03 to 26.65; P = 0.93; I2 = 62%; very low-certainty evidence, which means that people in the rapid testing group were less likely to develop complications of the index infection, but the evidence is very uncertain. Two trials (1161 participants) reported on the number of participants in need of re-consultation by the end of follow-up; the summary OR was 1.12, 95% CI 0.57 to 2.21; P = 0.74; I2 = 59%; low-certainty evidence, which means that participants in the rapid testing group were more likely to be in need of re-consultation by the end of the study follow-up, but the evidence is uncertain. Lack of data impeded assessment of other secondary outcomes (including safety outcomes) and of sources of heterogeneity.  AUTHORS' CONCLUSIONS: Rapid testing to guide antibiotic treatment for sore throat in primary care probably reduces antibiotic prescription rates by 25% (absolute risk difference), but may have little or no impact on antibiotic dispensing. More studies are needed to assess the efficacy and safety of rapid test-guided antibiotic prescribing, notably to evaluate patient-centred outcomes and variability across subgroups (e.g. adults versus children).


Assuntos
Antibacterianos/uso terapêutico , Faringite/tratamento farmacológico , Faringite/microbiologia , Infecções Estreptocócicas/diagnóstico , Streptococcus pyogenes/isolamento & purificação , Adulto , Técnicas Bacteriológicas , Criança , Prescrições de Medicamentos/estatística & dados numéricos , Feminino , Humanos , Masculino , Faringite/virologia , Ensaios Clínicos Controlados Aleatórios como Assunto , Infecções Estreptocócicas/microbiologia
7.
J Med Microbiol ; 69(6): 806-811, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32490793

RESUMO

Introduction. Bloodstream infections (BSI) are growing in incidence and present a serious health threat. Most patients wait up to 48 h before microbiological cultures can confirm a diagnosis. Low numbers of circulating bacteria in patients with BSI mean we need to develop new methods and optimize current methods to facilitate efficient recovery of bacteria from the bloodstream. This will allow detection of positive blood cultures in a more clinically useful timeframe. Many bacterial blood recovery methods are available and usually include a combination of techniques such as centrifugation, filtration, serum separation or lysis treatment. Here, we evaluate nine different bacteria recovery methods performed directly from blood culture.Aim. We sought to identify a bacterial recovery method that would allow for a cost-effective and efficient recovery of common BSI pathogens directly from blood culture.Methods. Simulated E. coli ATCC 25922 blood culture was used as a model system to evaluate nine different bacteria recovery methods. Each method was assessed on recovery yield, cost, hands-on time, risk of contamination and ease of use. The highest scoring recovery method was further evaluated using simulated blood cultures spiked with seven of the most frequently occurring bloodstream pathogens. The recovery yield was calculated based on c.f.u. count before and after each recovery method. Independent t-tests were performed to determine if the recovery methods evaluated were significantly different based on c.f.u. ml-1 log recovery.Results. All nine methods evaluated successfully recovered E. coli ATCC 25922 from simulated blood cultures although the bacterial yield differed significantly. The MALDI-TOF intact cell method offered the poorest recovery with a mean loss of 2.94±0.37 log c.f.u. ml-1. In contrast, a method developed by Bio-Rad achieved the greatest bacterial yield with a mean bacteria loss of 0.27±0.013 log c.f.u. ml-1. Overall, a low-speed serum-separation method was demonstrated to be the most efficient method in terms of time, cost and recovery efficiency and successfully recovered seven of the most frequent BSI pathogens with a mean bacteria loss of 0.717±0.18 log c.f.u. ml-1.Conclusion. The efficiency of bacterial recovery can vary significantly between different methods and thereby can have a critical impact on downstream analysis. The low-speed serum-separation method offered a simple and effective means of recovering common BSI pathogens from blood culture and will be further investigated for use in the rapid detection of bacteraemia and susceptibility testing in clinical practice.


Assuntos
Bactérias/isolamento & purificação , Técnicas Bacteriológicas/métodos , Hemocultura/métodos , Patógenos Transmitidos pelo Sangue/isolamento & purificação , Bacteriemia/diagnóstico , Escherichia coli/isolamento & purificação , Humanos
8.
Can J Microbiol ; 66(10): 586-592, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32497439

RESUMO

This study aimed to isolate and identify the population of filamentous fungi colonizing a cotton painting, whose conservation status was compromised and showed signs of biodeterioration due to dirt accumulation and microbial metabolism. In addition, microbiological techniques such as cultivation-dependent approach and molecular biology were used to identify microbial populations and to eliminate their metabolic action. For this, the nondestructive anoxic atmosphere technique was used, in which the microbial metabolism was affected by the absence of oxygen. Prior to exposure to an anoxic atmosphere, only one fungal species, Aspergillus niger, was identified at 12 points sampled in the obverse and reverse of the artwork; no fungal species persisted as a result of anoxic treatment. These results showed that exposure to anoxic conditions was effective for the total elimination of isolated fungal strains as well as their spores. In conclusion, this study proved the unprecedented effectiveness of a nondestructive technique for artwork on textile colonized by black fungi species. Thus, this interdisciplinary work involving conservation, microbiology, and chemistry presents a tool to eliminate microorganisms, while maintaining the integrity of artwork and safety of the restorer, that can be applied prior to artwork restoration.


Assuntos
Anaerobiose/fisiologia , Aspergillus niger/isolamento & purificação , Fibra de Algodão/microbiologia , Pinturas , Aspergillus niger/genética , Técnicas Bacteriológicas , DNA Bacteriano/genética , Reação em Cadeia da Polimerase , Células-Tronco
9.
PLoS One ; 15(6): e0234542, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32555702

RESUMO

Staphylococcus aureus is one of the principal causative agents of bacteremia which can progress to sepsis. Rapid diagnostic tests for identification and antibiotic resistance profiling of S. aureus would improve patient outcomes and antibiotic stewardship, but existing methods require a lengthy culture step to obtain enough material for testing. Complexity of the host matrix, where pathogenic microbes are often present, also interferes with many diagnostic methods. Here, we describe a straightforward and rapid method for enriching viable S. aureus using bio-orthogonal, or "click," chemistry methods. Bacteria labeled in this manner can potentially be cultured, interrogated using molecular methods for pathogen identification, or used to test antibiotic susceptibility.


Assuntos
Técnicas Bacteriológicas , Sepse/diagnóstico , Infecções Estafilocócicas/diagnóstico , Staphylococcus aureus/isolamento & purificação , Testes Diagnósticos de Rotina , Farmacorresistência Bacteriana , Humanos , Sepse/microbiologia , Infecções Estafilocócicas/microbiologia , Staphylococcus aureus/patogenicidade , Propriedades de Superfície
10.
BMC Infect Dis ; 20(1): 326, 2020 May 07.
Artigo em Inglês | MEDLINE | ID: mdl-32380973

RESUMO

BACKGROUND: T2Bacteria assay uses T2 magnetic resonance (T2MR) technology for the rapid diagnosis of bacterial bloodstream infections (BSIs). This FDA cleared technology can detect 5 of the most prevalent pathogens causing bacteremia (Escherichia coli, Staphylococcus aureus, Klebsiella pneumoniae, Pseudomonas aeruginosa, and Enterococcus faecium). Because the significance of discordant results between the T2Bacteria assay and blood culture (BC) remains a challenge, in this case series we reviewed the medical records of patients who had a positive T2Bacteria test and a concurrent negative BC. METHODS: Among 233 participants, we identified 20 patients with 21 (9%) discordant T2Bacteria-positive/BC-negative (T2+/BC-) results. We classified these results based on clinical cultures and clinical evidence. RESULTS: When we analyzed these 21 discordant results in-depth, 11 (52.5%) fulfilled criteria for probable BSI, 4 (19%) for possible BSI, and 6 (28.5%) were presumptive false positives. Among the probable/possible BSIs, discordant results were often associated with patients diagnosed with closed space and localized infections [pyelonephritis (n = 7), abscess (n = 4), pneumonia (n = 1), infected hematoma (n = 1), and osteomyelitis (n = 1)]. Also, within the preceding 2 days of the T2+/BC- blood sample, 80% (16/20) of the patients had received at least one dose of an antimicrobial agent which was active against the T2Bacteria-detected pathogen. CONCLUSIONS: In the majority of discrepant results, the T2Bacteria assay detected a plausible pathogen that was supported by clinical and/or microbiologic data. Discrepancies appear to be associated with closed space and localized infections and the recent use of effective antibacterial agents. The clinical significance and potential implications of such discordant results should be further investigated.


Assuntos
Bacteriemia/microbiologia , Técnicas Bacteriológicas/métodos , Adulto , Idoso , Idoso de 80 Anos ou mais , Antibacterianos/uso terapêutico , Bacteriemia/tratamento farmacológico , Hemocultura , Infecções por Escherichia coli/microbiologia , Reações Falso-Positivas , Feminino , Infecções por Bactérias Gram-Positivas/microbiologia , Humanos , Infecções por Klebsiella/microbiologia , Masculino , Pessoa de Meia-Idade , Infecções por Pseudomonas/microbiologia , Infecções Estafilocócicas/tratamento farmacológico , Infecções Estafilocócicas/microbiologia , Adulto Jovem
11.
Epidemiol Infect ; 148: e107, 2020 05 18.
Artigo em Inglês | MEDLINE | ID: mdl-32418555

RESUMO

Porphyromonas gingivalis has been linked to the development and progression of oesophageal squamous cell carcinoma (ESCC), and is considered to be a high-risk factor for ESCC. Currently, the commonly used methods for P. gingivalis detection are culture or DNA extraction-based, which are either time and labour intensive especially for high-throughput applications. We aimed to establish and evaluate a rapid and sensitive direct quantitative polymerase chain reaction (qPCR) protocol for the detection of P. gingivalis without DNA extraction which is suitable for large-scale epidemiological studies. Paired gingival swab samples from 192 subjects undergoing general medical examinations were analysed using two direct and one extraction-based qPCR assays for P. gingivalis. Tris-EDTA buffer-based direct qPCR (TE-direct qPCR), lysis-based direct qPCR (lysis-direct qPCR) and DNA extraction-based qPCR (kit-qPCR) were used, respectively, in 192, 132 and 60 of these samples for quantification of P. gingivalis. The sensitivity and specificity of TE-direct qPCR was 95.24% and 100% compared with lysis-direct qPCR, which was 100% and 97.30% when compared with kit-qPCR; TE-direct qPCR had an almost perfect agreement with lysis-direct qPCR (κ = 0.954) and kit-qPCR (κ = 0.965). Moreover, the assay time used for TE-direct qPCR was 1.5 h. In conclusion, the TE-direct qPCR assay is a simple and efficient method for the quantification of oral P. gingivalis and showed high sensitivity and specificity compared with routine qPCR.


Assuntos
Reação em Cadeia da Polimerase/métodos , Porphyromonas gingivalis/isolamento & purificação , Técnicas Bacteriológicas , Humanos , Sensibilidade e Especificidade
12.
PLoS One ; 15(5): e0232912, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32392236

RESUMO

The study of oral disease progression, in relation to the accumulation of subgingival biofilm in gingivitis and periodontitis is limited, due to either the ability to monitor plaque in vitro. When compared, optical spectroscopic techniques offer advantages over traditional destructive or biofilm staining approaches, making it a suitable alternative for the analysis and continued development of three-dimensional structures. In this work, we have developed a confocal Raman spectroscopy analysis approach towards in vitro subgingival plaque models. The main objective of this study was to develop a method for differentiating multiple oral subgingival bacterial species in planktonic and biofilm conditions, using confocal Raman microscopy. Five common subgingival bacteria (Fusobacterium nucleatum, Streptococcus mutans, Veillonella dispar, Actinomyces naeslundii and Prevotella nigrescens) were used and differentiated using a 2-way orthogonal Partial Least Square with Discriminant Analysis (O2PLS-DA) for the collected spectral data. In addition to planktonic growth, mono-species biofilms cultured using the 'Zürich Model' were also analyzed. The developed method was successfully used to predict planktonic and mono-species biofilm species in a cross validation setup. The results show differences in the presence and absence of chemical bands within the Raman spectra. The O2PLS-DA model was able to successfully predict 100% of all tested planktonic samples and 90% of all mono-species biofilm samples. Using this approach we have shown that Confocal Raman microscopy can analyse and predict the identity of planktonic and mono-species biofilm species, thus enabling its potential as a technique to map oral multi-species biofilm models.


Assuntos
Bactérias/isolamento & purificação , Gengivite/microbiologia , Microscopia Óptica não Linear/métodos , Periodontite/microbiologia , Actinomyces , Técnicas Bacteriológicas/métodos , Biofilmes/crescimento & desenvolvimento , Meios de Cultura , Fusobacterium nucleatum , Gengiva/microbiologia , Viabilidade Microbiana , Microbiota , Microscopia Confocal/métodos , Plâncton , Prevotella intermedia , Streptococcus mutans , Veillonella
13.
J Infect Chemother ; 26(8): 858-861, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32360135

RESUMO

In an 80-bed fee-based retirement home with nursing care, the dispatched-pharmacist has provided prescription recommendations to visiting physicians based on pathogen identification using Gram staining as part of an antimicrobial stewardship program. Thus, we evaluated the effects of pharmacist-supported antimicrobial stewardship. We calculated the total number of all antimicrobials and macrolides, fluoroquinolones, and cephalosporins prescriptions per 100 residents per month at the retirement home from January 2013 to December 2017. Using log-transformed monthly resident numbers with an offset before and after the intervention, we performed Poisson regression analyses that adjusted for monthly mean age. Interrupted time series analyses (ITSA) were conducted to examine the changes in the incidence rate ratios for the baseline and slope before and after the intervention. The total number of all antimicrobial prescriptions per 100 residents per month from 2013 to 2017 was 14.10, 18.51, 10.59, 5.41, and 3.90, respectively. Although there was a significant pre-intervention increase in the total number of all antimicrobial prescriptions, the intervention was followed by a significant decrease. There was also a significant reduction in the slope. ITSA of the changes in the prescription of macrolides and fluoroquinolones showed that there were significant pre-intervention increase and followed by a significant post-intervention decrease in the slope. There was no significant change in cephalosporin prescriptions by the intervention. Our study shows that pharmacist-supported AS can reduce antimicrobial prescriptions in a retirement home. Nevertheless, further studies are needed to collect and analyse more data on similar interventions.


Assuntos
Anti-Infecciosos/uso terapêutico , Gestão de Antimicrobianos/estatística & dados numéricos , Instituição de Longa Permanência para Idosos/estatística & dados numéricos , Casas de Saúde/estatística & dados numéricos , Farmacêuticos , Antibacterianos/uso terapêutico , Técnicas Bacteriológicas , Cefalosporinas/uso terapêutico , Prescrições de Medicamentos/estatística & dados numéricos , Fluoroquinolonas/uso terapêutico , Violeta Genciana , Humanos , Análise de Séries Temporais Interrompida , Japão , Macrolídeos/uso terapêutico , Fenazinas , Aposentadoria , Coloração e Rotulagem
14.
New Microbiol ; 43(1): 13-16, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-32334488

RESUMO

The long incubation time required for Mycobacteria detection may allow cultures to become overgrown by contaminating organisms. Therefore, samples need to be decontaminated before solid and liquid culture. MYCO-TB is a ready-to-use digestion and decontamination kit with single-sample formulation developed by Copan. Sample processing time (3 minutes) is shorter than that of other commercial kits. The aim of this study was to compare the performance of MYCO-TB with MycoPrep, both based on N-acetyl-Lcysteine and sodium hydroxide solution, in terms of culture contamination and Mycobacterial detection by culture. We tested 162 respiratory samples: the overall proportions of contamination of both liquid and solid media were 1.8% for MYCO-TB and 1.8% for MycoPrep. Mycobacterial growth was detected without significant differences in times to positivity (TTP) in liquid culture: 10.5 days for MYCO-TB and 11.1 days for MycoPrep. Samples decontaminated with MYCO-TB were suitable for molecular assays such as Xpert MTB/RIF Ultra and GenoType CMdirect. Extending decontamination times (up to 10 minutes) with MYCO-TB of 20 Mycobacteria-positive specimens did not produce any difference in TTP in liquid culture or in Ultra IS1081/IS6110 probe Ct values. In conclusion, the MYCO-TB kit proved to be effective for the rapid digestion and decontamination of respiratory materials for the detection of Mycobacteria, making it possible to reduce the manual skills required and lower the risk of contamination. Longer decontamination time could be used for samples with a high level of contamination, such as those from cystic fibrosis patients.


Assuntos
Técnicas Bacteriológicas , Mycobacterium tuberculosis , Kit de Reagentes para Diagnóstico , Tuberculose , Técnicas Bacteriológicas/métodos , Técnicas Bacteriológicas/normas , Descontaminação , Humanos , Mycobacterium tuberculosis/isolamento & purificação , Kit de Reagentes para Diagnóstico/normas , Sensibilidade e Especificidade , Tuberculose/diagnóstico , Tuberculose/microbiologia
15.
BMC Infect Dis ; 20(1): 253, 2020 Mar 30.
Artigo em Inglês | MEDLINE | ID: mdl-32228597

RESUMO

BACKGROUND: The aims of this study were to (1) evaluate the efficacy and safety of targeted antibiotics for the treatment of culture-negative prosthetic joint infection based on metagenomic next-generation sequencing results and (2) verify the accuracy and reliability of metagenomic next-generation sequencing for identifying pathogens related to culture-negative prosthetic joint infection. METHODS: Ninety-seven consecutive PJI patients, including 27 patients with culture-negative prosthetic joint infection, were treated surgically at our center. Thirteen of the 27 culture-negative prosthetic joint infection patients, who were admitted before June 2017 and treated with empirical antibiotics, comprised the empirical antibiotic group (EA group), and the other 14 patients, who were admitted after June 2017 and treated with targeted antibiotics according to their metagenomic next-generation sequencing results, were classified as the targeted antibiotic group (TA group). The short-term infection control rate, incidence of antibiotic-related complications and costs were compared between the two groups. RESULTS: Two of the patients in the EA group experienced debridement and prolonged antimicrobial therapy due to wound infection after the initial revision surgery. No recurrent infections were observed in the TA group; however, no significant difference in the infection control rate was found between the two groups (83.33% vs 100%, P = 0.217). More cases of antibiotic-related complications were recorded in the EA group (6 cases) than in the TA group (1 case), but the difference was not statistically significant (P = 0.0697). The cost of antibiotics obtained for the EA group was 20,168.37 Yuan (3236.38-45,297.16), which was higher than that found for the TA group (10,164.16 Yuan, 2959.54-16,661.04, P = 0.04). CONCLUSIONS: Targeted antibiotic treatment for culture-negative prosthetic joint infection based on metagenomic next-generation sequencing results is associated with a favorable outcome, and metagenomic next-generation sequencing is a reliable tool for identifying pathogens related to culture-negative prosthetic joint infection.


Assuntos
Sequenciamento de Nucleotídeos em Larga Escala/métodos , Metagenômica/métodos , Infecções Relacionadas à Prótese/microbiologia , Idoso , Antibacterianos/uso terapêutico , Técnicas Bacteriológicas , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Infecções Relacionadas à Prótese/tratamento farmacológico , Reoperação , Reprodutibilidade dos Testes , Infecção da Ferida Cirúrgica/tratamento farmacológico , Infecção da Ferida Cirúrgica/microbiologia , Resultado do Tratamento
16.
J Vet Sci ; 21(2): e30, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-32233136

RESUMO

Mycoplasma ovipneumoniae (Mo) is difficult to culture, resulting in many difficulties in related research and application. Since nucleotide metabolism is a basic metabolism affects growth, this study conducted a "point-to-point" comparison of the corresponding growth phases between the Mo NM151 strain and the Mycoplasma mycoides subsp. capri (Mmc) PG3 strain. The results showed that the largest difference in nucleotide metabolism was found in the stationary phase. Nucleotide synthesis in PG3 was mostly de novo, while nucleotide synthesis in NM151 was primarily based on salvage synthesis. Compared with PG3, the missing reactions of NM151 referred to the synthesis of deoxythymine monophosphate. We proposed and validated a culture medium with added serine to fill this gap and prolong the stationary phase of NM151. This solved the problem of the fast death of Mo, which is significant for related research and application.


Assuntos
Técnicas Bacteriológicas/veterinária , Meios de Cultura/química , Mycoplasma ovipneumoniae/crescimento & desenvolvimento , Transcriptoma , Técnicas Bacteriológicas/métodos , Mycoplasma ovipneumoniae/metabolismo , Nucleotídeos/metabolismo
17.
Anal Chim Acta ; 1113: 18-25, 2020 May 29.
Artigo em Inglês | MEDLINE | ID: mdl-32340665

RESUMO

Magnetic trapping has been employed in the development of analytical methods owing to its ease and simplicity in handling samples. Nevertheless, the generation of functional probes is usually time consuming. A new and simple affinity method that uses gadolinium ion (Gd3+), a magnetic ion, as affinity probe for magnetic tapping of pathogenic bacteria was demonstrated in the present study. Escherichia coli O157:H7, Staphylococcus aureus, and Acinetobacter baumannii were selected as model bacteria. The model bacteria were magnetically isolated after incubation in Tris buffer (pH 8) containing Gd3+ (0.1 M) under microwave heating (power: 180 W, 90 s × 3). The resultant Gd3+-bacterium conjugates possessed sufficient magnetism, resulting in magnetic aggregations by an external magnet (∼4,000 Gauss). For ease of magnetic isolation, the sample containing Gd3+-bacterium complexes was stirred by a small magnet. After 1 h, the magnet attached with precipitates, i.e., Gd3+-bacterium conjugates, was readily removed using a pair of tweezers. The bacteria in the resultant conjugates were characterized by matrix-assisted laser desorption/ionization mass spectrometry. The limits of detection of the current approach toward E. coli O157:H7, S. aureus, and A. baumannii in complex samples were ∼104-105 cells mL-1.


Assuntos
Acinetobacter baumannii/isolamento & purificação , Técnicas Bacteriológicas/métodos , Escherichia coli O157/isolamento & purificação , Gadolínio/química , Staphylococcus aureus/isolamento & purificação , Acinetobacter baumannii/química , Animais , Sangue/microbiologia , Bovinos , Complexos de Coordenação/química , Difosfatos/química , Escherichia coli O157/química , Limite de Detecção , Fenômenos Magnéticos , Microbiologia do Solo , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Staphylococcus aureus/química
18.
J Infect Chemother ; 26(8): 813-817, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32312620

RESUMO

The time to positivity (TTP) of blood culture has significant value for clinicians. However, almost all subjects of previous studies regarding TTP were adults and early infants. Therefore, careful attention is required when referring to previously published data, which might differ according to the age of subjects, as the tendency of infectious focus and pathogens identified from culture vary with age. In this study, we compared the TTP between two pediatric age groups (≤12 months and 13 months to 15 years [>12 months]) at a teaching hospital during a 5-year period. Of the 95 subjects, 41 and 54 were aged ≤12 and > 12 months, among whom true pathogenic bacteria were identified in 12 (29.3%) and 19 (35.2%), respectively. The median TTP for the younger group with pathogenic bacteria was 11.2 (interquartile range, 10.0-11.9) hours, which was significantly shorter than that for the older group (12.6 [interquartile range, 11.9-16.9] hours) (P = 0.01). At 12 h after the initiation of culture, the younger group with pathogenic bacteria had a significantly higher positivity rate (83.3%) than the older group (26.3%) (P < 0.01). The times required for the positivity to exceed 90% were 13.4 and 20.1 h for the younger and older pathogenic groups and 30.4 and 67.8 h for the younger and older contaminant groups, respectively. The range of TTP could be assessed more accurately by considering the effect of age on the infectious background.


Assuntos
Bacteriemia/epidemiologia , Bacteriemia/microbiologia , Hemocultura/métodos , Adolescente , Fatores Etários , Bacteriemia/diagnóstico , Bactérias/isolamento & purificação , Técnicas Bacteriológicas , Sangue/microbiologia , Criança , Pré-Escolar , Feminino , Humanos , Lactente , Masculino , Estudos Retrospectivos , Fatores de Tempo
19.
J Dairy Sci ; 103(6): 5043-5046, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32307175

RESUMO

Postpasteurization contamination (PPC) of fluid milk remains a challenge for some dairy processors. Pseudomonas is the most common contaminant of fluid milk after pasteurization, and therefore methods to detect PPC should be inclusive of Pseudomonas and other gram-negative contaminants (e.g., coliform bacteria). Our objective was to compare the ability of 3M (St. Paul, MN) coliform and Enterobacteriaceae (EB) Petrifilm to detect total gram-negative bacteria with that of the standard method, crystal violet tetrazolium agar. To that end, we evaluated coliform Petrifilm, EB Petrifilm, and crystal violet tetrazolium agar to detect gram-negative bacteria in naturally contaminated samples of fluid milk. A total of 92 observations derived from shelf-life testing of 33 milk samples from 5 different processing facilities were evaluated for (1) presence of coliforms on coliform Petrifilm at both 24 and 48 h of incubation; (2) presence of any growth, regardless of gas production, on coliform Petrifilm at both 24 and 48 h of incubation; (3) presence of EB on EB Petrifilm at both 24 and 48 h of incubation; (4) presence of any growth, regardless of gas or acid production, on EB Petrifilm at both 24 and 48 h of incubation; and (5) presence of gram-negative bacteria on crystal violet tetrazolium agar after 48 h of incubation. Sensitivity and specificity analysis of results indicated that compared with the standard method (i.e., crystal violet tetrazolium agar), the method that performed the best, based on balanced accuracy (i.e., the average of sensitivity and specificity), was coliform Petrifilm evaluated for the presence of any growth after 48 h of incubation (sensitivity = 0.787; specificity = 0.839). This method can be easily adopted by the dairy industry as many processing facilities already test for coliforms using coliform Petrifilm. Improving the ability of processors to detect PPC will improve the quality of the fluid milk supply.


Assuntos
Técnicas Bacteriológicas , Enterobacteriaceae/metabolismo , Microbiologia de Alimentos , Bactérias Gram-Negativas/isolamento & purificação , Leite/microbiologia , Animais , Contagem de Colônia Microbiana , Indústria de Laticínios , Pasteurização , Pseudomonas
20.
APMIS ; 128(5): 406-413, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32202675

RESUMO

In areas of low tuberculosis (TB) prevalence, laboratory diagnosis of TB may essentially cover non-tuberculous mycobacteria (NTM) in addition to Mycobacterium tuberculosis (MTB). In this study, a semi-automated PCR workflow distinguishing MTB and NTM (Anyplex™ MTB/NTMe, Seegene) and subsequently detecting MTB isoniazid/rifampicin resistance (Allplex™ MTB/MDRe, Seegene) was evaluated for replacing smear microscopy of acid-fast bacilli as the rapid screening method for TB. With 279 clinical samples, 47 cultures positive for MTB and 76 for NTM, the Anyplex™ MTB/NTMe assay and smear microscopy showed equal sensitivities (49.6% vs 50.8%, respectively) but Anyplex™ MTB/NTMe was more sensitive for MTB (63.8% vs 25.6%) than for NTM (40.8% vs 64.5%). Allplex™ MTB/MDRe showed a slightly higher sensitivity of 68.1% for MTB (32/47 positive, n = 222). Antibiotic resistance profiles were correctly identified for all MTB isolates (one MDR isolate). Specificity was 100% for both assays. Anyplex™ MTB/NTMe detected all the 18 NTM species present in the study. The analytical performance of the evaluated high-throughput workflow was relatively weak compared to culture but potentially adequate as a rapid screening method analogous to smear microscopy with additional differentiation between TB, MDR-TB, and NTM.


Assuntos
Automação Laboratorial , Farmacorresistência Bacteriana Múltipla , Mycobacterium tuberculosis/isolamento & purificação , Micobactérias não Tuberculosas/isolamento & purificação , Tuberculose Resistente a Múltiplos Medicamentos/diagnóstico , Técnicas Bacteriológicas , Humanos , Técnicas de Diagnóstico Molecular , Mycobacterium tuberculosis/genética , Micobactérias não Tuberculosas/genética , Valor Preditivo dos Testes , Reação em Cadeia da Polimerase em Tempo Real , Sensibilidade e Especificidade
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