Nachweis von Actinobacillus equuli ssp. equuli bei Saugferkeln mit eitriger Polyarthritis und Tendovaginitis

2020 ◽  
Vol 48 (01) ◽  
pp. 51-58
Author(s):  
Christiane Maul ◽  
Marcel Suchowski ◽  
Kristin Klose ◽  
Vladislav Antov ◽  
Martin Pfeffer ◽  
...  
Keyword(s):  
16S Rrna ◽  
Post Partum ◽  
Toxin Gene ◽  
Maldi Tof Ms ◽  
Maldi Tof ◽  
Molekularbiologische Methoden ◽  
Tof Ms

Zusammenfassung Actinobacillus equuli ssp. equuli ist ein opportunistischer Erreger des Pferdes, der u. a. als Auslöser der Fohlenfrühlähme bekannt ist. Beim Schwein gibt es vergleichsweise wenige Beschreibungen zu Erkrankungen bedingt durch diese gramnegative Bakterienspezies. Der Bericht beschreibt den Ausbruch einer Infektion in einem kombinierten Zuchtbetrieb mit angeschlossener Mast. Im September 2018 traten bei etwa einem Drittel neugeborener Saugferkel aus Würfen von Jung- und Altsauen folgende Symptome auf: Normal entwickelte Ferkel zeigten nach etwa 6–8 Stunden post partum umfangsvermehrte Gelenke in Zusammenhang mit gering- bis hochgradig ausgeprägten Stütz- und/oder Hangbeinlahmheiten. Die Ferkel wiesen ein reduziertes Allgemeinbefinden auf und befanden sich teilweise in Seitenlage. Innerhalb der ersten Lebenstage kam es zu einer erhöhten Mortalität. Bei 4 der zur Untersuchung eingesandten 7 Saugferkel (2./3. Lebenstag) erfolgte eine pathologisch-anatomische Untersuchung. Hauptbefunde waren eitrige Polyarthritiden und Tendovaginitiden. Bei einem Tier lagen zudem eitrige Entzündungsherde in Gehirn und Nieren vor. Bei der bakteriologischen Untersuchung ließ sich in 18 Lokalisationen (Gehirn, Gelenke, eitrig-abszedierende Veränderungen an Gliedmaßen) A. equuli ssp. equuli nachweisen, zum Teil in Reinkultur. Zur Identifizierung des Erregers wurden kulturell-biochemische Eigenschaften überprüft und ein Massenspektrometrie-Verfahren (MALDI-TOF-MS) angewendet. Als molekularbiologische Methoden dienten sowohl die Sequenzanalyse des 16S-rRNA-Gens als auch eine PCR zum Nachweis ausgewählter apx-Toxin-Gene sowie eine die beiden Subspezies differenzierende PCR. Die Eintragsquelle und die Erregerübertragung im Bestand konnten abschließend nicht aufgeklärt werden. Aus den Isolaten wurde eine bestandsspezifische Vakzine hergestellt und eingesetzt.

scholarly journals Identification and Quantification of Nonviable Lactobacillus pentosus Cells in a Health Food Product

2020 ◽  
Vol 103 (1) ◽  
pp. 223-226
Author(s):  
Na Liu ◽  
Wei Zhou ◽  
Chune Wang ◽  
Xiu Ren ◽  
Haipeng Luo ◽  
...  
Keyword(s):  
16S Rrna ◽  
Metagenomic Analysis ◽  
Lactobacillus Pentosus ◽  
Purity Determination ◽  
Maldi Tof Ms ◽  
Cell Product ◽  
Maldi Tof ◽  
Nonviable Cell ◽  
Tof Ms

Abstract Background: The lack of formal protocol to verify the nonviable cell probiotic product authenticity blocks its registration and supervision process. Objective: To develop a protocol for identification, enumeration, and purity determination of a Lactobacillus pentosus nonviable cell product. Methods: The 16S ribosomal RNA (rRNA) sequencing, 16S rRNA metagenomic analysis, whole-genome sequencing, matrix-assisted laser desorption ionization–time of flight MS (MALDI-TOF-MS), and FACSMicroCount™ system were applied to establish a protocol of identification, enumeration and purity determination of a Lactobacillus pentosus nonviable cell product. Results: The 1530 bp of 16S rRNA sequence could only identify the bacteria at genus level, but the MALDI-TOF-MS could identify both the nonviable cell and fresh culture to species level with high confidence. Metagenomic analysis of the 16S rRNA amplicon could recognize Lactobacillus as the dominant genus in the nonviable cell product. The total number of matching k-mers between the nonviable cell product and the L. pentosus BGM48 in the GenBank was the highest. The 95% confidence interval of the nonviable cell concentration in the product was determined as 4.31–4.68 × 1010 cells/g through the BD FACSMicroCount system. Conclusions: This validation protocol offers an executable approach that can verify microbial contents in nonviable cell products and ensure the compliance with label claims. Highlights: The established validation protocol could determine the nonviable cell species through MALDI-TOF-MS, the concentration through FACSMicroCount system, and the purity and strain level identification through metagenomic analysis of 16S rRNA and the genomic deoxyribonucleic acid.

scholarly journals Acinetobacter gandensis sp. nov. isolated from horse and cattle

2014 ◽  
Vol 64 (Pt_12) ◽  
pp. 4007-4015 ◽  
Author(s):  
Annemieke Smet ◽  
Piet Cools ◽  
Lenka Krizova ◽  
Martina Maixnerova ◽  
Ondrej Sedo ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Type Species ◽  
Rrna Gene ◽  
Content Type ◽  
Dna Relatedness ◽  
Maldi Tof Ms ◽  
Link Type ◽  
Maldi Tof ◽  
Tof Ms

We previously reported the presence of an OXA-23 carbapenemase in an undescribed species of the genus Acinetobacter isolated from horse dung at the Faculty of Veterinary Medicine, Ghent University, Belgium. Here we include six strains to corroborate the delineation of this taxon by phenotypic characterization, DNA–DNA hybridization, 16S rRNA gene and rpoB sequence analysis, % G+C determination, MALDI-TOF MS and fatty acid analysis. The nearly complete 16S rRNA gene sequence of strain UG 60467T showed the highest similarities with those of the type strains of Acinetobacter bouvetii (98.4 %), Acinetobacter haemolyticus (97.7 %), and Acinetobacter schindleri (97.2 %). The partial rpoB sequence of strain UG 60467T showed the highest similarities with ‘Acinetobacter bohemicus’ ANC 3994 (88.6 %), A. bouvetii NIPH 2281 (88.6 %) and A. schindleri CIP 107287T (87.3 %). Whole-cell MALDI-TOF MS analyses supported the distinctness of the group at the protein level. The predominant fatty acids of strain UG 60467T were C12 : 0 3-OH, C12 : 0, C16 : 0, C18 : 1ω9c and summed feature 3 (C16 : 1ω7c and/or iso-C15 : 0 2-OH). Strains UG 60467T and UG 60716 showed a DNA–DNA relatedness of 84 % with each other and a DNA–DNA relatedness with A. schindleri LMG 19576T of 17 % and 20 %, respectively. The DNA G+C content of strain UG 60467T was 39.6 mol%. The name Acinetobacter gandensis sp. nov. is proposed for the novel taxon. The type strain is UG 60467T ( = ANC 4275T = LMG 27960T = DSM 28097T).

scholarly journals Lactococcus garvieae endocarditis in a native valve identified by MALDI-TOF MS and PCR-based 16s rRNA in Spain: A case report

2015 ◽  
Vol 5 ◽  
pp. 13-15 ◽  
Author(s):  
V. Heras Cañas ◽  
M.D. Pérez Ramirez ◽  
F. Bermudez Jiménez ◽  
M.D. Rojo Martin ◽  
C. Miranda Casas ◽  
...  
Keyword(s):  
Case Report ◽  
16S Rrna ◽  
Native Valve ◽  
Maldi Tof Ms ◽  
Lactococcus Garvieae ◽  
Maldi Tof ◽  
Tof Ms

scholarly journals Efficacy of MALDI-TOF mass spectrometry as well as genotypic and phenotypic methods in identification of staphylococci other than Staphylococcus aureus isolated from intramammary infections in dairy cows in Poland

2019 ◽  
Vol 31 (4) ◽  
pp. 523-530 ◽  
Author(s):  
Anna Wanecka ◽  
Jarosław Król ◽  
Jan Twardoń ◽  
Jacek Mrowiec ◽  
Agnieszka Korzeniowska-Kowal ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
16S Rrna ◽  
Species Level ◽  
Confirmatory Test ◽  
Identification System ◽  
Maldi Tof Ms ◽  
Intramammary Infections ◽  
Maldi Tof ◽  
Staphylococcus Spp ◽  
Tof Ms

We compared the effectiveness of various methods for the identification of Staphylococcus spp. other than S. aureus isolated from intramammary infections of cows on 3 dairy farms in Lower Silesia, Poland. A total of 131 isolates belonging to 18 Staphylococcus species were identified by sequence analysis of the 16S rRNA and dnaJ genes, as well using a commercial identification system (ID 32 STAPH; bioMérieux) and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS; Bruker Daltonics). Sequencing of the 16S rRNA gene was found to have low discriminatory value because only 43% of isolates were recognized unequivocally. Much better results were obtained with the dnaJ gene (all isolates were correctly identified at the species level). However, some of these isolates achieved a low similarity level (<97%) and required a confirmatory test (sequencing of the rpoB gene). The performance of ID 32 STAPH was poor. Regardless of the probability level used (80% or 90%), the commercial system obtained identification rates <40%. Using MALDI-TOF MS and the commercial Bruker database, 67% of isolates were identified correctly with scores ≥2.0 (acceptable species-level identification) but this number increased to 97% after the database was expanded. The definitive identification of Staphylococcus spp. other than S. aureus causing intramammary infections in cattle often requires a combination of different procedures, and the existing databases should be updated.

scholarly journals Implementation of a Non-Invasive Bioprospecting Protocol for Isolation of Lactobacillus from Feces of Hens Under Foraging Conditions

2018 ◽  
Vol 14 (28) ◽  
pp. 93-111
Author(s):  
Simón Robledo-Cardona ◽  
Sabina Ramírez-Hincapié ◽  
Javier Correa-Álvarez
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Lactobacillus Species ◽  
Rrna Gene ◽  
Isolation And Identification ◽  
Maldi Tof Ms ◽  
Non Invasive ◽  
Maldi Tof ◽  
Isolation And Characterization ◽  
Tof Ms

In animal production, probiotics seek to replace the use of antibiotics, while diminishing mortality and morbidity rates to raise productivity. Probiotics constitute a natural alternative that, in contrast with antibiotics, neither produces pathogen resistance, nor leaves chemical residues in the final product. Several bacteria, including some belonging to the genus Lactobacillus have been described as probiotics with high potential. A non-invasive bioprospecting protocol aimed for the isolation and characterization of lactobacilli from chicken feces was established. Fecal samples were collected from the ground. These were diluted and cultured in LAB selective medium. Colonies were identified by three methods: Gram stain, MALDI-TOF MS and sequencing of 16S rRNA gene. An initial probiotic potential of lactobacilli isolates was determined via antagonism tests using five enteropathogen reference strains: Staphylococcus aureus, Enterococcus faecium, Candida albicans, Pseudomonas spp. and Salmonella spp. 24 isolates belonging to four Lactobacillus species were identified by MALDITOF MS. BLAST of 16S rRNA gene of eight randomly selected isolates, confirmed MALDI-TOF MS identification. Five of these eight isolates inhibited the growth of at least one of the pathogenic strains used, three isolates of Lactobacillus plantarum and two of Lactobacillus salivarius. Our protocol achieved 21 lactobacilli per 100 isolates performance, greatly surpassing the normal percentage of lactobacilli in chicken gut microbiome, that so, its implementation would facilitate the isolation and identification of new probiotic strains from feces.

scholarly journals MALDI-TOF MS versus 16S rRNA sequencing: Minor discrepancy between tools in identification of Bacteroides isolates

2018 ◽  
Vol 65 (2) ◽  
pp. 173-181
Author(s):  
Károly Péter Sárvári ◽  
József Sóki ◽  
Miklós Iván ◽  
Cecilia Miszti ◽  
Krisztina Latkóczy ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Sequencing ◽  
Maldi Tof Ms ◽  
Maldi Tof ◽  
Rrna Sequencing ◽  
Tof Ms
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