scholarly journals Comparison of Phenotypic and Genotypic Identification Methods of Pasteurella multocida Serotypes Isolated from Pigs

2018 ◽  
Vol 44 (1) ◽  
pp. 5
Author(s):  
Amanda Figueiredo Amaral ◽  
Raquel Rebelatto ◽  
Catia Silene Klein ◽  
Karine Ludwig Takeuti ◽  
David Emilio Santos Neves de Barcellos
Keyword(s):  
Staphylococcus Aureus ◽  
Multiplex Pcr ◽  
Pasteurella Multocida ◽  
Weighted Kappa ◽  
Type A ◽  
Kappa Coefficient ◽  
Identification Methods ◽  
Type D ◽  
Weighted Kappa Coefficient ◽  
Capsular Typing

Background: Pasteurella multocida serotypes A and D are commonly associated with pneumonia and pleuritis in pigs.  Different phenotypic techniques, such as hyaluronidase and acriflavine tests, and genotyping techniques, such as PCR, are used to distinguish between these serotypes. The objective of this study was to compare the capsular identification methods of type A and type D P. multocida isolated from pigs using both phenotypic (hyaluronidase and acriflavine tests) and genotypic (multiplex PCR) techniques.Materials, Methods & Results: A total of 44 lyophilized P. multocida isolates, obtained between 1981 and 1997 from pig farms at Rio Grande do Sul State, Brazil, were analyzed. The isolates were reactivated in Brain Heart Infusion (BHI) broth and cultured in BHI broth and blood agar supplemented with 5% sheep blood. Colony identity was further confirmed by evaluating colony morphology in blood agar and confirming the absence of growth on MacConkey agar. Bacteria in Tryptone Soy Agar (TSA) were used for the Triple Sugar Iron (TSI), Sulfide-Indole-Motility (SIM), and nitrate, glucose, lactose, sucrose and mannitol fermentation tests. For hyaluronidase test, P. multocida colonies were streaked transversally across the entire plate, approximately 3-5mm apart, in order to observe their lines of growth. Following this, a hyaluronidase producing strain of Staphylococcus aureus was heavily streaked at right angles to the P. multocida lines and the plates were incubated at 37°C for 24 h. Type A isolates were then identified as those with smaller colonies in the region adjacent to the Staphylococcus aureus streak (negative satellitism). For acriflavine test, the isolates were inoculated into tubes containing 2 mL of BHI, incubated at 37°C for 18-24 h, centrifuged (500 g for 15 min) and 1.5 mL of the supernatant was discarded. A 1:1000 solution of acriflavine neutral (0.5 mL) was then added to the residual broth containing bacteria and kept at room temperature. Solutions of acriflavine were freshly prepared each week and stored protected from light. Type D strains were identified by the appearance of a heavy flocculent precipitate within 5 min. DNA extraction by heat shock was performed prior to multiplex PCR for the detection of capsular genes hyaD-hyaC (capsular typing A) and dcbF (capsular typing D). Test of symmetry and a weighted kappa coefficient were used to evaluate correlations and to assess agreement of the  results between the identification methods, respectively. Phenotypic tests showed that two isolates were type D (4.55%), 40 were type A (90.9%) and two (4.55%) were untypable isolates (4.55%) while PCR showed that 38 isolates were type A (86.36%) and six were type D (13.64%). The correlation analysis between the phenotypic and genotypic tests showed that 90.9% of the strains were identified as belonging to the same serotype by both tests and the weighted kappa coefficient (K = 0.633) indicates a substantial agreement between the two tests.Discussion: There was a disagreement between the phenotypic and genotypic results in four of the isolates (9.09%). The phenotypically untypable isolates were classified as type D by multiplex PCR. Nonetheless, we conclude that PCR testing is a more reliable method to differentiate between P. multocida serotypes A and D.

Phenotypic and genotypic characterisation of Pasteurella multocida strains isolated from pigs in Hungary

2007 ◽  
Vol 55 (4) ◽  
pp. 425-434 ◽  
Author(s):  
Zsuzsanna Varga ◽  
Boglárka Sellyei ◽  
T. Magyar
Keyword(s):  
Multiplex Pcr ◽  
Ornithine Decarboxylase ◽  
Pasteurella Multocida ◽  
Regional Distribution ◽  
Type A ◽  
Capsule Type ◽  
Type D ◽  
Biochemical Characterisation ◽  
River Danube ◽  
Swine Herds

A total of 146 Pasteurella multocida strains isolated from swine in Hungary in the last 20 years were examined. Biochemical characterisation and PCR-based techniques were used to determine species, subspecies, biovar, capsule type and presence of the toxA gene. Eighty-seven percent of the isolates belonged to P. multocida ssp. multocida , and 98% of these had biovar 3 or were trehalose-or lactose-fermenting or ornithine decarboxylase negative variants of that. Ten percent of the strains were P. multocida ssp. septica , and within this group 80% of the strains showed sorbitol-negative biovars (5, 6 and 7). The rest of the strains (20%) were lactose positive. Only 3% of the porcine isolates were P. multocida ssp. gallicida and 3 out of the 4 strains belonged to the dulcitol-fermenting biovar 8. Using a capsule-specific multiplex PCR, 60% of the strains belonged to capsule type D, 38% to capsule type A, and only 1 isolate had capsule type F. In contrast with data published in the literature, only 3% of capsule type D isolates carried the toxA gene, while this ratio was 41% for the type A strains. A remarkable regional distribution of toxA gene positive strains was observed. All but two isolates were found in swine herds located in the Transdanubian region, separated from other parts of Hungary by the river Danube.

scholarly journals Phenotypic and Genotypic Characterisation of Pasteurella Multocida Strains Isolated from Pigs in Poland

2013 ◽  
Vol 57 (1) ◽  
pp. 29-34 ◽  
Author(s):  
Katarzyna Stępniwska ◽  
Iwona Markowska-Daniel
Keyword(s):  
Pasteurella Multocida ◽  
Type A ◽  
High Susceptibility ◽  
Biochemical Tests ◽  
Internal Organs ◽  
Type D ◽  
Phenotypic Characterisation ◽  
Nasal Swabs ◽  
Typing Methods ◽  
Capsular Typing

AbstractA total of 319Pasteurella multocida(Pm) strains isolated from pigs in Poland were examined. Phenotypic characterisation included: biochemical tests (to determine species, subspecies, and biovar), capsular typing, and antimicrobial susceptibility. Genotypic characterisation included detection of thetoxAgene by PCR. All testedPmstrains were classified asPmsubsp.multocida: 87.2% biovar 3, 10.7%-2 and 0.9%-12. One strain was classified as biovar 1. Three strains ofPmdid not suit any of the biovars. Using capsular typing methods, 77% ofPmstrains isolated from nasal swabs belonged to type D and 33% to type A. AmongPmstrains isolated from internal organs, 59.5% belonged to type A and 40.5% to type D. All the isolates showed a high susceptibility to β-lactams: ampicillin and amoxicilin with clavulonic acid (97.8%), penicillin (86.7%), doxicilline (100%), oxytetracycline (97.8%), and tetracycline (93.2%). It was found that all strains were susceptible to norfloxacin, 97.8% to enrofloxacin, and 95.6% to SxT. 24.4% and 15.6% of the strains were resistant to linco-spectin and tiamulin, respectively. The presence oftoxAgene was confirmed by PCR in 20.8% of the strains isolated from nasal swabs and 29.1% of isolates from internal organs.

scholarly journals Ανίχνευση ασβέστωσης των στεφανιαίων αρτηριών με ψηφιακή ακτινοσκόπηση

2016 ◽  
Author(s):  
Μαρία Παπαθανασίου
Keyword(s):  
Weighted Kappa ◽  
Kappa Coefficient ◽  
Weighted Kappa Coefficient

Εισαγωγή:Η μελέτη αυτή διερευνά τη διαγνωστική ακρίβεια της ψηφιακής ακτινοσκόπησης επίπεδων ανιχνευτών (FPDD) για την ανίχνευση ασβέστωσης των στεφανιαίων αρτηριών λαμβάνοντας ως μέθοδο αναφοράς την πολυτομική αξονική τομογραφία καρδιάς (MDCT).Μέθοδος:Μελετήθηκε η ασβέστωση των στεφανιαίων σε ασυμπτωματικούς ενήλικες χαμηλού έως μετρίου καρδιαγγειακού κινδύνου χωρίς γνωστή στεφανιαία νόσο με πολυτομική αξονική τομογραφία και με ψηφιακή ακτινοσκόπηση καρδιάς με την τεχνολογία FPDD. Η ασβέστωση κατηγοριοποιήθηκε με βάση την ακτινοσκοπική εξέταση ημιποσοτικά σε απούσα (0), ήπιου (1), μέτριου (2) και σοβαρού βαθμού (3). Εφαρμόζοντας τη μεθόδο Agatston υπολογίστηκε το σκορ ασβεστίου των στεφανιαίων αρτηριών στην MDCT.Αποτελέσματα:Στη μελέτη συμμετείχαν συνολικά 151 ασυμπτωματικοί εθελοντές, 40-60 ετών (μέση ηλικία 53,1 ± 7,4). Οι άνδρες συνιστούν το 76,5% του πληθυσμού της μελέτης. Η ύπαρξη στεφανιαίας ασβέστωσης ανιχνεύθηκε σε 79 (52,3%) και 69 (45,7%) άτομα με την MDCT και την ακτινοσκόπηση αντίστοιχα. Η συμφωνία ανάμεσα στην MDCT και την ακτινοσκόπηση ήταν 77,5% (weighted kappa coefficient 0,75). Η ακτινοσκόπηση ανίχνευσε ασβέστιο σε 50% των ατόμων με ελάχιστη ασβέστωση στην MDCT (σκορ ασβεστίου <10). Με τη χρήση ROC ανάλυσης και λαμβάνοντας ως ουδό σκορ ασβεστίου μηδέν, η AUC ήταν 0,89 (95% CI 0,83-0,93, p=0,0001) με ευαισθησία 82,3% και ειδικότητα 94,4%. Για σκορ ασβεστίου 10, η AUC ήταν 0,91 (95% CI 0,86-0,95 p=0.0001) με ευαισθησία 91,8% και ειδικότητα 85,6%. Για σκορ ασβεστίου 400, η AUC ήταν 0,97 (95% CI 0,94-0,99, p=0,0001) με ευαισθησία 100% και ειδικότητα 88,7%. Η ενεργός δόση ακτινοβολίας ήταν 1,8 ± 0,09 mSv για την MDCT και 0,26 ± 0,13 mSv για την ακτινοσκόπηση.Συμπεράσματα:Η ψηφιακή ακτινοσκόπηση με την τεχνολογία FPDD έχει εξαιρετική διαγνωστική ακρίβεια σε σύγκριση με την καθιερωμένη MDCT για τη διάγνωση της ασβέστωσης των στεφανιαίων σε ασυμπτωματικά άτομα μέσης ηλικίας, με χαμηλό έως μέτριο καρδιαγγειακό κίνδυνο, με το πλεονέκτημα της μειωμένης έκθεσης σε ακτινοβολία.

scholarly journals Adaptation of an instrument to classify neonatal patients into care categories

2021 ◽  
Vol 55 ◽  
Author(s):  
Ariane Polidoro Dini ◽  
Andrezza de Cassia Vannucci de Oliveira ◽  
Beatriz Pera de Almeida-Hamasaki ◽  
Norma Mejias Quinteiro ◽  
Elenice Valentim Carmona
Keyword(s):  
Intensive Care ◽  
Pediatric Patients ◽  
Weighted Kappa ◽  
Kappa Coefficient ◽  
Patient Classification ◽  
Methodological Study ◽  
Validity Index ◽  
Nursing Team ◽  
Agreement Coefficient ◽  
Weighted Kappa Coefficient

ABSTRACT Objective: To adapt and validate a patient classification instrument for neonatal units. Method: Methodological study, with adaptation of the Pediatric Patients Classification Instrument for neonatal patients. After content validation by judges, the instrument was tested for reliability, applied to 33 neonates by two nurses, simultaneously. To assess the agreement among nurses regarding the scores of each patient on the classification scale, an intraclass agreement coefficient was applied. To assess the agreement regarding the classification in care categories, the weighted Kappa coefficient was calculated. Results: The adapted instrument consisted of a total of nine indicators, with three care categories: high dependence on care, semi-intensive care and intensive care. The Content Validity Index varied between 0.85 and 0.92 for items of the instrument. The intraclass agreement was 0.87 and the weighted Kappa for care categories was 0.56. Conclusion: An instrument that allows neonatal patients to be classified into care categories, with satisfactory reliability was validated to support the dimensioning of the nursing team.

scholarly journals Comparison of Visual and Digital Color Measurement Methods on Anterior Natural Teeth

2018 ◽  
Vol 22 (2) ◽  
pp. 87-92
Author(s):  
Sait Ege Eryürük ◽  
Canan Hekimoğlu ◽  
Elif Tuba Akçin ◽  
Yeliz Çavuşoğlu
Keyword(s):  
Weighted Kappa ◽  
Kappa Coefficient ◽  
Measurement Methods ◽  
Color Measurement ◽  
Central Incisor ◽  
Digital Device ◽  
Natural Teeth ◽  
Digital Methods ◽  
Shade Guide ◽  
Weighted Kappa Coefficient

SummaryBackground/Aim: The purpose of this study was to evaluate compatibility between visual and digital color measurement methods.Material and Methods: The color measurement components of intact natural maxillary right central incisor and left canine teeth of eighty patients were measured with visually shade guide and, digitally, with digital device. The color matchings were performed for each teeth on labial surfaces divided into three regions: cervical, middle and incisal. Ligthness, MLR (M: means middle hue, L designates greener, R designates redder) and chroma levels were assessed for each tooth on divided regions respectively. Measurements were performed by the same and experinced dentist with healthy eyes. The data were analyzed with Kappa and weighted Kappa coefficient (p< 0.05).Results: The compatibility between visual and digital methods did not exist for MLR and chroma. The compatibility between both methods were determined only for ligthness of maxillary central and canine teeth at all regions of labial surfaces.Conclusions: There was compatibility only for ligthness of intact natural teeth at all regions with both methods.

scholarly journals Self-reported weight and height are valid measures to determine weight status: results from the Brazilian National Health Survey (PNS 2013)

2018 ◽  
Vol 34 (5) ◽  
Author(s):  
Naiara Ferraz Moreira ◽  
Verônica Gronau Luz ◽  
Caroline Camila Moreira ◽  
Rosângela Alves Pereira ◽  
Rosely Sichieri ◽  
...  
Keyword(s):  
National Health ◽  
Health Survey ◽  
Weight Status ◽  
Weighted Kappa ◽  
National Health Survey ◽  
Kappa Coefficient ◽  
Bland Altman Method ◽  
Epidemiological Surveys ◽  
Exact Agreement ◽  
Weighted Kappa Coefficient

Self-reported measures have been used to obtain weight and height information in some epidemiological surveys. The validation of such information is necessary to guarantee data quality. This study assessed the validity of self-reported weight and height to determine weight status. Data were obtained in the Brazilian National Health Survey, a Brazilian household-based nationwide survey carried out in 2013. In this survey, 40,366 individuals (aged ≥ 18 years) provided self-reported and measured information about weight and height. Student’s paired t-test was used to verify the differences between self-reported and measured data. The agreement between measurements was obtained using the intraclass correlation coefficient (ICC) and Bland-Altman method. To evaluate variations in weight status categorizations, the weighted kappa coefficient and exact agreement were used. Sensitivity and specificity were estimated for the self-reported information to classify overweight and obese individuals. There was high agreement between self-reported and measured weight, height, and body mass index (ICC > 0.88). The mean agreements estimated by the Bland-Altman method were 99.6% for weight and 100.6% for height. The weighted kappa coefficient showed substantial agreement among the weight status categories (> 0.66); the exact agreement was 77%. Sensibility and specificity for overweight (83% and 87.5%, respectively) and obesity (73.4% and 96.7%, respectively) were considered high for the sociodemographic characteristics evaluated. According to our results, self-reported measurements of weight and height can be used cautiously as valid alternatives to determine weight status.

scholarly journals Molecular Fingerprinting of Pasteurella Multocida Associated with Progressive Atrophic Rhinitis in Swine Herds

1994 ◽  
Vol 6 (4) ◽  
pp. 442-447 ◽  
Author(s):  
Ian A. Gardner ◽  
Rick Kasten ◽  
Graeme J. Eamens ◽  
Kurt P. Snipes ◽  
Randall J. Anderson
Keyword(s):  
Pasteurella Multocida ◽  
Uv Light ◽  
Type A ◽  
Atrophic Rhinitis ◽  
Capsular Type ◽  
Molecular Fingerprinting ◽  
Whole Cell ◽  
Type D ◽  
Progressive Atrophic Rhinitis ◽  
Swine Herds

Ninety-six nasal isolates of Pasteurella multocida from swine herds with progressive atrophic rhinitis were characterized by restriction endonuclease analysis (REA) of whole-cell DNA, ribotyping, and plasmid analysis. For REA, bacterial DNA was digested with SmaI and electrophoresed in 0.7% agarose, and fragments were visualized with UV light. For ribotyping, EcoRI-digested and electrophoresed restriction fragments of whole-cell DNA were transferred to nitrocellulose membranes, hybridized with γ-32P-labeled Escherichia coli ribosomal RNA, and visualized by autoradiography. Phenotypes of isolates were toxigenic capsular type D ( n = 51), nontoxigenic type D ( n = 28), nontoxigenic type A ( n = 16), and toxigenic type A ( n = 1). Plasmids of various sizes were evident in 92.2% and 17.9% of toxigenic and nontoxigenic D strains, respectively, but were absent from all type A strains. Among the 4 phenotypes, there were 17 REA profiles and 6 ribotypes. For 3 of 17 REA patterns, multiple ribotypes were evident, and several REA types were evident in 5 of 6 ribotypes. Thirty-seven isolates of toxigenic capsular type D from Australian herds were either SmaI type B or C and ribotype 2, whereas 14 toxigenic D isolates from the USA and other countries were more heterogeneous (7 REA types and 6 ribotypes). The fingerprinting results provided evidence in support of the hypothesis of a single source infection in Australia associated with the introduction of breeding pigs from overseas.

Development of Two Multiplex Polymerase Chain Reactions for the Detection of Enterotoxigenic Strains of Staphylococcus aureus Isolated from Foods

2003 ◽  
Vol 66 (6) ◽  
pp. 1055-1062 ◽  
Author(s):  
GABRIELA NÁJERA-SÁNCHEZ ◽  
ROGELIO MALDONADO-RODRÍGUEZ ◽  
PATRICIA RUÍZ OLVERA ◽  
LYDIA MOTA de la GARZA
Keyword(s):  
Staphylococcus Aureus ◽  
Simultaneous Detection ◽  
Type A ◽  
Enzyme Linked Immunosorbent Assay ◽  
Powdered Milk ◽  
Chain Reactions ◽  
Polymerase Chain Reactions ◽  
Type D ◽  
Multiplex Reaction ◽  
Polymerase Chain

Two multiplex polymerase chain reactions were developed for the detection of enterotoxigenic strains of Staphylococcus aureus: one multiplex reaction for the simultaneous detection of enterotoxigenic strains type A (entA), type B (entB), and type E (entE) and another for the simultaneous detection of enterotoxigenic strains type C (entC) and type D (entD). Both reactions were standardized with the use of the reference enterotoxigenic strains of S. aureus: FRI 722, producer of staphylococcal enterotoxin (SE) type A (SEA); FRI 1007, producer of SEB; FRI 137, producer of SEC1; FRI 472, producer of SED; and FRI 326, producer of SEE. Optimized methods were used to determine the presence of enterotoxigenic types for 51 S. aureus strains isolated from meat (sausage, ham, and chorizo) and dairy (powdered milk and cheese) products by the Baird-Parker technique. The enterotoxigenic capacities of the strains were determined by the indirect enzyme-linked immunosorbent assay (ELISA) with the use of reference staphylococcal toxins and antitoxins. Fifty of the 51 strains isolated were enterotoxigenic and produced one to four enterotoxin types, with the most frequently produced types being SEA and SED. Levels of correlation between the presence of genes that code for the production of SE (as determined by polymerase chain reaction) and the expression of these genes (as determined by the indirect ELISA) were 100% for SEA and SEE, 86% for SEC, 89% for SED, and 47% for SEB.

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