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1.
Arch Microbiol ; 206(6): 246, 2024 May 05.
Artigo em Inglês | MEDLINE | ID: mdl-38704767

RESUMO

Shake-flask culture, an aerobic submerged culture, has been used in various applications involving cell cultivation. However, it is not designed for forced aeration. Hence, this study aimed to develop a small-scale submerged shaking culture system enabling forced aeration into the medium. A forced aeration control system for multiple vessels allows shaking, suppresses volatilization, and is attachable externally to existing shaking tables. Using a specially developed plug, medium volatilization was reduced to less than 10%, even after 45 h of continuous aeration (~ 60 mL/min of dry air) in a 50 mL working volume. Escherichia coli IFO3301 cultivation with aeration was completed within a shorter period than that without aeration, with a 35% reduction in the time-to-reach maximum bacterial concentration (26.5 g-dry cell/L) and a 1.25-fold increase in maximum concentration. The maximum bacterial concentration achieved with aeration was identical to that obtained using the Erlenmeyer flask, with a 65% reduction in the time required to reach it.


Assuntos
Meios de Cultura , Escherichia coli , Escherichia coli/crescimento & desenvolvimento , Volatilização , Meios de Cultura/química , Reatores Biológicos/microbiologia , Técnicas Bacteriológicas/métodos
2.
Int J Mycobacteriol ; 13(1): 34-39, 2024 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-38771277

RESUMO

BACKGROUND: In patients with cystic fibrosis (CF), representatives of the fast-growing Mycobacterium abscessus complex (MABSc) are often distinguished, but the culture of the material taken from such patients increases the growth time. We analyzed the terms of cultivation of MABSc representatives on dense nutrient media and also evaluated the productivity of a modified nutrient medium based on agar for the isolation of Burkholderia cepacia complex (BCC). METHODS: Sixty-four strains of MABSc isolated from patients with CF and suspected tuberculosis were analyzed. The material from the patients was cultured on a universal chromogenic medium, 5% blood agar, yolk-salt agar, selective medium for isolation of BCC, and Löwenstein-Jensen medium. The cultures were incubated for 5 days (37°C, aerobic conditions), after for 23 days (28°C, aerobic conditions). The productivity of the developed nutrient medium was evaluated by the number of cells that gave visible growth after culturing 0.1 mL of a bacterial suspension of 103 CFU/mL. RESULTS: 76.8% of the strains grew in a 2-week period, and 23.2% of the strains were obtained at a later date from 18 to 28 days (average: 21.23 days). The modified medium with a concentration of 240 mg of iron (III) polymaltose hydroxide proved to be the most optimal for the isolation of MABSc. CONCLUSION: When using a chromogenic medium for culture material from patients with CF, it is necessary to extend incubation up to 28 days to increase the probability of MABSc isolation. The modified BCC medium showed a good selectivity result but required further investigation.


Assuntos
Meios de Cultura , Fibrose Cística , Infecções por Mycobacterium não Tuberculosas , Mycobacterium abscessus , Humanos , Fibrose Cística/microbiologia , Meios de Cultura/química , Mycobacterium abscessus/crescimento & desenvolvimento , Mycobacterium abscessus/isolamento & purificação , Infecções por Mycobacterium não Tuberculosas/microbiologia , Fatores de Tempo , Técnicas Bacteriológicas/métodos , Complexo Burkholderia cepacia/isolamento & purificação , Complexo Burkholderia cepacia/crescimento & desenvolvimento
3.
J Clin Microbiol ; 62(5): e0144523, 2024 May 08.
Artigo em Inglês | MEDLINE | ID: mdl-38557148

RESUMO

The virulence of methicillin-resistant Staphylococcus aureus (MRSA) and its potentially fatal outcome necessitate rapid and accurate detection of patients colonized with MRSA in healthcare settings. Using the BD Kiestra Total Lab Automation (TLA) System in conjunction with the MRSA Application (MRSA App), an imaging application that uses artificial intelligence to interpret colorimetric information (mauve-colored colonies) indicative of MRSA pathogen presence on CHROMagar chromogenic media, anterior nares specimens from three sites were evaluated for the presence of mauve-colored colonies. Results obtained with the MRSA App were compared to manual reading of agar plate images by proficient laboratory technologists. Of 1,593 specimens evaluated, 1,545 (96.98%) were concordant between MRSA App and laboratory technologist reading for the detection of MRSA growth [sensitivity 98.15% (95% CI, 96.03, 99.32) and specificity 96.69% (95% CI, 95.55, 97.60)]. This multi-site study is the first evaluation of the MRSA App in conjunction with the BD Kiestra TLA System. Using the MRSA App, our results showed 98.15% sensitivity and 96.69% specificity for the detection of MRSA from anterior nares specimens. The MRSA App, used in conjunction with laboratory automation, provides an opportunity to improve laboratory efficiency by reducing laboratory technologists' labor associated with the review and interpretation of cultures.


Assuntos
Automação Laboratorial , Técnicas Bacteriológicas , Staphylococcus aureus Resistente à Meticilina , Sensibilidade e Especificidade , Infecções Estafilocócicas , Staphylococcus aureus Resistente à Meticilina/isolamento & purificação , Humanos , Infecções Estafilocócicas/diagnóstico , Infecções Estafilocócicas/microbiologia , Automação Laboratorial/métodos , Técnicas Bacteriológicas/métodos , Automação/métodos , Colorimetria/métodos , Inteligência Artificial
4.
J Clin Microbiol ; 62(5): e0165123, 2024 May 08.
Artigo em Inglês | MEDLINE | ID: mdl-38572970

RESUMO

In clinical bacteriology laboratories, reading and processing of sterile plates remain a significant part of the routine workload (30%-40% of the plates). Here, an algorithm was developed for bacterial growth detection starting with any type of specimens and using the most common media in bacteriology. The growth prediction performance of the algorithm for automatic processing of sterile plates was evaluated not only at 18-24 h and 48 h but also at earlier timepoints toward the development of an early growth monitoring system. A total of 3,844 plates inoculated with representative clinical specimens were used. The plates were imaged 15 times, and two different microbiologists read the images randomly and independently, creating 99,944 human ground truths. The algorithm was able, at 48 h, to discriminate growth from no growth with a sensitivity of 99.80% (five false-negative [FN] plates out of 3,844) and a specificity of 91.97%. At 24 h, sensitivity and specificity reached 99.08% and 93.37%, respectively. Interestingly, during human truth reading, growth was reported as early as 4 h, while at 6 h, half of the positive plates were already showing some growth. In this context, automated early growth monitoring in case of normally sterile samples is envisioned to provide added value to the microbiologists, enabling them to prioritize reading and to communicate early detection of bacterial growth to the clinicians.


Assuntos
Inteligência Artificial , Bactérias , Sensibilidade e Especificidade , Humanos , Bactérias/crescimento & desenvolvimento , Bactérias/isolamento & purificação , Bactérias/classificação , Algoritmos , Técnicas Bacteriológicas/métodos , Processamento de Imagem Assistida por Computador/métodos , Infecções Bacterianas/diagnóstico , Infecções Bacterianas/microbiologia , Bacteriologia , Automação Laboratorial/métodos , Meios de Cultura/química
5.
PLoS One ; 19(3): e0300042, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38536821

RESUMO

BACKGROUND: Mycobacterium tuberculosis culturing remains the gold standard for laboratory diagnosis of tuberculosis. Tuberculosis remains a great public health problem in developing countries like The Gambia, as most of the methods currently used for bacterial isolation are either time-consuming or costly. OBJECTIVE: To evaluate the Kudoh swab method in a West African setting in Gambia, with a particular focus on the method's performance when culturing Mycobacterium africanum West Africa 2 (MAF2) isolates. METHOD: 75 sputum samples were collected in the Greater Banjul Area and decontaminated in parallel with both the standard N-acetyl-L-Cysteine-NaOH (NALC-NaOH) and the Kudoh swab method in the TB diagnostics laboratory in the Medical Research Council Unit The Gambia between 30th December 2017 and 25th February 2018. These samples were subsequently cultured on standard Löwenstein-Jensen and Modified Ogawa media respectively and incubated at 37°C for mycobacterial growth. Spoligotyping was done to determine if the decontamination and culture methods compared could equally detect Mycobacterium tuberculosis, Mycobacterium africanum West Africa 1 and Mycobacterium africanum West Africa 2. RESULT: Among the 50 smear positives, 35 (70%) were culture-positive with Kudoh and 32 (64%) were culture positive with NALC-NaOH, whilst 7(28%) of the 25 smear negative samples were culture positive with both methods (Table 2). There was no significant difference in recovery between both methods (McNemar's test, p-value = 0.7003), suggesting that the overall positivity rate between the two methods is comparable. There were no differences in time-to-positivity or contamination rate between the methods. However, Kudoh yielded positive cultures that were negative on LJ and vice versa. All findings were irrespective of mycobacterial lineages. CONCLUSION: The Kudoh method has comparable sensitivity to the NALC-NaOH method for detecting Mycobacterium tuberculosis complex isolates. It is easy to perform and could be an add on option for mycobacterial culture in the field in The Gambia, since it requires less biosafety equipment.


Assuntos
Mycobacterium tuberculosis , Tuberculose , Humanos , Gâmbia , Hidróxido de Sódio , Técnicas Bacteriológicas/métodos , Escarro/microbiologia , Tuberculose/diagnóstico , Tuberculose/microbiologia , Meios de Cultura
6.
Diagn Microbiol Infect Dis ; 109(2): 116275, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38537505

RESUMO

BACKGROUND: Tuberculosis (TB), caused by Mycobacterium tuberculosis (MTB), remains a global health threat, necessitating faster and more accessible diagnostic methods. This study investigates critical parameters in the application of a commercial ATP bioluminescence assay for the detection of MTB. METHOD: Our objective was to optimize the ATP bioluminescence protocol using BacTiter-Glo™ for MTB, investigating the impact of varying volumes of MTB suspension and reagent on assay sensitivity, evaluating ATP extraction methods, establishing calibration curves, and elucidating strain-specific responses to antimicrobial agents. RESULTS: ATP extraction methods showed no significant improvement over controls. Calibration curves revealed a linear correlation between relative light units (RLU) and colony-forming units (CFU/mL), establishing low detection limits. Antimicrobial testing demonstrated strain-specific responses aligning with susceptibility and resistance patterns. CONCLUSION: Our findings contribute to refining ATP bioluminescence protocols for enhanced MTB detection and susceptibility testing. Further refinements and validation efforts are warranted, holding promise for more efficient diagnostic platforms in the future.


Assuntos
Trifosfato de Adenosina , Medições Luminescentes , Mycobacterium tuberculosis , Tuberculose , Mycobacterium tuberculosis/efeitos dos fármacos , Trifosfato de Adenosina/análise , Trifosfato de Adenosina/metabolismo , Medições Luminescentes/métodos , Humanos , Tuberculose/diagnóstico , Tuberculose/microbiologia , Sensibilidade e Especificidade , Testes de Sensibilidade Microbiana/métodos , Técnicas Bacteriológicas/métodos
7.
Res Vet Sci ; 172: 105237, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38555775

RESUMO

This study includes the evaluation of multiplex real-time PCR (rPCR) kit, which was developed to provide rapid diagnosis of mastitis infections, by working with milk samples of 2 different sources of mastitis and comparing the results with the classical bacteriological culture method (BC). A total of 273 bacteria were isolated in 226 samples (47.88%) out of 472 samples by BC. These were 139 (50.91%) Staphylococcus spp., 61 (22.34%) Streptococcus spp., 15 (5.49%) E. coli, 8 (2.93%) Enterococcus spp., 50 (18.31%) other bacteria. When we look at the multiplex rPCR results; 1052 positive were obtained for the gene regions of 14 different bacteria, 1 yeast, and 1 ß-lactamase gene examined in 472 samples. While no searched gene region was found by rPCR in 78 (16.5%) of the 472 samples studied, at least 1 gene was detected in 394 (83.5%) samples. These 1052 positive samples by rPCR were; 263 (28.43%) Staphylococcus spp., 51 (5.51%) S. aureus, 57 (6.16%) Enterococcus spp., 49 (5.29%) C. bovis, 16 (1.73%) S. dysgalactiae, 84 (9.08%) S. agalactiae, 71 (7.67%) S. uberis, 73 (7.89%) E. coli, 14 (1.51%) Prototheca spp., 39 (4.21%) T. pyogenes/P. indolicus, 5 (0.54%) S. marcescens, 15 (1.62%) K. oxytoca/pneumonia, 117 (12.64%) Mycoplasma spp., 31 (3.35%) M. bovis, 40 (4.32%) yeast, and 127 samples (26.90%) were ß-lactamase positive. When the antibiotic resistance of the isolates was evaluated, 78 (31.96%) tetracycline, 72 (29.5%) penicillin, and 60 (24.59%) clindamycin resistance were observed predominantly in Gram-positive isolates, while 6 (23.07%) tigecycline, 6 (23.07%) netilmicin, 6 (23.07%) pipercillin resistance was found in gram-negative isolates. While a bacteria and/or yeast gene was found by rPCR in 187 of 246 (76.01%) samples with no bacterial growth, a bacterium was isolated with BC in only 20 (8.84%) samples whose gene region was not found by rPCR. As a result, the multiplex rPCR system used in the diagnosis of mastitis has been found to be quite reliable as it can detect a large number of bacteria in a very short time compared to classical methods. Therefore, we advise the use of rPCR and/or culture for confirmation of clinical signs in mastitis and at routine mastitis surveillance.


Assuntos
Mastite Bovina , Reação em Cadeia da Polimerase Multiplex , Reação em Cadeia da Polimerase em Tempo Real , Mastite Bovina/microbiologia , Mastite Bovina/diagnóstico , Feminino , Animais , Reação em Cadeia da Polimerase em Tempo Real/veterinária , Reação em Cadeia da Polimerase em Tempo Real/métodos , Bovinos , Reação em Cadeia da Polimerase Multiplex/veterinária , Reação em Cadeia da Polimerase Multiplex/métodos , Técnicas Bacteriológicas/métodos , Técnicas Bacteriológicas/veterinária , Leite/microbiologia , Bactérias/isolamento & purificação , Bactérias/genética , Bactérias/classificação
8.
Lett Appl Microbiol ; 77(3)2024 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-38364315

RESUMO

The objective of this study is to validate the US Food and Drug Administration (FDA) rea-time polymerase chain reaction (qPCR) assay, the Neogen Amplified Nucleic Single Temperature Reaction (ANSR) assay, and the Vitek ImmunoDiagnostic Assay System (VIDAS) SLM procedure against the FDA cultural procedure for Salmonella detection in green chile pepper. Green chile was artificially contaminated with Salmonella according to the FDA guidelines (FDA. Guidelines for the Validation of Microbiological Methods for the FDA Foods Program, 3rd Edition. 2019. www.fda.gov/media/83812/download?attachment (17 March 2024, date last accessed)) at a fractional recovery level (where 50%-25% tests positive and at a level +1 log greater for each organism tested). Enriched samples were tested directly by the ANSR Salmonella test and by qPCR, and were subcultured into Rappaport-Vassiliadis and tetrathionate brilliant green broth for cultural detection and qPCR. For the VIDAS-SLM assay, the selective enrichments were further cultured in M broth before testing. Presumptive salmonellae were confirmed with biochemical tests, serology, and qPCR. All three rapid assays were compared favorably with the FDA-BAM (Bacteriological Analytical Manual) method. No significant differences at P < .05 were found between the procedures using McNemar's χ2 test. The three procedures were found to be rapid and reliable alternatives to cultural detection of Salmonella enterica in green chile.


Assuntos
Microbiologia de Alimentos , Salmonella enterica , Meios de Cultura , Salmonella enterica/genética , Chile , Técnicas Bacteriológicas/métodos , Salmonella
10.
Clin Lab ; 70(2)2024 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-38345975

RESUMO

BACKGROUND: Bloodstream infections (BSI) represent a common cause of sepsis and mortality in children. Blood culture (BC) is the gold standard for diagnosis of BSI. The low sensitivity of BC in the pediatric population is usually due to the small volume of blood used for inoculation and to the antibiotics used before sampling. Here, we explore the ways to effectively reduce antibiotic activity to maximize the chances of pathogen recovery, and to enhance the growth of microorganisms in lower blood volume and bacterial counts. METHODS: The recovery of common pathogens causing blood stream infections was analyzed after exposure to cefo-perazone/sulbactam, vancomycin, and caspofung by using resin-containing or not BacT/Alert PF Plus and BD FX400 peds plus pediatric bottles. The microbial growth in the resin-containing bottles was assessed using 0.5 colony-forming units (CFU) bacterial inoculum to mimic the bacteremia/fungemia condition. The usefulness of a diagnosis to confirm or exclude BSI was evaluated by lower than recommended blood culture sampling (102 CFU/mL, 0.3 mL). RESULTS: Staphylococcus aureus (S. aureus), and Candida glabrata (C. glabrata) were recovered from 100% of two types of resin-containing bottles in the presence of a sufficient antibiotic dose, while Escherichia coli (E. coli) was not restored to 100% in BD FX400 peds plus pediatric bottles. The shorter TTD for S. aureus, C. glabrata, and E. coli were observed in antibiotic-containing BacT/Alert PF Plus bottles. Both the PF Plus and BD resin test bottles showed consistently good TTD performances to Gram-negative, Gram-positive, and yeast species in low inoculum levels, with the exception of S. aureus. The lower volume of blood inoculated into culture bottles hardly affected the growth of most bacteria, but optimized PF Plus resin-bottles accelerated the detection of infectious agents, especially S. aureus, Streptococcus pneumoniae, and C. glabrata. CONCLUSIONS: It is possible to enhance recovery from antibiotic-containing pediatric bottles and shorten TTD for the identification of pathogens by using the BacT/Alert blood culture system combination with new resin-containing media.


Assuntos
Bacteriemia , Sepse , Infecções Estafilocócicas , Criança , Humanos , Antibacterianos/farmacologia , Escherichia coli , Meios de Cultura , Staphylococcus aureus , Bacteriemia/diagnóstico , Bacteriemia/microbiologia , Bactérias , Técnicas Bacteriológicas
11.
J Clin Microbiol ; 62(1): e0109623, 2024 01 17.
Artigo em Inglês | MEDLINE | ID: mdl-38054730

RESUMO

Rapid diagnostic tests (RDTs) for bloodstream infections have the potential to reduce time to appropriate antimicrobial therapy and improve patient outcomes. Previously, an in-house, lipid-based, matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) method, Fast Lipid Analysis Technique (FLAT MS), has shown promise as a rapid pathogen identification method. In this study, FLAT MS for direct from blood culture identification was evaluated and compared to FDA-cleared identification methods using the Benefit-risk Evaluation Framework (BED-FRAME) analysis. FLAT MS was evaluated and compared to Bruker Sepsityper and bioMérieux BioFire FilmArray BCID2 using results from a previous study. For this study, 301 positive blood cultures were collected from the University of Maryland Medical Center. The RDTs were compared by their sensitivities, time-to-results, hands-on time, and BED-FRAME analysis. The overall sensitivity of all platforms compared to culture results from monomicrobial-positive blood cultures was 88.3%. However, the three RDTs differed in their accuracy for identifying Gram-positive bacteria, Gram-negative bacteria, and yeast. Time-to-results for FLAT MS, Sepsityper, and BioFire BCID2 were all approximately one hour. Hands-on times for FLAT MS, Sepsityper, and BioFire BCID2 were 10 (±1.3), 40 (±2.8), and 5 (±0.25) minutes, respectively. BED-FRAME demonstrated that each RDT had utility at different pathogen prevalence and relative importance. BED-FRAME is a useful tool that can used to determine which RDT is best for a healthcare center.


Assuntos
Bacteriemia , Sepse , Humanos , Bacteriemia/diagnóstico , Bacteriemia/microbiologia , Testes de Diagnóstico Rápido , Técnicas Bacteriológicas/métodos , Sepse/diagnóstico , Hemocultura , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos , Lipídeos
12.
J Vet Diagn Invest ; 36(1): 124-127, 2024 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-37919965

RESUMO

Carbapenemase-producing Enterobacterales (CPE) are one of the most urgent threats to human healthcare globally. Descriptions of CPE outbreaks in veterinary hospitals suggest the need for screening strategies for CPE from companion animals. Our aim was to optimize a chromogenic agar method with and without selective enrichment to isolate CPE from companion animal feces in an ongoing outbreak of New Delhi metallo-ß-lactamse-5 Escherichia coli. A limit of detection (LOD) assay for spiked canine and feline feces was performed for both methods using a carbapenamase-producing E. coli (24213-18); the LOD (1.5 × 103 cfu/g of feces) was equivalent to that reported for human fecal specimens. We screened 1,247 companion animal fecal specimens for carriage of CPE by 1) direct plating to chromogenic agar and 2) plating to chromogenic agar following selective enrichment. Twenty-one specimens were positive for CPE by both direct culture and enrichment culture. No specimens were positive with selective enrichment and negative by direct culture. A selective enrichment step did not result in any increased recovery of CPE from companion animals, which suggests that enrichment broth may not be necessary for outbreak surveillance testing. It is important to continue to validate methods for the detection of CPE in companion animals as outbreaks become more common in veterinary facilities.


Assuntos
Doenças do Gato , Doenças do Cão , Infecções por Enterobacteriaceae , Animais , Gatos , Cães , Humanos , Escherichia coli , Enterobacteriaceae , Ágar , Doenças do Gato/diagnóstico , Doenças do Gato/epidemiologia , Técnicas Bacteriológicas/veterinária , Técnicas Bacteriológicas/métodos , Doenças do Cão/diagnóstico , Doenças do Cão/epidemiologia , Proteínas de Bactérias , Surtos de Doenças/veterinária , Infecções por Enterobacteriaceae/diagnóstico , Infecções por Enterobacteriaceae/epidemiologia , Infecções por Enterobacteriaceae/veterinária , Testes de Sensibilidade Microbiana/veterinária
13.
J Clin Microbiol ; 62(1): e0115223, 2024 01 17.
Artigo em Inglês | MEDLINE | ID: mdl-38126761

RESUMO

The objective of this study was to assess the clinical performances of PhenoMATRIX and PhenoMATRIX PLUS for the screening of methicillin-resistant Staphylococcus aureus (MRSA) from nasal and inguinal/perineal ESwabs using chromogenic media. The automated performances were compared to the manual reading. Additionally, we evaluated PhenoMATRIX PLUS for the automatic release of the negative results to the Laboratory Information System (LIS) and the automatic discharge of the negative plates from the incubators. A total of 6,771 non-duplicate specimens were used by PhenoMATRIX as a machine learning model. The validation of these settings was performed on 17,223 non-duplicate specimens. The MRSA positivity rate was 5% (866/17,223). Validated settings were then used by PhenoMATRIX PLUS on another 1,409 non-duplicate specimens. The sensitivities of PhenoMATRIX and PhenoMATRIX PLUS were 99.8% [95% confidence interval (CI), 99.2%-99.9%] and 100% (95% CI, 92.1%-100%), respectively. The specificities of PhenoMATRIX and PhenoMATRIX PLUS were 99.1% (95% CI, 99.0%-99.2%) and 95.2% (95% CI, 93.8%-96.1%), respectively. All the 1,297 MRSA-negative specimens analyzed by PhenoMATRIX PLUS were automatically released and sent to the LIS immediately after availability of the culture image on the WASPLab (100% accuracy). All negative media plates were automatically discarded. PhenoMATRIX PLUS decreases the time spent by technologists on negative plates and ensures optimal usage of the incubators' capacity.


Assuntos
Staphylococcus aureus Resistente à Meticilina , Infecções Estafilocócicas , Humanos , Técnicas Bacteriológicas/métodos , Sensibilidade e Especificidade , Compostos Cromogênicos , Nariz , Infecções Estafilocócicas/diagnóstico , Meios de Cultura
14.
Transfusion ; 64(1): 104-115, 2024 01.
Artigo em Inglês | MEDLINE | ID: mdl-38098310

RESUMO

BACKGROUND AND OBJECTIVES: We evaluated the operational and safety impact of implementing anaerobic culture screening of apheresis and pooled platelets at the American Red Cross on the already established use of the aerobic culture screening of each donation performed no sooner than 24 h following collection. MATERIALS AND METHODS: Platelets were screened for bacterial contamination with the BACT/ALERT 3D® (bioMérieux, Durham, NC) microbial detection testing system. The addition of anaerobic culture to the already existing aerobic culture resulted in sampling an additional 8-10 mL from each donation. RESULTS: Implementation of anaerobic testing resulted in an approximate 3.5-fold increased rate of False Positive BACT/ALERT alarms. There was a modest increase in the rate of True Positive alarms of 1.4-fold with increased detection of Klebsiella and Propionibacterium species, including Cutibacterium acnes. In addition, there was an approximate 3.5-fold increase rate of False Positives and a 13.5-fold increase rate of Indeterminates, the majority (~57%) were due to Cutibacterium acnes. The combined costs and lost revenue associated with adding anaerobic screening increased by ~$1,000,000/year due to testing cost and product discards. CONCLUSION: The addition of anaerobic culture to aerobic culture to the original donation (without the introduction of sampling delay) resulted in a significant increase in the rate of alerts. The 40% increased rate of True Positive alarms may have modestly improved platelet safety. However, there was a disproportionate increase in the rate of False Positive and Indeterminate bacterial culture alarms, which added substantial cost and overall loss of platelet products.


Assuntos
Remoção de Componentes Sanguíneos , Plaquetas , Humanos , Anaerobiose , Plaquetas/microbiologia , Bactérias , Contaminação de Medicamentos , Técnicas Bacteriológicas
16.
Food Res Int ; 174(Pt 2): 113665, 2023 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-37981373

RESUMO

As outbreaks of foodborne illness caused by the opportunistic pathogen Cronobacter sakazakii (Cs) continue to occur, particularly in infants consuming powdered infant formula (PIF), the need for sensitive, rapid, and easy-to-use detection of Cs from food and food processing environments is increasing. Here, we developed bioluminescent reporter bacteriophages for viable Cs-specific, substrate-free, rapid detection by introducing luciferase and its corresponding substrate-providing enzyme complex into the virulent phage ΦC01. Although the reporter phage ΦC01_lux, constructed by replacing non-essential genes for phage infectivity with a luxCDABE reporter operon, produced bioluminescence upon Cs infection, the emitted signal was quickly decayed due to the superior bacteriolytic activity of ΦC01. By truncating the membrane pore-forming protein holin and thus limiting its function, the bacterial lysis was delayed and the resultant engineered reporter phage ΦC01_lux_Δhol could produce a more stable and reliable bioluminescent signal. Accordingly, ΦC01_lux_Δhol was able to detect at least an average of 2 CFU/ml of Cs artificially contaminated PIF and Sunsik and food contact surface models within a total of 7 h of assays, including 5 h of pre-enrichment for Cs amplification. The sensitive, easy-to-use, and specific detection of live Cs with the developed reporter phage could be applied as a novel complementary tool for monitoring Cs in food and food-related environments for food safety and public health.


Assuntos
Técnicas Bacteriológicas , Bacteriófagos , Cronobacter sakazakii , Microbiologia de Alimentos , Medições Luminescentes , Proteínas Virais , Cronobacter sakazakii/genética , Cronobacter sakazakii/isolamento & purificação , Técnicas Bacteriológicas/métodos , Bacteriófagos/genética , Bacteriófagos/metabolismo , Proteínas Virais/genética , Proteínas Virais/metabolismo , Microbiologia de Alimentos/métodos , Genoma Viral/genética , Deleção de Sequência , Medições Luminescentes/métodos , Sensibilidade e Especificidade
17.
Rev Esp Quimioter ; 36(6): 621-624, 2023 Dec.
Artigo em Espanhol | MEDLINE | ID: mdl-37786957

RESUMO

OBJECTIVE: The prevalence of carbapenemase-producing Enterobacterales has increased in recent years and is considered an important public health problem. METHODS: A total of 106 clinical samples were analyzed by different carbapenemase detection techniques: inhibition discs (ID), immunochromatographic test (ICT) and a genotypic method, comparing them with a multiplex RT-PCR as a reference method. RESULTS: Overall, all 3 techniques exceeded 90% sensitivity, although with differences in the performance of some of them by carbapenemase type. DI had low specificity (62%) for OXA-48, while with TIC the sensitivity for NDM-type metallo-beta-lactamase (93%) was slightly lower than for OXA-48 (95%). The best results were obtained with the genotypic technique (100% overall performance). CONCLUSIONS: Despite the lower sensitivity of TICs (especially in NDM carbapenemases) compared to molecular techniques, with the modification of the protocol we managed to increase this sensitivity and, together with the lower price, simplicity and speed, it makes this technique a good screening option.


Assuntos
Técnicas Bacteriológicas , beta-Lactamases , Humanos , Técnicas Bacteriológicas/métodos , beta-Lactamases/genética , beta-Lactamases/análise , Proteínas de Bactérias/genética , Proteínas de Bactérias/análise , Enterobacteriaceae/genética , Sensibilidade e Especificidade , Testes de Sensibilidade Microbiana
18.
Diagn Microbiol Infect Dis ; 107(4): 116085, 2023 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-37801888

RESUMO

BACKGROUND: We evaluated pre- and postimplementation of Virtuo on outcome in patients with gram-negative bacteremia using a quasiexperimental time-in-motion design. METHODS: Becton Dickinson BACTEC™ 9000 series (Bactec) (2018) and Virtuo system (2020) were utilized in a decentralized and centralized process, respectively. Data collected in August-December in 2018 and 2020 were analyzed with SPSS (ver 28). RESULTS: For 185 patients in each time period, patient age, gender, length of hospitalization were not different. However, blood culture (BC) volume was significantly lower in 2020 (7.1 ± 2.6 mL) compared to 2018 (8.9 ± 1.9 mL). Time from BC draw and time from pathogen identification (ID) to treatment change were both significantly faster in 2020 (52.9 ± 38.3 hours; 15.1 ± 27.4 hours), compared to 2018 (65.0 ± 46.3 hours; 23.8 ± 33.8), respectively. CONCLUSIONS: Replacement of decentralized Bactec with centralized Virtuo, resulted in significant improvement in management of patients gram-negative bacteremia.


Assuntos
Bacteriemia , Hemocultura , Humanos , Hemocultura/métodos , Fatores de Tempo , Bacteriemia/diagnóstico , Bacteriemia/tratamento farmacológico , Meios de Cultura , Técnicas Bacteriológicas/métodos
19.
J Biol Chem ; 299(11): 105286, 2023 11.
Artigo em Inglês | MEDLINE | ID: mdl-37742925

RESUMO

The twin arginine translocation (Tat) pathway transports folded protein across the cytoplasmic membrane in bacteria, archaea, and across the thylakoid membrane in plants as well as the inner membrane in some mitochondria. In plant chloroplasts, the Tat pathway utilizes the protonmotive force (PMF) to drive protein translocation. However, in bacteria, it has been shown that Tat transport depends only on the transmembrane electrical potential (Δψ) component of PMF in vitro. To investigate the comprehensive PMF requirement in Escherichia coli, we have developed the first real-time assay to monitor Tat transport utilizing the NanoLuc Binary Technology in E. coli spheroplasts. This luminescence assay allows for continuous monitoring of Tat transport with high-resolution, making it possible to observe subtle changes in transport in response to different treatments. By applying the NanoLuc assay, we report that, under acidic conditions (pH = 6.3), ΔpH, in addition to Δψ, contributes energetically to Tat transport in vivo in E. coli spheroplasts. These results provide novel insight into the mechanism of energy utilization by the Tat pathway.


Assuntos
Proteínas de Escherichia coli , Escherichia coli , Sistema de Translocação de Argininas Geminadas , Escherichia coli/metabolismo , Proteínas de Escherichia coli/metabolismo , Sinais Direcionadores de Proteínas/fisiologia , Transporte Proteico/fisiologia , Força Próton-Motriz , Medições Luminescentes , Técnicas Bacteriológicas/instrumentação , Técnicas Bacteriológicas/métodos , Metabolismo Energético , Esferoplastos/efeitos dos fármacos , Esferoplastos/metabolismo , Ionóforos/farmacologia
20.
Diagn Microbiol Infect Dis ; 107(3): 116043, 2023 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-37657230

RESUMO

The Resist Acineto from Coris Bioconcept is a novel immunochromatographic test for detection of the major acquired carbapenemases (OXA-23, OXA-40, OXA-58, and NDM) identified in Acinetobacter spp. This rapid and easy-to-perform test showed an excellent specificity and sensitivity, with positive and negatives predictive values of 100% in both cases.


Assuntos
Acinetobacter , Enterobacteriaceae , Humanos , Técnicas Bacteriológicas/métodos , Sensibilidade e Especificidade , beta-Lactamases/análise , Proteínas de Bactérias/análise , Testes Imunológicos , Testes de Sensibilidade Microbiana
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