scholarly journals A Fluorescent Gram Stain for Flow Cytometry and Epifluorescence Microscopy

1998 ◽  
Vol 64 (7) ◽  
pp. 2681-2685 ◽  
Author(s):  
David J. Mason ◽  
Subo Shanmuganathan ◽  
Fiona C. Mortimer ◽  
Vanya A. Gant
Keyword(s):  
Flow Cytometry ◽  
Epifluorescence Microscopy ◽  
Nucleic Acid Binding ◽  
Gram Stain ◽  
Green Fluorescence ◽  
Gram Positive ◽  
Gram Negative ◽  
Syto 13 ◽  
Gram Negative Organisms ◽  
Green Fluorescent

ABSTRACT The fluorescent nucleic acid binding dyes hexidium iodide (HI) and SYTO 13 were used in combination as a Gram stain for unfixed organisms in suspension. HI penetrated gram-positive but not gram-negative organisms, whereas SYTO 13 penetrated both. When the dyes were used together, gram-negative organisms were rendered green fluorescent by SYTO 13; conversely, gram-positive organisms were rendered red-orange fluorescent by HI, which simultaneously quenched SYTO 13 green fluorescence. The technique correctly predicted the Gram status of 45 strains of clinically relevant organisms, including several known to be gram variable. In addition, representative strains of gram-positive anaerobic organisms, normally decolorized during the traditional Gram stain procedure, were classified correctly by this method.

scholarly journals Assessing Gram-stain error rates within the pharmaceutical microbiology laboratory

2020 ◽  
Author(s):  
Tim Sandle
Keyword(s):  
Error Rates ◽  
Microbiology Laboratory ◽  
Gram Stain ◽  
Clinical Context ◽  
Determinative Bacteriology ◽  
Gram Positive ◽  
Gram Negative ◽  
Microbial Identification ◽  
Gram Staining ◽  
Gram Negative Organisms

Gram-staining remains the fundamental method for determinative bacteriology, dividing bacteria into Gram-positive and Gram-negative organisms. This test provides information as to the origin of any contamination and is a pre-requisite for many microbial identification methods. Despite the longevity of the test, the test is highly reliant upon analyst technique and therefore errors occur. While there are a few studies looking at errors in the clinical context, research has not been extended to the pharmaceutical microbiology laboratory context. In this study, we present a review of over 6,000 Gram-stains and establish an error rate of around 3%, with the most common reason for error being an over-decolourisation step resulting in organisms that should be Gram-positive appearing as Gram-negative. The analysis enables others to benchmark their facilities against.

scholarly journals Linearized esculentin-2EM shows pH dependent antibacterial activity with an alkaline optimum

2021 ◽  
Author(s):  
Erum Malik ◽  
David A. Phoenix ◽  
Timothy J. Snape ◽  
Frederick Harris ◽  
Jaipaul Singh ◽  
...  
Keyword(s):  
Helical Structure ◽  
Food Preservation ◽  
Forming Process ◽  
Gram Positive ◽  
Gram Negative ◽  
Bacterial Membranes ◽  
Gram Positive Bacteria ◽  
Alkaline Conditions ◽  
Ph Dependent ◽  
Gram Negative Organisms

AbstractHere the hypothesis that linearized esculentin 2EM (E2EM-lin) from Glandirana emeljanovi possesses pH dependent activity is investigated. The peptide showed weak activity against Gram-negative bacteria (MLCs ≥ 75.0 μM) but potent efficacy towards Gram-positive bacteria (MLCs ≤ 6.25 μM). E2EM-lin adopted an α-helical structure in the presence of bacterial membranes that increased as pH was increased from 6 to 8 (↑ 15.5–26.9%), whilst similar increases in pH enhanced the ability of the peptide to penetrate (↑ 2.3–5.1 mN m−1) and lyse (↑ 15.1–32.5%) these membranes. Theoretical analysis predicted that this membranolytic mechanism involved a tilted segment, that increased along the α-helical long axis of E2EM-lin (1–23) in the N → C direction, with −  < µH > increasing overall from circa − 0.8 to − 0.3. In combination, these data showed that E2EM-lin killed bacteria via novel mechanisms that were enhanced by alkaline conditions and involved the formation of tilted and membranolytic, α-helical structure. The preference of E2EM-lin for Gram-positive bacteria over Gram-negative organisms was primarily driven by the superior ability of phosphatidylglycerol to induce α-helical structure in the peptide as compared to phosphatidylethanolamine. These data were used to generate a novel pore-forming model for the membranolytic activity of E2EM-lin, which would appear to be the first, major reported instance of pH dependent AMPs with alkaline optima using tilted structure to drive a pore-forming process. It is proposed that E2EM-lin has the potential for development to serve purposes ranging from therapeutic usage, such as chronic wound disinfection, to food preservation by killing food spoilage organisms.

scholarly journals 1554. Epidemiology of Adult Bacterial Hand Infections at Two Urban Hospitals

2020 ◽  
Vol 7 (Supplement_1) ◽  
pp. S777-S778
Author(s):  
Arsheena Yassin ◽  
Christine Stavropoulos ◽  
Krystina L Woods ◽  
Jiashan Xu ◽  
Justin Carale ◽  
...  
Keyword(s):  
New York ◽  
Staphylococcus Aureus ◽  
Antibiotic Regimen ◽  
Gram Positive ◽  
Gram Negative ◽  
Streptococcus Species ◽  
Urban Hospitals ◽  
Haemophilus Parainfluenzae ◽  
Empiric Antibiotic ◽  
Gram Negative Organisms

Abstract Background Hand infections represent a major source of morbidity, which can result in hand stiffness and amputation. Early appropriate empiric antibiotic regimen may reduce the associated morbidity, hence the importance to examine local epidemiology. The aim of this study was to define the current epidemiology of adult hand infections at two urban hospitals in New York City. Methods We performed a double center, retrospective study of adult patients hospitalized from March 2018 to May 2020. Patients with positive cultures associated with the hand infections were included. Retrospectively, 100 patients were reviewed. Data on baseline demographic, clinical, surgical, microbiology, and treatment parameters were collected. Results Of the 100 patients, 76% were male, with median age of 47.5 years (35, 58.25) and average C-reactive protein (CRP) of 50.66 mg/L (± 64.64) on admission (see Table 1). Previous hospitalization within 1 year (38%), previous surgical procedures (39%) and recent IV medication use (26%) were common. 130 bacterial isolates were identified (see Table 2). The most frequent organisms were Gram-positive, with Methicillin susceptible Staphylococcus aureus (MSSA, 25.38%), Streptococcus species (20.08%), and Methicillin resistant Staphylococcus aureus (MRSA, 15.38%) being the most common. Gram-negative organisms were infrequent, with Haemophilus parainfluenzae (3.85%), Enterobacter cloacae (3.85) and Pseudomonas aeruginosa (3.08%) being the most prevalent. Of the 100 patients, 27% had polymicrobial infections, associated with trauma (6%), illicit IV use (6%) and unknown (7%) etiologies. Table 1: Baseline demographics and co-morbid conditions Table 2: Types and numbers of organisms in relation to etiologies Conclusion Within our population, the most common organisms associated with hand infections were Gram-positive, with Staphylococcus aureus and Streptococcus species being the most prevalent. Gram-negative pathogens were infrequently isolated. The results within this study can provide guidance to clinicians on assessing the appropriate empiric antibiotic regimen in patients with hand infections, and can serve as a basis for further studies identifying risk factors associated with isolation of organisms associated with hand infections. Disclosures All Authors: No reported disclosures

scholarly journals Evaluation of MicroScan Bacterial Identification Panels for Low-Resource Settings

Diagnostics ◽  
2021 ◽  
Vol 11 (2) ◽  
pp. 349
Author(s):  
Sien Ombelet ◽  
Alessandra Natale ◽  
Jean-Baptiste Ronat ◽  
Olivier Vandenberg ◽  
Liselotte Hardy ◽  
...  
Keyword(s):  
Bacterial Species ◽  
Ease Of Use ◽  
Bacterial Identification ◽  
Gram Positive ◽  
Gram Negative ◽  
Low Resource Settings ◽  
Low Resource ◽  
Bacillus Spp ◽  
Gram Negative Organisms ◽  
Instructions For Use

Bacterial identification is challenging in low-resource settings (LRS). We evaluated the MicroScan identification panels (Beckman Coulter, Brea, CA, USA) as part of Médecins Sans Frontières’ Mini-lab Project. The MicroScan Dried Overnight Positive ID Type 3 (PID3) panels for Gram-positive organisms and Dried Overnight Negative ID Type 2 (NID2) panels for Gram-negative organisms were assessed with 367 clinical isolates from LRS. Robustness was studied by inoculating Gram-negative species on the Gram-positive panel and vice versa. The ease of use of the panels and readability of the instructions for use (IFU) were evaluated. Of species represented in the MicroScan database, 94.6% (185/195) of Gram-negative and 85.9% (110/128) of Gram-positive isolates were correctly identified up to species level. Of species not represented in the database (e.g., Streptococcus suis and Bacillus spp.), 53.1% out of 49 isolates were incorrectly identified as non-related bacterial species. Testing of Gram-positive isolates on Gram-negative panels and vice versa (n = 144) resulted in incorrect identifications for 38.2% of tested isolates. The readability level of the IFU was considered too high for LRS. Inoculation of the panels was favorably evaluated, whereas the visual reading of the panels was considered error-prone. In conclusion, the accuracy of the MicroScan identification panels was excellent for Gram-negative species and good for Gram-positive species. Improvements in stability, robustness, and ease of use have been identified to assure adaptation to LRS constraints.

THE EFFECT OF SALMINE ON BACTERIA

1958 ◽  
Vol 4 (2) ◽  
pp. 65-71 ◽  
Author(s):  
Thomas D. Brock
Keyword(s):  
Sodium Chloride ◽  
Gram Negative Bacteria ◽  
Bacteriostatic Activity ◽  
Bactericidal Action ◽  
Gram Stain ◽  
Gram Positive ◽  
Gram Negative ◽  
Bacterial Surface ◽  
Bactericidal Effects

The bacteriostatic and bactericidal effects of salmine on various bacteria have been studied. Salmine has more bacteriostatic activity against Gram-positive than against Gram-negative bacteria. It is bactericidal in water but not in broth, and this bactericidal action occurs against both Gram-positive and Gram-negative bacteria. It has been shown that salmine causes agglutination of washed suspensions of certain bacteria and this agglutination is not correlated directly with the Gram stain. Salmine causes an increase in the turbidity of washed cells of all bacteria, Gram-positive and Gram-negative, and differs in this respect from the solutes sodium chloride and glucose, which affect only Gram-negative species.A comparison has been made of the effects of salmine and polymyxin and it has been concluded that salmine may also act by attachment to the bacterial surface.

scholarly journals Comparing the Performance of the New Fully Automated Urine Particle Analyzer UF-5000 with UF-1000i and Gram Stain in Predicting Bacteria Growth Patterns in Women with Uncomplicated Urinary Tract Infections

2020 ◽  
Author(s):  
Stephen Shei-Dei Yang ◽  
Chun-Chun yang ◽  
Yi-Shen Chen ◽  
Shangjen chang
Keyword(s):  
Urinary Tract ◽  
Urine Analysis ◽  
Growth Patterns ◽  
Gram Stain ◽  
Gram Positive ◽  
Gram Negative ◽  
Bacterial Patterns ◽  
Tract Infections ◽  
Sensitivity Specificity ◽  
Gram Positive Cocci

Abstract BackgroudTo compare the performance of the new flow cytometer UF-5000 with UF-1000i (Sysmex, Kobe, Japan) and Gram stain in predicting the bacterial patterns in urine samples MethodsWomen with symptoms suggestive of urinary tract infection were enrolled. Mid-stream urine sample was collected for gram staining, urine analysis and urine culture. Bacterial patterns were classified though UF1000i (none, cocci bacteria or rods/mixed growth), UF-5000 (none, cocci, rods or mixed growth) and Gram stain. Results Among the 102 samples, there were 10 gram-positive cocci, 2 gram-positive bacilli, 66 gram-negative rods, and 24 mixed growth. The sensitivity/specificity of the UF-1000i was 81.8/91.1% for gram-negative rods and 23.5/96.9% for cocci/mixed. The sensitivity/specificity of the UF-5000 was 80.0/88.2% for gram negative rods and 70.0/86.5% for gram-positive cocci.ConclusionsThe UF-5000 demonstrated the good sensitivity and specificity for Gram-negative bacilli bacteria and demonstrated an improved sensitivity for detecting Gram-positive cocci.

A 10-YEAR REVIEW OF COMPARISON BETWEEN GRAM-POSITIVE AND GRAM-NEGATIVE ORGANISMS IN ACUTE THORACIC EMPYEMA

CHEST Journal ◽  
2009 ◽  
Vol 136 (4) ◽  
pp. 45S
Author(s):  
Abhijit Duggal ◽  
Konstantinos Marmagkiolis ◽  
Lawrence Goldstein
Keyword(s):  
Gram Positive ◽  
Gram Negative ◽  
Thoracic Empyema ◽  
Gram Negative Organisms

P1173PERITONEAL DIALYSIS INFECTIONS IN MALTA - TRENDS OVER ELEVEN YEARS

2020 ◽  
Vol 35 (Supplement_3) ◽  
Author(s):  
Angela Borg Cauchi ◽  
Maria Angela Gauci ◽  
Theresia Dalli ◽  
James Gauci ◽  
James Farrugia ◽  
...  
Keyword(s):  
Peritoneal Dialysis ◽  
Coagulase Negative Staphylococcus ◽  
Distributed Data ◽  
Dialysis Modality ◽  
Gram Positive ◽  
Gram Negative ◽  
Male Predominance ◽  
E Coli ◽  
Number Of Patients ◽  
Gram Negative Organisms

Abstract Background and Aims Infections related to peritoneal dialysis (PD) are still a cause of morbidity and mortality. We describe an overview of PD peritonitis and catheter-related infections (CRI) in Malta over a period of eleven years. We also describe trends in dialysis modality over the years. Method All patients undergoing PD in Malta during 2008 and 2018 were analysed. Data from 2008-2012 was retrospective, shown as mean, that from 2013-2018 prospective. International Society for Peritoneal Dialysis (ISPD) definitions were used. Results for categorical responses were summarized using absolute numbers and percentages. Medians (range) were used to describe continuous non-normally distributed data. Results The total number of patients undergoing PD from 2008 till 2018 were 137 (2008-2012), 91, 80, 126, 117, 102, 103 respectively. There was an overall male predominance of 63.5% (61-67). Patient years at risk were 85.80, 85.25, 89.71, 83.70, 79.69, 72.88 since 2013 respectively. The overall incidence of diabetes mellitus was 45.3% (41.8-50), cardiovascular disease 34.2% (33.8-35), hypertension 79.3% (73.8-84.6). PD was used in 50% of dialysis modality prior to 2012, 39% in 2018. Initially 51% used Automated PD (APD), with 21% assisted PD, in 2018 39% used APD, with 6% assisted PD. PD peritonitis rates from 2008 were 0.38, 0.31, 0.35, 0.46, 0.43, 0.57, 0.54, 0.43, 0.39, 0.40, 0.46 episodes/patient year respectively There was marked dominance of Gram-positive peritonitis, mainly Staphylococcal, with a reduction of coagulase-negative-Staphylococcus from 0.26 episodes/patient in 2013 to 0.03 in 2017, 0.11 in 2018. Methicillin-resistant S. aureus (MRSA) peritonitis decreased from 0.03 episodes/patient to nil in 2016, 2017, 0.01 episodes/patient in 2018. Amongst Gram-negative peritonitis, Pseudomonas rates decreased from 0.06 to 0.03 episodes/patient in 2018, nil in 2016. Escherichia coli rates decreased from 0.02 episodes/patient to nil in the last three years. Fungal rates from 0.03 to 0.01 episodes/patient/year, with nil in 2016, 2017. Catheter-related infection rates were 0.39 (2008-2012), 0.35, 0.91, 0.37, 0.38, 0.25, 0.50 episodes/patient/year respectively. There was a higher incidence of recurrent infections in 2014, none in 2015 and 2016. Gram-negative organisms accounted for 57% of all CRI, predominantly Pseudomonas at 0.12 (2008-2012), 0.06, 0.09, 0.09, 0.14, 0.03, 017 episodes/patient/year respectively. Gram-positive CRI were mostly Staphylococcus aureus, peaking in 2014 at 0.38 episodes/patient/year. MRSA rates declined from 0.15 to 0.01 episodes/patient/year in 2018. Conclusion PD peritonitis rates in Malta between 2008 and 2018 were below the ISPD recommended threshold. There were no episodes of MRSA in 2016, 2017, no Pseudomonas in 2016, no E coli in the last three years and no fungal PD peritonitis in 2016, 2017. CRI rates also declined, with an overall predominance of Gram-negative infections.

Fusobacterium nucleatum, the first Gram-negative bacterium demonstrated to produce polyglutamate

2009 ◽  
Vol 55 (5) ◽  
pp. 627-632 ◽  
Author(s):  
Thomas Candela ◽  
Marie Moya ◽  
Michel Haustant ◽  
Agnès Fouet
Keyword(s):  
In Silico Analysis ◽  
Fusobacterium Nucleatum ◽  
Gram Negative Bacteria ◽  
Negative Bacterium ◽  
Gram Positive ◽  
Gram Negative ◽  
Gram Positive Bacteria ◽  
The Third ◽  
Gram Negative Organisms ◽  
First Time

Poly-γ-glutamate has been described in many Gram-positive organisms. When anchored to the surface, it is a capsule and as such a virulence factor. Based on sequence similarities, few Gram-negative organisms have been suggested to synthesize poly-γ-glutamate. For the first time, a Gram-negative bacterium, Fusobacterium nucleatum , is shown to produce and secrete poly-γ-glutamate. Putative poly-γ-glutamate-synthesizing genes from Gram-negative organisms have been compared with their Gram-positive homologs by in silico analysis, i.e., gene sequence and phylogenetic analysis. Clusters of three instead of four genes were highlighted by our screen. The products of the first two genes display similarity with their Gram-positive equivalents, yet the sequences from the Gram-negative organisms can be distinguished from those of the Gram-positives. Interestingly, the sequence of the predicted product of the third gene is conserved among Gram-negative bacteria but displays no similarity to that of either the third or fourth gene of the Gram-positive operons. It is suggested that, like for Gram-positive bacteria, poly-γ-glutamate has a role in virulence for pathogens and one in survival for other Gram-negative bacteria.

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