scholarly journals Identification of Bacteria in the Sputum of a Cystic Fibrosis patient; A Comparison of Phenotypic and Molecular Methods

2017 ◽  
Vol 11 (1) ◽  
pp. 384-386 ◽  
Author(s):  
Mubarak Alfaresi ◽  
Bassam Mahboub
Keyword(s):  
Cystic Fibrosis ◽  
Bacterial Species ◽  
Transmembrane Conductance Regulator ◽  
Bacterial Identification ◽  
Molecular Method ◽  
Isolation And Identification ◽  
Pcr Product ◽  
Clinical Culture ◽  
Identification Of Bacteria ◽  
Cf Transmembrane Conductance Regulator

Background: Cystic fibrosis (CF), caused by mutations in the CF transmembrane conductance regulator gene, is a common autosomal recessive disease. Accurate isolation and identification of the bacteria underlying these infections are is critical to the therapeutic management of CF. Objective: To compare phenotypic bacterial identification with a molecular method in a CF patient sputum. Methods: Bacterial identification done by standard microbiological method from a CF patient. Same sample underwent a molecular method involving 16S rDNA amplification, cloning, and sequencing. Results: All isolated bacteria from culture were also found after cloning PCR Product. Conversely, 9 pathogenic bacterial species were only detected after PCR and cloning. Conclusion: This study supports prior suggestions that a sequence-based molecular approach to clinical microbiology can significantly enhance the standard clinical culture-based view.

scholarly journals Isolation and identification of bacteria and parasites in glass eel (Anguilla spp.)

2021 ◽  
Vol 322 ◽  
pp. 02012
Author(s):  
Septyan Andriyanto ◽  
Hessy Novita ◽  
Tuti Sumiati ◽  
Taukhid
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Prevalence Rate ◽  
Bacterial Species ◽  
Bacterial Identification ◽  
Biochemical Method ◽  
Isolation And Identification ◽  
Glass Eel ◽  
Pathogenic Agent ◽  
Identification Of Bacteria ◽  
Glass Eels

The disease is the main agent that causes mortality of fish, especially during seed stages. The research aimed to find out bacteria and parasitic speciesin glass eel, Anguilla spp. Bacterial identification was carried out by a biochemical method. The prevalence of bacterial species was calculated using the El-Gohary et al. (2020) formula, while the results of bacterial identification from glass eel were Aeromonas spp., Vibrio spp., Enterococcus spp., Staphylococcus spp., Planococcus spp., Lactobacillus spp., Listeria spp., Citrbacterfreundii, Neisseria spp., Pseudomonas aeruginosa, Kurthia spp., Streptococcus spp., and Corynebacterium spp. It was found that the five highest prevalence rate was for Listeria spp. (39.64%), followed by Aeromonas spp. (26.13%), Staphylococcus spp. (16.22%), Corynebacterium spp. (5.41%), Lactobacillus spp. (2.70%), and the lowest prevalence rate was Streptococcus spp. (0.90%). The type of parasitic pathogen obtained was Trichodina spp. (2,70%), Dactylogyrus spp. (2,70%) and Gyrodactylus spp. (2,70%). Bacterial and parasites identified in glass eels need further verification on the epizootiology characteristic of each pathogenic agent.

scholarly journals VX-809 corrects folding defects in cystic fibrosis transmembrane conductance regulator protein through action on membrane-spanning domain 1

2013 ◽  
Vol 24 (19) ◽  
pp. 3016-3024 ◽  
Author(s):  
Hong Yu Ren ◽  
Diane E. Grove ◽  
Oxana De La Rosa ◽  
Scott A. Houck ◽  
Pattarawut Sopha ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Chloride Transport ◽  
Genetic Disorder ◽  
Missense Mutations ◽  
Transmembrane Conductance Regulator ◽  
Transmembrane Conductance ◽  
Regulator Protein ◽  
Folding Defect ◽  
Membrane Spanning ◽  
Cf Transmembrane Conductance Regulator

Cystic fibrosis (CF) is a fatal genetic disorder associated with defective hydration of lung airways due to the loss of chloride transport through the CF transmembrane conductance regulator protein (CFTR). CFTR contains two membrane-spanning domains (MSDs), two nucleotide-binding domains (NBDs), and a regulatory domain, and its channel assembly requires multiple interdomain contacts. The most common CF-causing mutation, F508del, occurs in NBD1 and results in misfolding and premature degradation of F508del-CFTR. VX-809 is an investigational CFTR corrector that partially restores CFTR function in people who are homozygous for F508del-CFTR. To identify the folding defect(s) in F508del-CFTR that must be repaired to treat CF, we explored the mechanism of VX-809 action. VX-809 stabilized an N-terminal domain in CFTR that contains only MSD1 and efficaciously restored function to CFTR forms that have missense mutations in MSD1. The action of VX-809 on MSD1 appears to suppress folding defects in F508del-CFTR by enhancing interactions among the NBD1, MSD1, and MSD2 domains. The ability of VX-809 to correct F508del-CFTR is enhanced when combined with mutations that improve F508del-NBD1 interaction with MSD2. These data suggest that the use of VX-809 in combination with an additional CFTR corrector that suppresses folding defects downstream of MSD1 may further enhance CFTR function in people with F508del-CFTR.

scholarly journals Maturation and function of cystic fibrosis transmembrane conductance regulator variants bearing mutations in putative nucleotide-binding domains 1 and 2.

1991 ◽  
Vol 11 (8) ◽  
pp. 3886-3893 ◽  
Author(s):  
R J Gregory ◽  
D P Rich ◽  
S H Cheng ◽  
D W Souza ◽  
S Paul ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Amino Acid ◽  
Functional Activity ◽  
Nucleotide Binding ◽  
Missense Mutations ◽  
Transmembrane Conductance Regulator ◽  
Transmembrane Conductance ◽  
Fluorescence Measurements ◽  
Cf Transmembrane Conductance Regulator ◽  
Altered Proteins

One feature of the mutations thus far found to be associated with the disease cystic fibrosis (CF) is that many of them are clustered within the first nucleotide-binding domain (NBD) of the CF transmembrane conductance regulator (CFTR). We sought to discover the molecular basis for this clustering by introducing into the two NBDs of CFTR mutations either mimicking amino acid changes associated with CF or altering residues within highly conserved motifs. Synthesis and maturation of the mutant CFTR were studied by transient expression in COS cells. The ability of the altered proteins to generate cyclic AMP-stimulated anion efflux was assessed by using 6-methoxy-N-(sulfopropyl) quinolinium (SPQ) fluorescence measurements in HeLa cells expressing mutated plasmids. The results show that (i) all CF-associated mutants, with one exception, lack functional activity as measured in the SPQ assay, (ii) mutations in NBD1 are more sensitive to the effects of the same amino acid change than are the corresponding mutations in NBD2, (iii) cells transfected with plasmids bearing CF-associated mutations commonly but not exclusively lack mature CFTR, (iv) NBD mutants lacking mature CFTR fail to activate Cl- channels, and (v) the glycosylation of CFTR, per se, is not required for CFTR function. We reason that the structure of NBD1 itself or of the surrounding domains renders it particularly sensitive to mutational changes. As a result, most NBD1 mutants, but only a few NBD2 mutants, fail to mature or lack functional activity. These findings are consistent with the observed uneven distribution of CFTR missense mutations between NBD1 and NBD2 of CF patients.

scholarly journals Dysfunctional cystic fibrosis transmembrane conductance regulator inhibits phagocytosis of apoptotic cells with proinflammatory consequences

2009 ◽  
Vol 297 (4) ◽  
pp. L677-L686 ◽  
Author(s):  
R. William Vandivier ◽  
Tiffany R. Richens ◽  
Sarah A. Horstmann ◽  
Aimee M. deCathelineau ◽  
Moumita Ghosh ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Rho Kinase ◽  
Negative Regulator ◽  
Apoptotic Cells ◽  
Transmembrane Conductance Regulator ◽  
Transmembrane Conductance ◽  
Altered Expression ◽  
Proinflammatory Mediators ◽  
Downstream Effector ◽  
Cf Transmembrane Conductance Regulator

Cystic fibrosis (CF) is caused by mutated CF transmembrane conductance regulator (CFTR) and is characterized by robust airway inflammation and accumulation of apoptotic cells. Phagocytosis of apoptotic cells (efferocytosis) is a pivotal regulator of inflammation, because it prevents postapoptotic necrosis and actively suppresses release of a variety of proinflammatory mediators, including IL-8. Because CF is associated with accumulation of apoptotic cells, inappropriate levels of IL-8, and robust inflammation, we sought to determine whether CFTR deficiency specifically impairs efferocytosis and its regulation of inflammatory mediator release. Here we show that CFTR deficiency directly interferes with efferocytosis by airway epithelium, an effect that is not due to altered binding of apoptotic cells to epithelial cells or altered expression of efferocytosis receptors. In contrast, expression of RhoA, a known negative regulator of efferocytosis, is substantially increased in CFTR-deficient cells, and inhibitors of RhoA or its downstream effector Rho kinase normalize efferocytosis in these cells. Impaired efferocytosis appears to be mediated through an amiloride-sensitive ion channel, because amiloride restores phagocytic competency in CFTR-deficient cells. Finally, ineffective efferocytosis in CFTR-deficient cells appears to have proinflammatory consequences, because apoptotic cells enhance IL-8 release by these cells, but not by wild-type controls. Therefore, in CF, dysregulated efferocytosis may lead to accumulation of apoptotic cells and impaired regulation of the inflammatory response and, ultimately, may suggest a new therapeutic target.

scholarly journals Elevation of hepatic sulphotransferase activities in mice with resistance to cystic fibrosis

2002 ◽  
Vol 364 (1) ◽  
pp. 115-120 ◽  
Author(s):  
Josie L. FALANY ◽  
Heather GREER ◽  
Timea KOVACS ◽  
Eric J. SORSCHER ◽  
Charles N. FALANY
Keyword(s):  
Cystic Fibrosis ◽  
Mouse Liver ◽  
Northern Blot Analysis ◽  
Transmembrane Conductance Regulator ◽  
Male Mice ◽  
Transmembrane Conductance ◽  
Female Adult ◽  
Cf Transmembrane Conductance Regulator ◽  
Different Levels

The severity of intestinal disease in the cystic fibrosis (CF) transmembrane conductance regulator (CFTR) (-/−) mice has been reported to co-segregate with gene loci which contain the genes for hydroxysteroid sulphotransferase (SULT). Because of the potential involvement of steroid hormones in CF, we investigated levels of steroid SULT activity in the livers of CFTR mice to determine whether the levels of SULT activity correlate with the occurrence or severity of CF. To elucidate the possible role of SULT activity in ameliorating the deleterious effects of CF in CFTR (-/−) mice, we determined the levels of phenol SULT (PST), hydroxysteroid SULT [dehydroepiandrosterone (DHEA)-ST] and oestrogen SULT (EST) activity in control CFTR (+/+), heterozygous CFTR (+/−) and homozygous CFTR (-/−) mice, which survive to adulthood. The level of PST activity was not significantly different between any of the groups of mice, regardless of sex or genotype. Although DHEA-ST activity was significantly higher in female mice than in male mice, there was no difference in DHEA-ST activity that could be correlated with genotype. In contrast with PST and DHEA-ST activities, we found that some male and all female adult CFTR (-/−) mice had elevated, dramatically different levels of EST from both CFTR (+/+) and CFTR (+/−) mice. Results from these SULT activity experiments were confirmed by Northern-blot analysis of mouse-liver RNA. Subsequent studies with preweanling mice revealed no differences in the levels of EST that could be correlated with genotype. Thus this study indicates that EST is elevated significantly in CFTR (-/−) mice which survive to adulthood and provides important biochemical information that EST levels may be protective in CF.

scholarly journals Hyperglycemia impedes lung bacterial clearance in a murine model of cystic fibrosis-related diabetes

2014 ◽  
Vol 306 (1) ◽  
pp. L43-L49 ◽  
Author(s):  
William R. Hunt ◽  
Susu M. Zughaier ◽  
Dana E. Guentert ◽  
Melissa A. Shenep ◽  
Michael Koval ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Murine Model ◽  
Lavage Fluid ◽  
Transmembrane Conductance Regulator ◽  
Bacterial Clearance ◽  
Cell Counts ◽  
Elevated Glucose ◽  
Clinically Significant ◽  
Cf Transmembrane Conductance Regulator ◽  
And Control

Cystic fibrosis-related diabetes (CFRD) is the most common comorbidity associated with cystic fibrosis (CF), impacting more than half of patients over age 30. CFRD is clinically significant, portending accelerated decline in lung function, more frequent pulmonary exacerbations, and increased mortality. Despite the profound morbidity associated with CFRD, little is known about the underlying CFRD-related pulmonary pathology. Our aim was to develop a murine model of CFRD to explore the hypothesis that elevated glucose in CFRD is associated with reduced lung bacterial clearance. A diabetic phenotype was induced in gut-corrected CF transmembrane conductance regulator (CFTR) knockout mice (CFKO) and their CFTR-expressing wild-type littermates (WT) utilizing streptozotocin. Mice were subsequently challenged with an intratracheal inoculation of Pseudomonas aeruginosa (PAO1) (75 μl of 1–5 × 106 cfu/ml) for 18 h. Bronchoalveolar lavage fluid was collected for glucose concentration and cell counts. A portion of the lung was homogenized and cultured as a measure of the remaining viable PAO1 inoculum. Diabetic mice had increased airway glucose compared with nondiabetic mice. The ability to clear bacteria from the lung was significantly reduced in diabetic WT mice and control CFKO mice. Critically, bacterial clearance by diabetic CFKO mice was significantly more diminished compared with nondiabetic CFKO mice, despite an even more robust recruitment of neutrophils to the airways. This finding that CFRD mice boast an exaggerated, but less effective, inflammatory cell response to intratracheal PAO1 challenge presents a novel and useful murine model to help identify therapeutic strategies that promote bacterial clearance in CFRD.

scholarly journals BACTERIA IN ANIMAL FEED AND THEIR SUSCEPTIBILITY TO ANTIMICROBIAL DRUGS

2012 ◽  
Vol 5 (2) ◽  
pp. 27-33
Author(s):  
Igor M. Stojanov ◽  
Milica M. Živkov Baloš ◽  
Miloš C. Kapetanov ◽  
Jasna Z. Prodanov Radulović ◽  
Jelena M. Petrović
Keyword(s):  
Animal Feed ◽  
Bacterial Species ◽  
Antibiotic Sensitivity ◽  
Diffusion Method ◽  
Isolation And Identification ◽  
Antimicrobial Drugs ◽  
Microbiological Safety ◽  
Feed Control ◽  
Identification Of Bacteria ◽  
Feed Safety

Control of microbiological safety of animal feed is regulated by a law. The regulation is based on methodological standards for estimation of feed quality. Those methods are permanently amended and strive for continuous improvement of microbiological safety of feed. The question remains whether the presence or absence of some bacterial species is sufficient for proper assessment of feed safety. The answer to this question can be found not only in the examination of the number of bacteria in animal feed, but also in qualitative analysis of the isolated microorganisms. The sensitivity of bacteria from animal feed to antimicrobials is one of the qualitative characteristics of microorganisms that can affect food safety even if the feed meets the standards and is within the maximum permitted level. The subject of our work was to determine the presence of bacteria in animal feed with a purpose to examine if they possess lower sensibility to antibiotics. For testing antibiotic sensitivity the isolates from routine microbiological feed control were used. The animal feed was analysed by the standard methods for the isolation and identification of bacteria and determination of susceptibility was done using disc diffusion method according to CLSI. The following bacteria were detected: Escherichia coli, Enterobacter sp., Staphylococcus sp., Micrococcus sp., Bacillus cereus, Bacillus sp. The results showed that some isolates have reduced susceptibility to certain antimicrobial drugs (The resistance is different for different antibiotics and microorganism - from 8,33% to 100%). This finding gives a reason for further examination of certain characteristics of the bacteria found in animal feed and to detect their possible influence on ecology.

In vivo analysis of fluid transport in cystic fibrosis airway epithelia of bronchial xenografts

1996 ◽  
Vol 270 (5) ◽  
pp. C1326-C1335 ◽  
Author(s):  
Y. Zhang ◽  
J. Yankaskas ◽  
J. Wilson ◽  
J. F. Engelhardt
Keyword(s):  
Cystic Fibrosis ◽  
Fluid Transport ◽  
Xenograft Model ◽  
Na Channel ◽  
Transmembrane Conductance Regulator ◽  
Fluid Absorption ◽  
Cystic Fibrosis Airway ◽  
In Vivo Analysis ◽  
Cf Transmembrane Conductance Regulator

An in vivo human bronchial xenograft model system was used to simultaneously analyze electrolyte and fluid transport defects in fully differentiated human cystic fibrosis (CF) and non-CF proximal airways. CF airways demonstrated three discernible defects when compared with non-CF, including 1) a lack of adenosine 3',5'-cylic monophosphate (cAMP)-inducible Cl- secretion, 2) a fourfold higher basal fluid absorption rate, and 3) an altered regulation of fluid absorption in response to amiloride-stimulated changes in Na+ transport. A unique finding in this study demonstrated that treatment of epithelia with amiloride led to a greater than threefold decrease in the rate of fluid absorption in CF tissues as contrasted to a greater than threefold increase in the rate of fluid absorption in non-CF tissues. The removal of apical Na+ from amiloride-treated non-CF xenografts was capable of ablating this amiloride-induced increase in fluid absorption. In light of the recent interactions demonstrated between CF transmembrane conductance regulator (CFTR) and the rat epithelial, amiloride-sensitive Na+ channel, these findings implicate additional complexities between the Na+ conductance pathways and fluid transport in normal and CF proximal airways. Such findings suggest that CFTR may also regulate amiloride-insensitive Na+ channels.

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